RU2786722C1 - Nitrile-containing antiviral compounds - Google Patents

Abstract

FIELD: pharmaceuticals.

SUBSTANCE: invention relates to (1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, a solvate or hydrate thereof, pharmaceutical compositions containing said compound, and to a method for treating a coronavirus infection such as COVID-19.

EFFECT: creation of new compounds for treating a coronavirus infection.

26 cl, 15 dwg, 41 tbl, 98 ex

Description

State of the art

The invention relates to compounds and methods for inhibiting viral replication activity, comprising contacting a 3C-like ("3CL") proteinase related to SARS-CoV-2 with a therapeutically effective amount of an inhibitor of a 3C-like protease related to SARS-CoV-2. The invention also relates to methods for treating coronavirus disease 2019 ("COVID-19") in a patient by administering a therapeutically effective amount of a SARS-CoV-2 related 3C-like protease inhibitor to a patient in need thereof. The invention also relates to methods for treating COVID-19 in a patient, wherein the method comprises administering to a patient in need thereof a pharmaceutical composition containing a therapeutically effective amount of an inhibitor of a 3C-like protease related to SARS-CoV-2.

The global outbreak of coronavirus disease 2019 (“COVID-19”) has been linked to infections that emerged in late 2019 in Wuhan, Hubei Province, China. By mid-2020, the COVID-19 outbreak had grown into a global pandemic when millions of people were confirmed to be infected, resulting in hundreds of thousands of deaths. The causative agent of COVID-19 has been identified as a novel coronavirus, named SARS coronavirus 2 (“SARS-CoV-2”). The SARS-CoV-2 genome sequence was sequenced from isolates obtained from nine patients in Wuhan, China and found to belong to the Sarbecovirus subgenus of the Betacoronavirus genus. Lu, R. et al. The Lancet, 395, 10224, 565-574; online January 29, 2020. The SARS-CoV-2 sequence was found to have 88% homology with two bat-derived SARS-type coronaviruses, bat-SL-CoVZC45 and bat-SL-CoVZXC21, which were collected in 2018 in Zhoushan, eastern China. SARS-CoV-2 was also found to share approximately 79% homology with the SARS corona virus (“SARS-CoV”), the causative agent of the SARS outbreak in 2002-2003, and approximately 50% homology with the Middle East respiratory syndrome coronavirus (“MERS”). -CoV"), the causative agent of the respiratory virus outbreak that originated in the Middle East in 2012. Based on a recent analysis of 103 sequenced SARS-CoV-2 genomes, it has been suggested that SARS-CoV-2 can be divided into two main types (L and S types), with the S type being inherited and the L type being descended from the S type. Lu, J.; Cui, J. et al. On the origin and continuing evolution of SARS-CoV-2; National Science Review , 7(6), June 2020, 1012-1023, http://doi.org/10.1093/nsr/nwaa036. Types S and L can only be clearly defined by two closely related SNPs at positions 8782 ( orf1ab : T8517C, synonymous) and 28144 ( ORF8 : C251T, S84L). Of the 103 genomes analyzed, approximately 70% belonged to the L type and approximately 30% to the S type. It is not clear whether L type evolved from S type in humans or through a zoonotic intermediate, but it appears that L type is more aggressive than S type, and human intervention in attempts to contain the outbreak may have shifted the relative abundance of L and S types shortly after the outbreak began. SARS-CoV-2. The discovery of the proposed S and L subtypes of SARS-CoV-2 raises the possibility that a person could potentially become infected with separate subtypes sequentially or become infected with both subtypes at the same time. In view of this growing threat, there is an urgent need in the art for effective treatment of COVID-19 and methods for suppressing the replication of the SARS-CoV-2 coronavirus.

Recent evidence clearly shows that the recently emerged coronavirus SARS-CoV-2, the causative agent of COVID-19 (Centers for Disease Control, CDC), has acquired the ability to transmit from person to person, leading to the spread of the virus in the population. The sequence of the receptor-binding domain of the SARS-CoV-2 spike protein (“RBD”), including its receptor-binding motif (RBM), which directly contacts the angiotensin-converting enzyme 2 receptor, ACE2, is similar to the RBD and RBM of SARS-CoV, which strongly suggests that SARS-CoV-2 uses ACE2 as its receptor. Wang, Y.; Shang, J.; Graham, R.; Baric, RS; Li, F.; Receptor recognition by the novel coronavirus from Wuhan: An analysis based on decade-long structural studies of SARS coronavirus; J. Virol. 2020; doi:10.1128/JVI.00127-20. Several critical residues in the SARS-CoV-2 RBM (especially Gln ⁴⁹³ ) provide favorable interactions with human ACE2, consistent with the ability of SARS-CoV-2 to infect human cells. Several other critical residues in the SARS-CoV-2 RBM (notably Asn ⁵⁰¹ ) are compatible but not ideal for human ACE2 binding, suggesting that SARS-CoV-2 uses ACE2 binding to some extent for person-to-person transmission. .

The replication and transcription function of the coronavirus is encoded by the so-called “replicase” gene (Ziebuhr, J., Snijder, E.J., and Gorbalenya, A.E.; Virus-encoded proteinases and proteolytic processing in theNidovirales. J. Gen. Virol. 2000, 81, 853-879; and Fehr, A.R.; Perlman, S.; Coronaviruses: An Overview of Their Replication and Pathogenesis, Methods Mol. Biol. 2015; 1282:1-23. doi:10.1007/978-1-4939-2438-7_1), which consists of two overlapping polyproteins that are extensively processed by viral proteases. The C-proximal region is processed at eleven conserved interdomain junction sites by the major or "3C-like" coronavirus protease (Ziebuhr, Snijder, Gorbalenya, 2000 and Fehr, Perlman et al., 2015). The name "3C-like" protease comes from certain similarities between the coronavirus enzyme and well-known picornavirus 3C proteases. These include substrate preferences, the use of cysteine as the active site nucleophile in catalysis, and the similarity of their putative shared polypeptide folds. The sequence of the SARS-CoV-2 3CL protease (accession number YP_009725301.1) was found to have 96.08% homology compared to the SARS-CoV 3CL protease (accession number YP_009725301.1) Xu, J.; Zhao, S.; Teng, T.; Abdalla, A.E.; Zhu, W.; Xie, L.; Wang, Y.; Guo, X.; Systematic Comparison of Two Animal-to-Human Transmitted Human Coronaviruses: SARS-CoV-2 and SARS-CoV; Viruses 2020, 12, 244; doi:10.3390/v12020244. More recently, Hilgenfeld and colleagues published a high-resolution X-ray structure of the SARS-CoV-2 (3CL) coronavirus core protease Zhang, L.; Lin, D.; Sun, X.; Rox, K.; Hilgenfeld, R.; X-ray Structure of Main Protease of the Novel Coronavirus SARS-CoV-2 Enables Design ofα-Ketoamide Inhibitors; bioRxiv preprint doi: https://doi.org/10.1101/2020.02.17.952879. The structure indicates differences when comparing the 3CL proteases of SARS-CoV-2 and SARS-CoV. In SARS-CoV, but not in the SARS-CoV-2 3CL protease dimer, there is a polar interaction between the two domains III, involving a 2.60-Å hydrogen bond between the side chain hydroxyl groups of the Thr residue²⁸⁵ of each protomer, and is maintained by hydrophobic contact between the Ile side chain²⁸⁶ and Thr²⁸⁵ Cγ2. In SARS-CoV-2 3CL, threonine is replaced by alanine and isoleucine by leucine compared to the same residues in SARS-CoV 3CL. The Thr285Ala substitution observed in the 3CL protease of SARS-CoV-2 allows two domains III to approach each other somewhat (the distance between the C atomsα 285 residues in molecules A and B is 6.77 Å in SARS-CoV 3CL protease and 5.21 Å in SARS-CoV-2 3CL protease, and the distance between the centers of mass of two domains III is reduced from 33.4 Å to 32.1 E). In the active site 3CL of SARS-CoV-2, Cys¹⁴⁵ and His⁴¹ form a catalytic dyad which, when taken together with a hidden water molecule hydrogen-bonded to His⁴¹, can be considered a catalytic triad of SARS-CoV-2 3CL protease. In view of the ongoing spread of SARS-CoV-2, which has caused the current worldwide COVID-19 outbreak, it is desirable to have new ways to suppress SARS-CoV-2 virus replication and treat COVID-19 in patients.

The essence of the invention

The present invention provides novel compounds that act to inhibit or prevent SARS-CoV-2 virus replication and thus are useful in the treatment of COVID-19. The present invention also relates to pharmaceutical compositions containing the compounds and methods for treating COVID-19 and inhibiting the replication of the SARS-CoV-2 virus by administering the compounds of the present invention or pharmaceutical compositions containing the compounds of the present invention. It should be understood that each of the following embodiments of the treatment method can also be formulated as corresponding embodiments of the application. For example, any of the compounds, or their pharmaceutically acceptable salts, or solvates, or hydrates, or pharmaceutically acceptable salts of compounds, solvates, or hydrates, as defined in any of Embodiments E1-E30, E45-E46, E50, E50a, E59-E68 and E80-E83 can be used for use as a drug, or alternatively for use in a method of treatment, as described in any of the embodiments of E36-E41, E47-E49, E52-E58a, E69-E74, E77-R79, E85-E93 and E95-E98.

E1 is a compound according to E45 or E59, below, of formula I

I;

I;

or a pharmaceutically acceptable salt thereof; where R ¹ is selected from the group consisting of C ₁ -C ₆ alkyl, which is optionally substituted with cyano or from one to five fluorine atoms; C ₂ -C ₆ alkynyl; and (C ₃ -C ₆ cycloalkyl)-C ₁ -C ₃ alkyl which is optionally substituted with one to two substituents selected from trifluoromethyl and C ₁ -C ₃ alkyl, or one to five fluorine atoms; R ² is hydrogen, or R ² and R ¹ , taken together with the nitrogen and carbon atoms to which they are attached, are a pyrrolidine or piperidine ring which is optionally substituted with one to four R ^2a ; R ^2a is at each occurrence independently selected from the group consisting of fluorine, C ₁ -C ₆ alkyl optionally substituted with one to three fluorine atoms, and C ₁ -C ₆ alkoxy optionally substituted with one to three fluorine atoms; or two R ^2a groups attached to adjacent carbon atoms and taken together with the atoms to which they are attached are fused C ₃ -C ₆ cycloalkyl which is optionally substituted with one to four R ^2b ; or two R ^2a groups attached to the same carbon atom and taken together with the atoms to which they are attached are spiro C ₃ -C ₆ cycloalkyl which is optionally substituted with one to four R ^2b ; R ^2b is at each occurrence independently selected from fluorine, C ₁ -C ₃ alkyl optionally substituted with one to three fluorine atoms, and C ₁ -C ₃ alkoxy optionally substituted with one to three fluorine atoms; R ³ is selected from the group consisting of C ₁ -C ₈ alkyl, C ₁ -C ₈ alkoxy, (C ₁ -C ₆ alkoxy)-C ₁ -C ₆ alkyl, C ₂ -C ₆ alkynyl, C ₂ -C ₆ alkynyloxy, C ₃ -C ₁₂ cycloalkyl optionally fused with 5-6 membered heteroaryl or phenyl, (C ₃ -C ₁₂ cycloalkyl)-C ₁ -C ₆ alkyl, C ₃ -C ₁₂ cycloalkoxy, (C ₃ -C ₁₂ cycloalkoxy)-C ₁ -C ₆ alkyl, 4-12 membered heterocycloalkyl which is optionally fused with 5-6 membered heteroaryl or phenyl, and wherein said heterocycloalkyl contains one to four heteroatoms independently selected from N, O and S( O) _n , (4-12 membered heterocycloalkyl)-C ₁ -C ₆ alkyl, wherein said heterocycloalkyl group contains one to four heteroatoms independently selected from N, O and S(O) _n , C ₆ -C ₁₀ aryl , optionally fused with C ₄ -C ₆ cycloalkyl or 4-7 membered heterocycloalkyl, (C ₆ -C ₁₀ aryl)-C ₁ -C ₆ alkyl, 5-10 membered heteroaryl containing from one to five heteroatoms, independently selected from N, O and S, which is optionally fused with C ₅ -C ₆ cycloalkyl; (5-10-membered heteroaryl)-C ₁ -C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; (C ₆ -C ₁₀ aryl)-(5-10 membered heteroaryl)- wherein the heteroaryl group contains one to five heteroatoms independently selected from N, O and S, (5-10 membered heteroaryloxy)-C ₁ -C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; (5-6 membered heteroaryl)-(5-6 membered heteroaryl)-, where each heteroaryl group contains from one to four heteroatoms independently selected from N, O and S; (4-7-membered heterocycloalkyl)-(5-6-membered heteroaryl)-, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O) _n and the heteroaryl group contains one to four heteroatoms , independently selected from N, O and S; (5-6-membered heteroaryl)-(4-7-membered heterocycloalkyl)-, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O) _n and the heteroaryl group contains one to four heteroatoms , independently selected from N, O and S; where each R ³ group is optionally substituted with one to five R ⁴ ; R ⁴ at each occurrence is independently selected from the group consisting of oxo, halogen, hydroxy, cyano, phenyl, benzyl, amino, (C ₁ -C ₆ alkyl)amino optionally substituted with one to five fluorine atoms, di(C ₁ - C ₆ alkyl)amino optionally substituted with one to ten fluorine atoms, C ₁ -C ₆ alkyl optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkoxy optionally substituted with one to five fluorine atoms, C ₁ - C ₃ alkoxy-C ₁ -C ₃ alkyl optionally substituted with one to five fluorine atoms, C ₃ -C ₆ cycloalkyl optionally substituted with one to three fluorine atoms or C ₁ -C ₃ alkyl, C ₁ -C ₆ alkyl- C(O)NH- optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkyl-S(O) ₂ NH- optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkyl-C(O )- optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkyl-S(O) _n - optionally substituted with one to five fluorine atoms; and

n is independently selected from 0, 1 and 2 in each case.

E2 is a compound according to any of E1, E45 and E59, where R ¹ is selected from the group consisting of (CH ₃ ) ₂ CHCH ₂ -, (CH ₃ ) ₃ CCH ₂ -, cyanomethyl, 2-cyanoethyl, 2,2-difluoroethyl , 2,2,2-trifluoroethyl, 3,3-difluoropropyl, 3,3,3-trifluoropropyl, 3,3,3-trifluoro-2-methylpropyl, cyclopropylmethyl, (2,2-difluorocyclopropyl)methyl, [1-( trifluoromethyl)cyclopropyl]methyl, (2-methylcyclopropyl)methyl, (3,3-difluorocyclobutyl)methyl, cyclopentylmethyl and propynyl; and R ² is hydrogen; or a pharmaceutically acceptable salt thereof.

E3 is a compound according to any of E1, E45 and E59, where R ² and R ¹ taken together with the nitrogen and carbon atoms to which they are attached, are a pyrrolidine or piperidine ring which is optionally substituted with one to four R ^2a ; or a pharmaceutically acceptable salt thereof.

E4 is a compound according to any one of E1, E45, E59 and E3 wherein R ^2a is at each occurrence independently selected from the group consisting of fluoro, methyl, isopropyl, trifluoromethyl and t -butoxy; or two R ^2a groups attached to adjacent carbon atoms and taken together with the atoms to which they are attached are fused cyclopentane or cyclopropane which is optionally substituted with one to four R ^2b ; or two R ^2a groups attached to the same carbon atom and taken together with the atoms to which they are attached is a spirocyclopropane ring which is optionally substituted with one to four R ^2b ; or a pharmaceutically acceptable salt thereof.

E5 is a compound according to E1, E3, E4, E45 and E59, where R ^2b in each case is independently selected from the group consisting of fluorine, methyl and methoxy; or a pharmaceutically acceptable salt thereof.

E6 is a compound according to any one of E1, E2, E45 and E59 selected from the group consisting of formulas Ia - Ig

;and

;

or a pharmaceutically acceptable salt thereof.

E7 is a compound according to any of E1, E3, E4, E45 and E59 selected from the group consisting of formulas Ih - Ik

or a pharmaceutically acceptable salt thereof.

E8 is a compound according to any one of E1, E3, E4, E7, E45 and E59 selected from the group consisting of

or a pharmaceutically acceptable salt thereof.

E9 is a compound according to any of E1, E3, E4, E7, E8, E45 and E59 where R ³ is selected from the group consisting of C ₁ -C ₆ alkyl and (C ₃ -C ₆ cycloalkyl)-C ₁ -C ₃ alkyl ; each of which is substituted with one to four R ⁴ ; or a pharmaceutically acceptable salt thereof.

E10 is a compound according to any of E1, E3, E4, E7 to E9, E45 and E59 where R ³ is selected from the group consisting of (CH ₃ ) ₂ CHCH(R ⁴ )-, (CH ₃ ) ₃ CCH(R ⁴ ) - and (cyclohexyl)CH(R ⁴ )-; or a pharmaceutically acceptable salt thereof.

E11 is a compound according to any one of E1, E3, E4, E7-E10, E45 and E59 selected from the group consisting of

,

,

or a pharmaceutically acceptable salt thereof.

E12 is a compound according to any one of E1, E3, E4, E7-E11, E45 and E59, where R ⁴ is selected from the group consisting of (C ₁ -C ₆ alkyl)amino optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkyl-C(O)NH- optionally substituted with one to five fluorine atoms, and C ₁ -C ₆ alkyl-S(O) ₂ NH- optionally substituted with one to five fluorine atoms; or a pharmaceutically acceptable salt thereof.

E13 is a compound according to any of E1, E3, E4, E7-E12, E45 and E59, where R ⁴ is selected from the group consisting of CF ₃ C(O)NH-, CF ₃ S(O) ₂ NH-, CH ₃ C(O)NH-, CH ₃ CH ₂ C(O)NH- and CF ₃ CH ₂ NH-; or a pharmaceutically acceptable salt thereof.

E14 is a compound according to any of E1, E3, E4, E7-E13, E45 and E59, where R ⁴ is CF ₃ C(O)NH- or CF ₃ S(O) ₂ NH-; or a pharmaceutically acceptable salt thereof.

E15 is a compound according to any one of E1-E8, E45 and E59, where R ³ is a 4-12 membered heterocycloalkyl which is optionally fused to a 5-6 membered heteroaryl or phenyl, and wherein said heterocycloalkyl contains from one to four heteroatoms, independently selected from N, O and S(O) _n , or is (4-12-membered heterocycloalkyl)-C ₁ -C ₆ alkyl, where the specified heterocycloalkyl group contains from one to four heteroatoms independently selected from N, O and S( O) _n ; each of which is optionally substituted with one to five R ⁴ ; or a pharmaceutically acceptable salt thereof.

E16 is a compound according to any one of E1-E8, E15, E45 and E59, wherein the 4-12 membered heterocycloalkyl group in ^R3 is selected from the group consisting of acetidinyl, pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl, oxetanyl, tetrahydrofuranyl, pyranyl, 2 -oxo-1,3-oxazolidinyl, oxabicyclo[2.2.1]heptyl, 1-oxa-8-azaspiro[4.5]decyl, 1,1-dioxido-1,2-thiazolidinyl and 1,1-dioxido-1,2 -thiazinanil; each of which is optionally substituted with one to three R ⁴ ; or a pharmaceutically acceptable salt thereof.

E17 is a compound according to any one of E1-E8, E45 and E59, where R ³ is selected from the group consisting of phenyl, benzyl, phenethyl, 5-10 membered heteroaryl containing from one to five heteroatoms independently selected from N, O and S; (5-10-membered heteroaryl)-C ₁ -C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; and (5-10-membered heteroaryloxy)-C ₁ -C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; each of which is optionally substituted with one to five R ⁴ ; or a pharmaceutically acceptable salt thereof.

E18 is a compound according to any one of E1-E8, E17, E45 and E59, where the 5-10 membered heteroaryl group in R³ selected from the group consisting of imidazolyl, pyrazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, triazolyl, pyridinyl, pyrimidinyl, pyrazinyl, pyridazinyl, indolyl, benzimidazolyl, pyridinopyrrolyl, quinolinyl, quinoxalinyl, benzotriazolyl, imidazo[1,2-a] pyridinyl, imidazo[2,1-b][1,3]thiazolyl, 4H-furo[3,2-b]pyrrolyl, 4H-thieno[3,2-b]pyrrolyl, [1,2,4]triazolo[ 1,5-a]pyrimidinyl, [1,2,3]triazolo[1,5-a]pyridinyl and naphthyridinyl; each of which is optionally substituted with one to four R⁴; or a pharmaceutically acceptable salt thereof.

E19 is a compound according to any of E1-E8, E17-E18, E45 and E59, where R ³ is indolyl; which is optionally substituted with one to four R ⁴ ; or a pharmaceutically acceptable salt thereof.

E20 is a compound according to any of E1-E8, E17-E19, E45 and E59, where R ³ is indol-2-yl; which is optionally substituted with one to four R ⁴ ; and R ⁴ at each occurrence is independently selected from the group consisting of fluorine, chlorine, bromine, hydroxy, methyl, ethyl, propyl, isopropyl, 1-methylpropyl, butyl, t -butyl, acetyl, methoxy, ethoxy, propoxy, butoxy, trifluoromethyl , trifluoromethoxy, cyclohexyl and diethylamino; or a pharmaceutically acceptable salt thereof.

E21 is a compound according to any one of E1, E2, E6, E9-E10, E12-E20, E45 and E59 of the formula

;

;

or a pharmaceutically acceptable salt thereof.

E22 is a compound according to any of E1, E2, E6, E9-E10, E12-E21, E45 and E59, where R ³ is selected from the group consisting of 1H-indol-2-yl, 7-fluoro-4-methoxy-1H -indol-2-yl, 4-methoxy-7-(trifluoromethyl)-1H-indol-2-yl, 4-methoxy-1H-indol-2-yl, 4-(trifluoromethoxy)-1H-indol-2-yl , 6-(trifluoromethyl)-1H-indol-2-yl, 4-methoxy-3,6,7-tris(trifluoromethyl)-1H-indol-2-yl, 3-fluoro-4-methoxy-1H-indol- 2-yl and 3,5-difluoro-4-methoxy-1H-indol-2-yl; or a pharmaceutically acceptable salt thereof.

E23 is a compound according to any of E1-E8, E21, E45 and E59, where R ³ is C ₁ -C ₆ alkoxy; or a pharmaceutically acceptable salt thereof.

E24 is a compound according to any one of E1-E8, E21, E23, E45 and E59, where R ³ is selected from the group consisting of methoxy, ethoxy and prop-2-oxy; or a pharmaceutically acceptable salt thereof.

E25 is a compound according to any one of E1-E8, E21, E45 and E59, where R ³ is selected from the group consisting of C ₃ -C ₁₂ cycloalkyl optionally fused with 5-6 membered heteroaryl or phenyl, (C ₃ -C ₁₂ cycloalkyl)-C ₁ -C ₆ alkyl, C ₃ -C ₁₂ cycloalkoxy and (C ₃ -C ₁₂ cycloalkoxy)-C ₁ -C ₆ alkyl; each of which is optionally substituted with one to three R ⁴ ; or a pharmaceutically acceptable salt thereof.

E26 is a compound according to any one of E1-E8, E21, E25, E45 and E59, where R ³ is selected from the group consisting of cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1-(cyclohexyloxy)ethyl, cyclohexoxymethyl, cyclopropylmethyl, cyclopropylethyl, cyclobutylmethyl, cyclobutylethyl, cyclopentylmethyl, cyclopentylethyl, cyclohexylmethyl and cyclohexylethyl; each of which is optionally substituted with one to three R ⁴ ; or a pharmaceutically acceptable salt thereof.

E27 is a compound according to any one of E1-E8, E17, E45 and E59, where R ³ is selected from the group consisting of phenyl, benzyl and phenethyl, each of which is optionally substituted with one to three R ⁴ ; or a pharmaceutically acceptable salt thereof.

E28 is a compound according to any of E1-E8, E17, E27, E45 and E59, where R ⁴ is selected from the group consisting of fluorine, chlorine, dimethylamino, trifluoromethyl, CF ₃ C(O)NH- and CF ₃ S(O) _2NH- ; or a pharmaceutically acceptable salt thereof.

E29 is a compound according to any one of E1, E45 and E59 selected from the group consisting of

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -{(2R)-2-(dimethylamino)-2-[4-(trifluoromethyl )phenyl]acetyl}-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -{(2R)-2-(dimethylamino)-2-[3-(trifluoromethyl )phenyl]acetyl}-4-methyl-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-6-(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-3,6,7-tris(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-(trifluoromethoxy)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-(trifluoromethoxy)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-3-fluoro-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-3,5-difluoro-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5,7-difluoro-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-fluoro-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-3,5,7-tris(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-3,7-bis(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-(trifluoromethyl)-1H-indole-2-carboxamide;

7-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methoxy-7-methyl-1H-indole-2-carboxamide;

6-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

4-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-7-(trifluoromethyl)-1H-indole-2-carboxamide;

4,6-dichloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl -1-oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-(trifluoromethyl)-1H-indole-2-carboxamide;

7-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-7-methyl-1H-indole-2-carboxamide;

6-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

4-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

5,7-dichloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-7-(trifluoromethyl)-1H-indole-2-carboxamide;

4,6-dichloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-3-methyl-5-(trifluoromethyl)imidazo[2,1-b][1,3]thiazole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[4-methyl-2-(trifluoromethyl)-1, 3-thiazol-5-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[5-methyl-2-(trifluoromethyl)-1, 3-thiazol-4-yl]carbonyl}-L-leucinamide;

N ² -[(4-bromo-1-ethyl-3-methyl-1H-pyrazol-5-yl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine- 3-yl]ethyl}-L-leucinamide;

N ² -[(4-chloro-1,3-dimethyl-1H-pyrazol-5-yl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine-3- yl]ethyl}-L-leucinamide;

3-acetyl-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3R)-2,5-dioxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2 -yl]-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-hydroxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-hydroxy-4-methoxy-1H-indole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(3,3-difluorocyclobutyl)acetyl]-4-methyl-L-leucinamide ;

N ² -[(trans-4-cyanocyclohexyl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide ;

N ² -[(trans-4-cyanocyclohexyl)carbonyl]-N-{(1R)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide ;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[2-(cyclohexyloxy)propanoyl]-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(2S)-2-(dimethylamino)-2-phenylacetyl]-4- methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -(pyrrolidin-1-ylacetyl)-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(2R)-2-(dimethylamino)-2-phenylacetyl]-4- methyl-L-leucinamide;

N ² -[(4-chloro-1,3-dimethyl-1H-pyrazol-5-yl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine-3- yl]ethyl}-4-methyl-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methoxy-3-(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methoxy-7-(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methoxy-3,7-bis(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methoxy-3,5-bis(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methoxy-3,6-bis(trifluoromethyl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-3-(trifluoromethyl)-1H-indole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -(cyclohexylcarbonyl)-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -(cyclohexylcarbonyl)-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[2-(trifluoromethyl)-1,3-thiazol- 4-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -[(propan-2-yloxy)acetyl]-L-leucinamide ;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(cyclohexyloxy)acetyl]-4-methyl-L-leucinamide;

N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -(4,4,4-trifluoro-3-methylbutanoyl )-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-3-methylimidazo[2,1-b][1,3]thiazole-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-5,5,5-trifluoro-1-oxopentane -2-yl]-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-7-fluoro-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methoxy-1H-indole-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(trifluoroacetyl )-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

N ² -[(4-bromo-1-ethyl-3-methyl-1H-pyrazol-5-yl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine- 3-yl]ethyl}-4-methyl-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-7-(trifluoromethyl)-1H-indole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -(2,6-dichlorobenzoyl)-4-methyl-L-leucinamide;

(2S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4,4-dimethyl-1-[3-methyl-N-(trifluoroacetyl )-L-valyl]piperidine-2-carboxamide;

3-methyl-N-(trifluoroacetyl)-L-valyl-(4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl -4-(trifluoromethyl)-L-prolinamide;

(2S,4S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-{3-methyl-N-[( trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -{[2-(trifluoromethyl)-1,3-thiazol-5-yl] carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(2S)-2-(dimethylamino)-2-phenylacetyl]-4- methyl-L-leucinamide;

N ² -[(trans-4-cyanocyclohexyl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide ;

N ² -[(trans-4-cyanocyclohexyl)carbonyl]-N-{(1R)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide ;

N-{(1R)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[2-(cyclohexyloxy)propanoyl]-4-methyl-L-leucinamide;

(2S,4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[N-(trifluoroacetyl)-L- valyl]piperidine-2-carboxamide;

5-(butan-2-yl)-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4 ,4-dimethyl-1-oxopentan-2-yl]-1H-indole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(4,5-dichloro-1H-imidazol-2-yl)carbonyl] -4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(4,5-dichloro-1H-pyrazol-3-yl)carbonyl] -4-methyl-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-2,3-dimethyl-4H-furo[3,2-b]pyrrole-5-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-pyrrolo[2,3-b]pyridine-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-(trifluoromethyl)-1H-benzimidazole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-methoxy-1H-pyrrolo[3,2-b]pyridine-2-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-pyrrolo[3,2-b]pyridine-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-fluoro-1H-benzimidazole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[3-(propan-2-yl)-1H- pyrazol-5-yl]carbonyl}-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-fluoro-1H-benzimidazole-2-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-benzimidazole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5,6-difluoro-1H-benzimidazole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4H-thieno[3,2-b]pyrrole-5-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[3-(2-methylpropyl)-1H-pyrazole- 5-yl]carbonyl}-L-leucinamide;

N ² -{[4-(3-chlorophenyl)-1H-imidazol-2-yl]carbonyl}-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl] ethyl}-4-methyl-L-leucinamide;

N ² -[(3-tert-butyl-1H-pyrazol-5-yl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl} -4-methyl-L-leucinamide;

6-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-benzimidazole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-methyl-1H-benzimidazole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4,5,6,7-tetrahydro-1H-indazole-3-carboxamide;

4,6-dichloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl -1-oxopentan-2-yl]-1H-benzimidazole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-(1-methylcyclopropyl)-4-(trifluoromethyl)-1H-pyrrolo[2,3-b]pyridine-2-carboxamide;

N ² -{[5-(2-chlorophenyl)-4-fluoro-1H-pyrazol-3-yl]carbonyl}-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine- 3-yl]ethyl}-4-methyl-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-2-methyl-4H-thieno[3,2-b]pyrrole-5-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -{[3-(4-methoxyphenyl)-1H-pyrazol-5-yl] carbonyl}-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -{[3-(2-methoxyphenyl)-1H-pyrazol-5-yl] carbonyl}-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -{[4-(4-methoxyphenyl)-1H-imidazol-2-yl] carbonyl}-4-methyl-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[3-(4-methylphenyl)-1H-pyrazole- 5-yl]carbonyl}-L-leucinamide;

7-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-5-methyl-1H-indole-2-carboxamide;

7-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

(2S,4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[3-methyl-N-(methylsulfonyl )-L-valyl]piperidine-2-carboxamide;

(2S,4S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[N-(trifluoroacetyl)-L- valyl]piperidine-2-carboxamide;

(2S,4S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[3-methyl-N-(trifluoroacetyl )-L-valyl]piperidine-2-carboxamide;

(2S,4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[3-methyl-N-(trifluoroacetyl )-L-valyl]piperidine-2-carboxamide;

5-[(2S)-butan-2-yl]-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl} amino)-4,4-dimethyl-1-oxopentan-2-yl]-1H-indole-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3',3 ',3'-trifluoro-N-(trifluoroacetyl)-L-isoleucyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-{(2S)-2-cyclohexyl-2- [(trifluoroacetyl)amino]acetyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-{(2S)-2-cyclopentyl-2- [(trifluoroacetyl)amino]acetyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[4-methyl- N-(trifluoroacetyl)-L-leucyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-{(2S)-2-(4.4 -difluorocyclohexyl)-2-[(trifluoroacetyl)amino]acetyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclopentyl-N-(trifluoroacetyl)- L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclohexyl-N-(trifluoroacetyl)- L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(trifluoroacetyl )-L-leucyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-[6,6-difluoro-N-(trifluoroacetyl )-L-norleucyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-{(2S)- 4,4,4-trifluoro-2-[(trifluoroacetyl)amino]butanoyl}-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-[3-fluoro-N-(trifluoroacetyl)- L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-{(2S)-2-cyclopropyl-2- [(trifluoroacetyl)amino]acetyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-[3-(3,3-difluorocyclobutyl)- N-(trifluoroacetyl)-L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(trifluoroacetyl )-O-(trifluoromethyl)-L-seryl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-{(2S)- 2-phenyl-2-[(trifluoroacetyl)amino]acetyl}-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(trifluoroacetyl )-L-phenylalanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-[3,5-difluoro-N-(trifluoroacetyl )-L-phenylalanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(trifluoroacetyl )-3-(trifluoromethyl)-L-phenylalanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(2 ,2,2-trifluoroethyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(2S,4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-{(2S)-3-methyl- 2-[(trifluoroacetyl)amino]butyl}piperidine-2-carboxamide;

(2S,4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-{(2S)-3-methyl- 2-[(2,2,2-trifluoroethyl)amino]butyl}piperidine-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(3 ,3,3-trifluoropropanoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-(N-propanoyl- L-valyl)-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(2S,4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[N-(2,2,2 -trifluoroethyl)-L-valyl]piperidine-2-carboxamide;

N ² -[(4-chloro-1-ethyl-3-methyl-1H-pyrazol-5-yl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine- 3-yl]ethyl}-L-leucinamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-3-ethyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-cyclohexyl-1H-indole-2-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-3-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-3,5-dimethyl-1H-indole-2-carboxamide;

5-tert-Butyl-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-(propan-2-yl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-3-ethyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-6-ethyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-ethyl-1H-indole-2-carboxamide;

4-butoxy-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-(trifluoromethoxy)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-6-(diethylamino)-1H-indole-2-carboxamide;

4-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

5-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

6-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-3-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-6-propoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-7-fluoro-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-7-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-6-fluoro-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-fluoro-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4,5-dimethoxy-1H-indole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -[(4-methyl-1,3-thiazol-5- yl)carbonyl]-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -(ethoxycarbonyl)-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -(ethoxycarbonyl)-4-methyl-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-5,5,5-trifluoro-1-oxopentane -2-yl]-4-methoxy-1H-indole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[2-(trifluoromethyl)-1,3-oxazol- 4-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[3-(trifluoromethyl)-1,2-thiazol- 4-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[3-(trifluoromethyl)-1,2-oxazol- 4-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -{[2-(trifluoromethyl)-1,3-thiazol-4-yl] carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-5,5,5-trifluoro-N ² -{[2-(trifluoromethyl)-1, 3-thiazol-4-yl]carbonyl}-L-norvalinamide;

(4S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-5,5,5-trifluoro-N ² -{[2-(trifluoromethyl )-1,3-thiazol-4-yl]carbonyl}-L-leucinamide;

(4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-5,5,5-trifluoro-N ² -{[2-(trifluoromethyl )-1,3-thiazol-4-yl]carbonyl}-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-3-cyclopentyl-1-oxopropan-2-yl ]-2-(trifluoromethyl)-1,3-thiazole-4-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[5-(trifluoromethyl)-1,2-thiazol- 4-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[5-(trifluoromethyl)-1,2-oxazol- 4-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[2-(trifluoromethyl)-1,3-oxazol- 5-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[2-(trifluoromethyl)-1,3-thiazol- 5-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-5,5,5-trifluoro-4-methyl-N ² -{[2-(trifluoromethyl )-1,3-thiazol-4-yl]carbonyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-N ² -{[(2S)-2-methyltetrahydrofuran-2-yl] carbonyl}-L-leucinamide;

N-[(2S,4R)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-5,5,5-trifluoro-4 -methyl-1-oxopentan-2-yl]-4-methoxy-1H-indole-2-carboxamide;

N-[(2S,4S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-5,5,5-trifluoro-4 -methyl-1-oxopentan-2-yl]-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-5,5,5-trifluoro-4,4 -dimethyl-1-oxopentan-2-yl]-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-3-cyclopentyl-1-oxopropan-2-yl ]-4-methoxy-1H-indole-2-carboxamide;

5,7-dichloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl -1-oxopentan-2-yl]-1H-indole-2-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-3-ethyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-cyclohexyl-1H-indole-2-carboxamide;

5-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-3-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-3,5-dimethyl-1H-indole-2-carboxamide;

5-tert-butyl-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl -1-oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-(propan-2-yl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-7-(propan-2-yl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-3-ethyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-ethyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-ethyl-1H-indole-2-carboxamide;

4-butoxy-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-(trifluoromethoxy)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-(diethylamino)-1H-indole-2-carboxamide;

4-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

5-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

6-bromo-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1 -oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-3-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-propoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-7-fluoro-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-7-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-fluoro-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-fluoro-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-6-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-5-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-4,5-dimethoxy-1H-indole-2-carboxamide;

5-(butan-2-yl)-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4 -methyl-1-oxopentan-2-yl]-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-7-(propan-2-yl)-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-5-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-6-methyl-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-6-methoxy-1H-indole-2-carboxamide;

5-(butan-2-yl)-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4 ,4-dimethyl-1-oxopentan-2-yl]-1H-indole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(2R)-2-cyclohexyl-2-methoxyacetyl]-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(2R)-2-(cyclohexyloxy)propanoyl]-L-leucinamide;

N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -(4,4,4-trifluoro-3-methylbutanoyl )-L-leucinamide;

N ² -[(trans-4-cyanocyclohexyl)carbonyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(1-ethyl-4-methyl-1H-pyrazol-5-yl) carbonyl]-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -(cyclohexylcarbonyl)-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(cyclohexyloxy)acetyl]-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(3,3-difluorocyclobutyl)acetyl]-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(propan-2-yloxy)acetyl]-L-leucinamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-3-methylimidazo[2,1-b][1,3]thiazole-2-carboxamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(2R)-2-cyclohexyl-2-methoxyacetyl]-4-methyl- L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -[(1-ethyl-4-methyl-1H-pyrazol-5-yl) carbonyl]-4-methyl-L-leucinamide;

N ² -[2-chloro-4-(methylsulfonyl)benzoyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-L-leucinamide;

N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ² -(2,6-dichlorobenzoyl)-L-leucinamide;

(1R,2S,5S)-3-[N-(tert-butylsulfonyl)-3-methyl-L-valyl]-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidine- 3-yl]ethyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-3-{[(3R)-1-benzyl-5-oxopyrrolidin-3-yl]carbonyl}-N-{(1S)-1-cyano-2-[(3S)-2 -oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-{[(3R) -5-oxo-1-phenylpyrrolidin-3-yl]carbonyl}-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-3-{[(3R)-1-tert-butyl-5-oxopyrrolidin-3-yl]carbonyl}-N-{(1S)-1-cyano-2-[(3S) -2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[(3-methylimidazo [2,1-b][1,3]thiazol-2-yl)carbonyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-{[2-( trifluoromethyl)-1,3-thiazol-4-yl]carbonyl}-3-azabicyclo[3.1.0]hexane-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-3-cyclopropyl-1-oxopropan-2-yl ]-4-methoxy-1H-indole-2-carboxamide; and

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-3-cyclopropyl-1-oxopropan-2-yl ]-1H-indole-2-carboxamide;

or a pharmaceutically acceptable salt thereof.

E30 is a compound according to any one of E1, E45 and E59 selected from the group consisting of

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2 -yl]-7-fluoro-4-methoxy-1H-indole-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-4-methoxy-7-(trifluoromethyl)-1H-indole-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[N-(trifluoroacetyl )-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide;

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl ]-7-fluoro-4-methoxy-1H-indole-2-carboxamide;

(2S,4S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[N-(trifluoroacetyl)-L- valyl]piperidine-2-carboxamide;

(2S,4S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-[3-methyl-N-(trifluoroacetyl )-L-valyl]piperidine-2-carboxamide;

(1R,2S,5S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-{(2S)-2-cyclohexyl-2- [(trifluoroacetyl)amino]acetyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide;

(2S,4S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-1-{3-methyl-N-[( trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide;

3-methyl-N-(trifluoroacetyl)-L-valyl-(4R)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4-methyl -4-(trifluoromethyl)-L-prolinamide; and

(2S)-N-{(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-4,4-dimethyl-1-[3-methyl-N-(trifluoroacetyl )-L-valyl]piperidine-2-carboxamide;

or a pharmaceutically acceptable salt thereof.

E31 is a pharmaceutical composition containing a pharmaceutically effective amount of a compound according to any of E1-E30, or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier.

E32 is a pharmaceutical composition according to E31, where the composition is in intravenous, subcutaneous, inhalable or oral dosage form

E33 is a pharmaceutical composition according to E31 or E32, where the composition is in an oral dosage form.

E34 is a pharmaceutical composition according to any of E31-E33, additionally containing an additional therapeutic agent.

E35 is a pharmaceutical composition according to any one of E31-E34, wherein the pharmaceutical composition further comprises one or more of dexamethasone, azithromycin, and remdesivir.

E36 is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of a compound of any of E1-E30, or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

E37 is the way of E36 where the coronavirus infection is COVID-19.

E38 is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a pharmaceutical composition according to any of E31-E35 to a patient in need thereof.

E39 is the way of E38 where the coronavirus infection is COVID-19.

E40 is a method for inhibiting or preventing SARS-CoV-2 virus replication, comprising contacting SARS-CoV-2 coronavirus 3CL protease with a therapeutically effective amount of a compound or pharmaceutically acceptable salt thereof according to any one of E1-E30.

E41 is a method for inhibiting or preventing SARS-CoV-2 virus replication in a patient, comprising administering to a patient in need of inhibiting or preventing SARS-CoV-2 virus replication a therapeutically effective amount of a compound or a pharmaceutically acceptable salt thereof according to any one of E1-E30.

E42 is the use of a compound or a pharmaceutically acceptable salt thereof according to any one of E1-E30 for the treatment of a coronavirus infection.

E43 is an application of E42 where the coronavirus infection is COVID-19.

E44 is the use of a compound or a pharmaceutically acceptable salt thereof according to any of E1-E30 for the preparation of a medicament that is used to treat a coronavirus infection.

E44a is the application of E44 where the coronavirus infection is COVID-19.

E45 is a compound of formula I '

or a pharmaceutically acceptable salt thereof; where R is independently at each occurrence hydroxy or oxo; p is 0, 1, or 2; R ¹ is selected from the group consisting of C ₁ -C ₆ alkyl which is optionally substituted with cyano or from one to five fluorine atoms; C ₂ -C ₆ alkynyl; and (C ₃ -C ₆ cycloalkyl)-C ₁ -C ₃ alkyl which is optionally substituted with one to two substituents selected from trifluoromethyl and C ₁ -C ₃ alkyl or one to five fluorine atoms; R ² is hydrogen, or R ² and R ¹ taken together with the nitrogen and carbon atoms to which they are attached, are a pyrrolidine or piperidine ring which is optionally substituted with one to four R ^2a ; R ^2a is at each occurrence independently selected from the group consisting of fluorine, hydroxy, C ₁ -C ₆ alkyl optionally substituted with one to three fluorine atoms, and C ₁ -C ₆ alkoxy optionally substituted with one to three fluorine atoms; or two R ^2a groups attached to adjacent carbon atoms and taken together with the atoms to which they are attached are fused C ₃ -C ₆ cycloalkyl which is optionally substituted with one to four R ^2b ; or two R ^2a groups attached to the same carbon atom and taken together with the atoms to which they are attached are spiro C ₃ -C ₆ cycloalkyl which is optionally substituted with one to four R ^2b ; R ^2b is at each occurrence independently selected from fluorine, hydroxy, C ₁ -C ₃ alkyl optionally independently substituted with one to three fluorine or hydroxy, and C ₁ -C ₃ alkoxy optionally independently substituted with one to three fluorine or hydroxy; R ³ is selected from the group consisting of C ₁ -C ₈ alkyl, C ₁ -C ₈ alkoxy, (C ₁ -C ₆ alkoxy)-C ₁ -C ₆ alkyl, C ₂ -C ₆ alkynyl, C ₂ -C ₆ alkynyloxy, C ₃ -C ₁₂ cycloalkyl optionally fused with 5-6 membered heteroaryl or phenyl, (C ₃ -C ₁₂ cycloalkyl)-C ₁ -C ₆ alkyl, C ₃ -C ₁₂ cycloalkoxy, (C ₃ -C ₁₂ cycloalkoxy)-C ₁ -C ₆ alkyl, 4-12 membered heterocycloalkyl which is optionally fused with 5-6 membered heteroaryl or phenyl and wherein said heterocycloalkyl contains one to four heteroatoms independently selected from N, O and S(O ) _n , (4-12-membered heterocycloalkyl)-C ₁ -C ₆ alkyl, where the specified heterocycloalkyl group contains from one to four heteroatoms independently selected from N, O and S(O) _n , C ₆ -C ₁₀ aryl, optionally fused with C ₄ -C ₆ cycloalkyl or 4-7 membered heterocycloalkyl, (C ₆ -C ₁₀ aryl)-C ₁ -C ₆ alkyl, 5-10 membered heteroaryl containing from one to five heteroatoms independently selected from N, O and S, which is optionally fused with C ₅ -C ₆ cycloalkyl; (5-10-membered heteroaryl)-C ₁ -C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; (C ₆ -C ₁₀ aryl)-(5-10 membered heteroaryl)-, where the heteroaryl group contains one to five heteroatoms independently selected from N, O and S, (5-10 membered heteroaryloxy)-C ₁ - C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; (5-6 membered heteroaryl)-(5-6 membered heteroaryl)-, where each heteroaryl group contains from one to four heteroatoms independently selected from N, O and S; (4-7-membered heterocycloalkyl)-(5-6-membered heteroaryl)-, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O) _n and the heteroaryl group contains one to four heteroatoms , independently selected from N, O and S; (5-6-membered heteroaryl)-(4-7-membered heterocycloalkyl)-, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O) _n and the heteroaryl group contains one to four heteroatoms , independently selected from N, O and S; where each R ³ group is optionally substituted with one to five R ⁴ ; R ⁴ at each occurrence is independently selected from the group consisting of oxo, halo, hydroxy, cyano, phenyl, benzyl, amino, (C ₁ -C ₆ alkyl)amino optionally substituted with one to five fluorine atoms, di(C ₁ - C ₆ alkyl)amino optionally substituted with one to ten fluorine atoms, C ₁ -C ₆ alkyl optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkoxy optionally substituted with one to five fluorine atoms, C ₁ - C ₃ alkoxy-C ₁ -C ₃ alkyl optionally substituted with one to five fluorine atoms, C ₃ -C ₆ cycloalkyl optionally substituted with one to three fluorine atoms or C ₁ -C ₃ alkyl, C ₁ -C ₆ alkyl- C(O)NH- optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkyl-OC(O)NH- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl- NHC(O)NH- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl-S(O) ₂ NH- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl-C(O)- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl-S(O) _n - optionally substituted with one to five fluorine atoms or one R ⁵ ; R ⁵ is selected from phenyl, phenoxy, C ₃ -C ₆ cycloalkyl, C ₃ -C ₆ cycloalkoxy, 4-7 membered heterocycloalkyl-, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O ) _n , and 5-6-membered heteroaryl-, where the heteroaryl group contains from one to four heteroatoms independently selected from N, O and S; where each R ⁵ is optionally independently substituted with one to three halo, C ₁ -C ₃ alkyl and C ₁ -C ₃ alkoxy; and n in each case is independently selected from 0, 1 and 2.

E46 is a compound selected from the group consisting of ( 2S , 4R )-4- tert -butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidine-3- yl]ethyl}-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide; ( 2R , 4S )-4- tert -butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1-{ N- [(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide; 3-methyl- N- (trifluoroacetyl)-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- 4-(trifluoromethyl)-L-prolinamide; (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; methyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3,3-dimethyl-1-oxobutan-2-yl}carbamate; and N- (trifluoroacetyl)-L- valyl- (4R) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-4-( trifluoromethyl)-L-prolinamide; or a pharmaceutically acceptable salt thereof.

E47 is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of a compound of any of E45 and E46, or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

E48 is the way according to E47, where the coronavirus infection is COVID-19.

E49 is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of a compound according to any of E1-E30 and E45-E46, or a pharmaceutically acceptable salt thereof, wherein an additional therapeutic agent is administered and the additional therapeutic agent is selected from the group consisting of remdesivir , galidesivir, favilavir/avifavir, molnupiravir, AT-527, AT-301, BLD-2660, favipiravir, camostat, SLV213, emtrictabine/tenofivir, clevudine, dalcetrapib, boceprevir, ABX464, dexamethasone, hydrocortisone, convalescent plasma (Rhu, g-solin p65N), regdanvimab (Regkirova), ravulizumab (Ultomiris), VIR-7831/VIR-7832, BRII-196/BRII-198, COVI-AMG/COVI DROPS (STI-2020), bamlanivimab (LY-CoV555), mavrilimab, leronlimab (PRO140), AZD7442, lenzilumab, infliximab, adalimumab, JS 016, STI-1499 (COVIGUARD), lanadelumab (Takhzyro), canakinumab (Ilaris), gimsilumab, otilimab, kasirivimab/imdevimab (REGN-Cov2), MK-7110 ( CD24Fc/SACC OVID), heparin, apixaban, tocilizumab (Actemra), sarilumab (Kevzara), apilimod dimesylate, DNL758, DC402234, PB1046, dapaglifosin, abivertinib, ATR-002, bemcentinib, acalabrutinib, baricitinib, tofacitinib, losmapidine, nicloviridine, nicloviramide, famotidine diminazene.

E50 is the compound (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl -3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof.

E50a is the compound (1R, 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl -3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide.

E51 is a pharmaceutical composition containing a therapeutically effective amount of (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier.

E51a is a pharmaceutical composition containing a therapeutically effective amount of (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide together with a pharmaceutically acceptable carrier.

E52 is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine -3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof to a patient in need of its treatment.

E52a is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of (1R, 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidine -3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide to a patient in need of her treatment.

E53 is the way according to E52, where the coronavirus infection is COVID-19.

E53a is the method of E52a, where the coronavirus infection is COVID-19.

E54 is the method according to E52 or E53, where (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof is administered orally.

E54a is the method according to E52a or E53a where (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide is administered orally.

E55 is the method according to E54, where from 50 mg to 1500 mg (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof is administered every day.

E55a is the method according to E54a, where from 50 mg to 1500 mg (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide was administered every day.

E56 is the method according to E55, where 380 mg (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof is administered three times a day.

E56a is the method according to E55a, where 380 mg (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof is administered three times a day.

E57 is the method according to E55, where from 50 mg to 1500 mg (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof is administered daily as an oral suspension, capsule or tablet.

E57a is the method according to E55a, where from 50 to 1500 mg (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl ]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a pharmaceutically acceptable salt thereof is administered daily as an oral suspension, capsule or tablet.

E58 is the method of E57 where a tablet is administered.

E58a is the method of E57a where a tablet is administered.

E59 is a compound of formula I''

or a solvate or hydrate thereof, or a pharmaceutically acceptable salt of said compound, solvate or hydrate thereof;

where

R is independently at each occurrence hydroxy or oxo;

q and q ' are each independently selected from 0, 1 and 2;

p is 0, 1 or 2;

R ¹ is selected from the group consisting of C ₁ -C ₆ alkyl which is optionally substituted with cyano or from one to five fluorine atoms; C ₂ -C ₆ alkynyl; and (C ₃ -C ₆ cycloalkyl)-C ₁ -C ₃ alkyl which is optionally substituted with one to two substituents selected from trifluoromethyl and C ₁ -C ₃ alkyl, or one to five fluorine atoms;

R ² is hydrogen, or R ² and R ¹ taken together with the nitrogen and carbon atoms to which they are attached, are a pyrrolidine or piperidine ring which is optionally substituted with one to four R ^2a ;

R ^2a is at each occurrence independently selected from the group consisting of fluorine, hydroxy, C ₁ -C ₆ alkyl optionally substituted with one to three fluorine atoms, and C ₁ -C ₆ alkoxy optionally substituted with one to three fluorine atoms; or two R ^2a groups attached to adjacent carbon atoms and taken together with the atoms to which they are attached are fused C ₃ -C ₆ cycloalkyl which is optionally substituted with one to four R ^2b ; or two R ^2a groups attached to the same carbon atom and taken together with the atoms to which they are attached are spiro C ₃ -C ₆ cycloalkyl which is optionally substituted with one to four R ^2b ;

R ^2b is at each occurrence independently selected from fluorine, hydroxy, C ₁ -C ₃ alkyl optionally independently substituted with one to three fluorine or hydroxy atoms, and C ₁ -C ₃ alkoxy optionally independently substituted with one to three fluorine or hydroxy atoms;

R ³ is selected from the group consisting of C ₁ -C ₈ alkyl, C ₁ -C ₈ alkoxy, (C ₁ -C ₆ alkoxy)-C ₁ -C ₆ alkyl, C ₂ -C ₆ alkynyl, C ₂ -C ₆ alkynyloxy, C ₃ -C ₁₂ cycloalkyl optionally fused with 5-6 membered heteroaryl or phenyl, (C ₃ -C ₁₂ cycloalkyl)-C ₁ -C ₆ alkyl, C ₃ -C ₁₂ cycloalkoxy, (C ₃ -C ₁₂ cycloalkoxy)-C ₁ -C ₆ alkyl, 4-12 membered heterocycloalkyl which is optionally fused with 5-6 membered heteroaryl or phenyl, and wherein said heterocycloalkyl contains one to four heteroatoms independently selected from N, O and S( O) _n , (4-12-membered heterocycloalkyl)-C ₁ -C ₆ alkyl, where the specified heterocycloalkyl group contains from one to four heteroatoms independently selected from N, O and S(O) _n , C ₆ -C ₁₀ aryl , optionally fused with C ₄ -C ₆ cycloalkyl or 4-7 membered heterocycloalkyl, (C ₆ -C ₁₀ aryl)-C ₁ -C ₆ alkyl, 5-10 membered heteroaryl containing from one to five heteroatoms, independently selected from N, O and S, which is optionally fused with C ₅ -C ₆ cycloalkyl; (5-10-membered heteroaryl)-C ₁ -C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; (C ₆ -C ₁₀ aryl)-(5-10 membered heteroaryl)-, where the heteroaryl group contains one to five heteroatoms independently selected from N, O and S, (5-10 membered heteroaryloxy)-C ₁ - C ₆ alkyl, where the heteroaryl group contains from one to five heteroatoms independently selected from N, O and S; (5-6 membered heteroaryl)-(5-6 membered heteroaryl)-, where each heteroaryl group contains from one to four heteroatoms independently selected from N, O and S; (4-7-membered heterocycloalkyl)-(5-6-membered heteroaryl)-, where the heterocycloalkyl group contains from one to three heteroatoms independently selected from N, O and S(O) _n and the heteroaryl group contains from one to four heteroatoms , independently selected from N, O and S; (5-6-membered heteroaryl)-(4-7-membered heterocycloalkyl)-, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O) _n and the heteroaryl group contains one to four heteroatoms , independently selected from N, O and S; where each R ³ group is optionally substituted with one to five R ⁴ ;

R ⁴ at each occurrence is independently selected from the group consisting of oxo, halo, hydroxy, cyano, phenyl, benzyl, amino, (C ₁ -C ₆ alkyl)amino optionally substituted with one to five fluorine atoms, di(C ₁ - C ₆ alkyl)amino optionally substituted with one to ten fluorine atoms, C ₁ -C ₆ alkyl optionally substituted with one to five fluorine atoms, (5-6 membered heteroaryl)amino-, where the heteroaryl group contains from one to four heteroatoms independently selected from N, O and S; (4-7-membered heterocycloalkyl)amino-, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O) _n , C ₁ -C ₆ alkoxy optionally substituted with one to five fluorine atoms, C ₁ -C ₃ alkoxy-C ₁ -C ₃ alkyl optionally substituted with one to five fluorine atoms, C ₃ -C ₆ cycloalkyl optionally substituted with one to three fluorine atoms, or C ₁ -C ₃ alkyl, C ₁ -C ₆ alkyl-C(O)NH- optionally substituted with one to five fluorine atoms, C ₁ -C ₆ alkyl-OC(O)NH- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl-NHC(O)NH- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl-S(O) ₂ NH- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl-C(O)- optionally substituted with one to five fluorine atoms or one R ⁵ , C ₁ -C ₆ alkyl-S(O) _n - optionally substituted with one to five fluorine atoms or one R ⁵ ;

R ⁵ is selected from phenyl, phenoxy, C ₃ -C ₆ cycloalkyl, C ₃ -C ₆ cycloalkoxy, 4-7 membered heterocycloalkyl, where the heterocycloalkyl group contains one to three heteroatoms independently selected from N, O and S(O) _n , and 5-6-membered heteroaryl, where the heteroaryl group contains from one to four heteroatoms independently selected from N, O and S; where each R ⁵ is optionally independently substituted with one to three halogen, C ₁ -C ₃ alkyl and C ₁ -C ₃ alkoxy; and

n is independently selected from 0, 1 and 2 in each case.

E60 is the compound according to E59 which is (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- 6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a solvate or hydrate thereof, or a pharmaceutically acceptable salt of said compound, solvate or hydrate.

E61 is the compound (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl -3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide having the structure

;

;

or its solvate or hydrate.

E62 is a compound according to E61 which is crystalline ( 1R , 2S , 5S ) -N -{( 1S )-1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide.

E63 is a compound according to E62 which is crystalline ( 1R , 2S , 5S ) -N -{( 1S )-1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 1.

E64 is the compound at E62 which is crystalline (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 4.

E65 is a compound according to E61 which is amorphous ( 1R , 2S , 5S ) -N -{( 1S )-1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide.

E66 is the compound according to E61 which is ( 1R , 2S , 5S ) -N -{( 1S )-1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}- 6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl solvate.

E67 is a compound according to E66 which is crystalline ( 1R , 2S , 5S ) -N -{( 1S )-1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl solvate.

E68 is a compound according to E67 which is crystalline ( 1R , 2S , 5S ) -N -{( 1S )-1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl solvate, solid form 2.

E69 is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of a compound of any E61-E68 to a patient in need of its treatment.

E70 is the way according to E69, where the coronavirus infection is COVID-19.

E71 is the method according to E70, where ritonavir is also administered to the patient.

E72 is the method of E71 wherein the compound of any of E61-E68 and ritonavir are administered orally to the patient.

E73 is the method of E72 wherein about 10 mg to about 1500 mg per day of a compound of any of E61-E68 and about 10 mg to about 1000 mg per day of ritonavir are administered.

E74 is the method of E73 wherein about 50 mg of a compound of any of E61-E68 and about 100 mg of ritonavir are each administered to the patient twice daily.

E75 is a pharmaceutical composition containing a therapeutically effective amount of ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a solvate or hydrate thereof, or a pharmaceutically acceptable salt of said compound, solvate or hydrate, together with a pharmaceutically acceptable carrier.

E75a is a pharmaceutical composition containing a therapeutically effective amount of (1R, 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide; or a solvate or hydrate thereof, together with a pharmaceutically acceptable carrier.

E76 is a pharmaceutical composition according to E75a containing a compound according to any of E62-E68.

E77 is the method according to E69 or E70, where about 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg or 750 mg of a compound according to any of E61-E68 is administered to the patient orally twice a day.

E78 is the method according to E77, where ritonavir is co-administered orally to the patient twice a day.

E79 is the method of E78 wherein about 300 mg of a compound of any of E61-E68 and about 100 mg of ritonavir are co-administered to the patient twice daily.

E80 is an E63 compound that has a ¹⁹ F peak with a chemical shift of -73.3±0.1 ppm. and ¹³ C peaks with chemical shifts of 31.0±0.1 ppm, 27.9±0.1 ppm. and 178.9±0.2 ppm

E81 is a compound according to E64, which is characterized by one or more peaks selected from the group consisting of a ¹⁹ F peak with a chemical shift of -73.6±0.1 ppm. and ¹³ C peaks at 26.9±0.1 ppm, 21.6±0.1 ppm. and 41.5±0.1 ppm

E82 is N- (Methoxycarbonyl)-3-methyl-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}-4-(trifluoromethyl)-L-prolinamide having the structure

;

;

or its solvate or hydrate.

E83 is the compound according to E82, which is N- (Methoxycarbonyl)-3-methyl-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine -3-yl]ethyl}-4-(trifluoromethyl)-L-prolinamide.

E84 is a pharmaceutical composition containing a therapeutically effective amount of N- (Methoxycarbonyl)-3-methyl-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2- oxopyrrolidin-3-yl]ethyl}-4-(trifluoromethyl)-L-prolinamide; or a solvate or hydrate thereof, together with a pharmaceutically acceptable carrier.

E85 is a method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of an E82 or E83 compound to a patient in need of its treatment.

E86 is the way according to E85, where the coronavirus infection is COVID-19.

E87 is the method according to E85 or E86, where 10 mg to 1500 mg per day of compound E82 or E83 is administered.

E88 is the method of any one of E85-E87 wherein the compound is administered orally.

E89 is the method according to E88, where 200 mg of the compound is administered twice a day.

E90 is a method of targeting the inhibition of SARS-CoV-2 with a compound of any of E1-E30, E45-E46, E50, E50a, E59-E68 and E80-E83 as a treatment for indications caused by SARS-CoV-2 related viral infections.

E91 is a method for identifying cellular or viral pathways interfering with the function of members that can be used to treat indications caused by SARS-CoV-2 infections through the administration of a SARS-CoV-2 protease inhibitor compound according to any of E1-E30, E45-E46, E50 , E50a, E59-E68 and E80-E83.

E92 is a method of using a SARS-CoV-2 protease inhibitor compound of any of E1-E30, E45-E46, E50, E50a, E59-E68, and E80-E83 as a tool to understand the mechanism of action of other SARS-CoV-2 inhibitors.

E93 is a method of using a SARS-CoV-2 3C-like protease inhibitor compound of any of E1-E30, E45-E46, E50, E50a, E59-E68, and E80-E83 to perform gene profiling experiments to monitor gene activation or suppression for the purposes of identifying inhibitors for the treatment of indications caused by SARS-CoV-2 infections such as COVID-19.

E94 is a pharmaceutical composition for the treatment of COVID-19 in a mammal, comprising an amount of a SARS-CoV-2 3C-like protease inhibitor compound of any one of E1-E30, E45-E46, E50, E50a, E59-E68, and E80-E83 that is effective for the treatment of COVID-19, together with a pharmaceutically acceptable carrier.

E95 is a method of treating MERS in a patient, wherein the method comprises administering a therapeutically effective amount of a compound of any of E1-E30, E45-E46, E50, E50a, E59-E68, and E80-E83 to a patient in need thereof.

E96 is a method of treating MERS in a patient, wherein the method comprises administering a pharmaceutical composition of any of E31-E35, E51, E51a, E75, E75a, E84 and E94 to a patient in need thereof.

E97 is a method for inhibiting or preventing viral replication of MERS, comprising contacting a SARS-CoV-2 coronavirus 3CL protease with a therapeutically effective amount of a compound according to any one of E1-E30, 45-46, 50, 50a, 59-68 and 80-83.

E98 is a method for inhibiting or preventing MERS viral replication in a patient, comprising administering to a patient in need of inhibiting or preventing MERS viral replication a therapeutically effective amount of a compound according to any one of E1-E30, 45-46, 50, 50a, 59-68 and 80- 83.

E99 the use of a compound according to any of E1-E30, 45-46, 50, 50a, 59-68 and 80-83 for the treatment of a coronavirus infection.

E100 application of E99, where the coronavirus infection is COVID-19.

E101 the use of a compound according to any of E1-E30, 45-46, 50, 50a, 59-68 and 80-83 in the preparation of a medicament.

E102 is a compound according to any one of embodiments E1-E30, or a pharmaceutically acceptable salt thereof, for use as a drug.

E103 is a compound of any of the embodiments of E1-E30, or a pharmaceutically acceptable salt thereof, for use in a method of treatment, wherein the method is as described in any of the embodiments of E36-E41.

Brief description of the drawings

Figure 1: Powder X-ray diffraction pattern of 13, methyl t-butyl ether solvate , Solid Form 2, from the alternative synthesis of Example 13, methyl t- butyl ether solvate; obtaining a solid form 2.

Figure 2: Powder X-ray diffraction pattern of 13, methyl t-butyl ether solvate , Solid Form 2, from the second alternative synthesis from Example 13, methyl t- butyl ether solvate; obtaining a solid form 2

Figure 3: Powder X-ray diffraction pattern of Example 13, Solid Form 1, from recrystallization from Example 13; obtaining a solid form 1

Figure 4: Single crystal x-ray structure determination of Example 13, Solid Form 1. The ORTEP diagram is shown with substitution parameters at 50% probability.

Figure 5: Overlay of the X-ray powder pattern obtained for Example 13, Solid Form 1, from the recrystallization from Example 13; obtaining a solid form 1 (Figure 3) and a calculated X-ray powder diffraction pattern obtained using the Mercury program from the resolving single-crystal x-ray pattern data of Form 1 (see Single-Crystal X-ray Structure Determination of Example 13, Solid Form 1).

Figure 6: Powder X-ray diffraction pattern of Example 13, Solid Form 4, from an alternative recrystallization of Example 13; obtaining a solid form 4.

Figure 7: Single crystal x-ray determination of the structure of example 13, Solid form 4. ORTEP diagram depicted with substitution parameters at 50% probability.

Figure 8: Overlay of the X-ray powder diffraction obtained for Example 13, Solid Form 4, from an alternative recrystallization of Example 13; obtaining a solid form 4 (Figure 6) and a calculated X-ray powder diffraction pattern obtained using the Mercury program from the resolving single-crystal X-ray pattern data of Form 4 (see Single-Crystal X-ray Determination of the Structure of Example 13, Solid Form 4).

Figure 9: Powder X-ray diffraction pattern of Example 13, Solid Form 5, from Example 96.

Figure 10: Powder X-ray diffraction pattern of intermediate C16, HCl salt .

Figure 11: Powder X-ray diffraction pattern of intermediate C91 .

Figure 12: Single crystal x-ray structure determination of intermediate C91 . ORTEP diagram depicted with substitution parameters at 50% probability.

Figure 13: Powder X-ray diffraction pattern of intermediate C92 .

Figure 14: Powder X-ray diffraction pattern of intermediate C42 .

Figure 15: Single crystal x-ray structure determination of intermediate C42 . ORTEP diagram depicted with substitution parameters at 50% probability.

Detailed description of the invention

For the purposes of the present invention, as described and claimed herein, the following terms are defined as follows:

As used herein, the terms "comprising" and "including" are used in their open, non-limiting sense. As used herein, the term "treatment", unless otherwise indicated, means the treatment, alleviation, inhibition of progression, or prevention of the disorder or condition to which such term applies, or one or more of the symptoms of such disorder or condition. In methods of treating COVID-19, it should be understood that COVID-19 is a disease caused in patients by infection with the SARS-CoV-2 virus. The SARS-CoV-2 virus is to be understood as the originally discovered strain of the virus, as well as emerging mutant strains, such as, but not limited to, strains such as B.1.1.7 (variant from the UK), B.1.351 (variant from the South Africa), P.1 (variant from Brazil) and B.1.427/B.1.429 (variant from California). Used in this document, the term "treatment", unless otherwise indicated, refers to the act of treatment, in the sense that "treatment" is defined immediately above.

The term "patient" refers to warm-blooded animals such as, for example, guinea pigs, mice, rats, gerbils, cats, rabbits, dogs, cattle, goats, sheep, horses, monkeys, chimpanzees, and humans. With respect to the treatment of COVID-19, the methods of the invention are particularly useful in the treatment of humans.

The term "pharmaceutically acceptable" means that the substance or composition must be compatible, chemically and/or toxicologically, with the other ingredients in the formulation and/or with the mammal being treated with them.

The term "therapeutically effective amount" means an amount of a compound of the present invention that (i) treats or prevents a particular disease, condition, or disorder, (ii) relieves, alleviates, or eliminates one or more symptoms of a particular disease, condition, or disorder, or (iii) prevents or delays the onset of one or more symptoms of a particular disease, condition, or disorder described herein.

As used herein, the term "alkyl" refers to a straight or branched saturated hydrocarbon substituent (ie, a substituent derived from a hydrocarbon by removal of hydrogen); in one embodiment, containing from one to eight carbon atoms, in another, up to six carbon atoms, and in another, up to three carbon atoms. Non-limiting examples of such substituents include methyl, ethyl, propyl (including n- propyl and isopropyl), butyl (including n -butyl, isobutyl, sec -butyl and t -butyl), pentyl, isoamyl, hexyl, heptyl, octyl, and the like. In another embodiment, containing from one to three carbon atoms and consisting of methyl, ethyl, n -propyl and isopropyl.

As used herein, the term "alkynyl" refers to a straight or branched saturated hydrocarbon substituent that contains a carbon-carbon triple bond (ie, a substituent derived from a hydrocarbon containing a triple bond by removal of hydrogen); in one embodiment, containing from two to six carbon atoms. Non-limiting examples of such substituents include prop-2-yn-1-yl, but-3-yn-1-yl, pent-4-yn-1-yl, and hex-5-yn-1-yl.

The term "alkoxy" refers to a linear or branched saturated hydrocarbyl substituent attached to an oxygen radical (ie, a substituent derived from a hydrocarbon alcohol by removing hydrogen from OH); in one embodiment, containing from one to six carbon atoms. Non-limiting examples of such substituents include methoxy, ethoxy, propoxy (including n- propoxy and isopropoxy), butoxy (including n -butoxy, isobutoxy, sec -butoxy and tert -butoxy), pentoxy, hexoxy, and the like. In another embodiment, containing from one to three carbon atoms and consisting of methoxy, ethoxy, n- propoxy and isopropoxy. An alkoxy group that is attached to an alkyl group is called alkoxyalkyl. An example of an alkoxyalkyl group is methoxymethyl.

The term "alkynyloxy" refers to a linear or branched saturated hydrocarbyl substituent containing a carbon-carbon triple bond attached to an oxygen radical (i.e., a substituent derived from a hydrocarbon alcohol containing a triple bond by removing hydrogen from the OH group); in one embodiment, containing from three to six carbon atoms. Non-limiting examples of such substituents include propynyloxy, butynyloxy and pentynyloxy and the like.

In some cases, the number of carbon atoms in a hydrocarbyl substituent (for example, alkyl, cycloalkyl, etc.) is indicated by the prefix "C _x -C _y - " or "C _xy ", where x is the minimum and y is the maximum number of carbon atoms in deputy. Thus, for example, "C ₁ -C ₈ alkyl" or "C _1-8 alkyl" refers to an alkyl substituent containing from 1 to 8 carbon atoms, "C ₁ -C ₆ alkyl" or "C _1-6 alkyl" refers to an alkyl substituent containing 1 to 6 carbon atoms, "C ₁ -C ₃ alkyl" or "C _1-3 alkyl" refers to an alkyl substituent containing 1 to 3 carbon atoms. Illustrating further, C ₃ -C ₆ cycloalkyl or C _3-6 -cycloalkyl refers to a saturated cycloalkyl group containing from 3 to 6 carbon atoms in the ring.

The term "cycloalkyl" refers to a carbocyclic substituent obtained by removal of hydrogen from a saturated carbocyclic molecule, for example having three to seven carbon atoms. The term "cycloalkyl" includes monocyclic saturated carbocycles. The term "C ₃ -C ₇ cycloalkyl" means a radical of a three to seven membered ring system which includes cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl groups. The term "C ₃ -C ₆ cycloalkyl" means a radical of a three to six membered ring system which includes cyclopropyl, cyclobutyl, cyclopentyl and cycloalkyl groups. Cycloalkyl groups can also be bicyclic or spirocyclic carbocycles. For example, the term "C ₃ -C ₁₂ cycloalkyl" includes monocyclic carbocycles and bicyclic and spirocyclic groups such as bicyclopentyl, bicyclohexyl, bicycloheptyl, bicyclooctyl, bicyclononyl, spiropentyl, spirohexyl, spiroheptyl, spirooctyl, and spirononyl.

The term "C ₃ -C ₆ cycloalkoxy" refers to a three to six membered cycloalkyl group attached to an oxygen radical. Examples include cyclopropoxy, cyclobutoxy, cyclopentoxy and cyclohexoxy.

The term "aryl" refers to a carbocyclic aromatic system. The term "C ₆ -C ₁₀ aryl" refers to carbocyclic aromatic systems with 3-10 atoms and includes phenyl and naphthyl.

In some cases, the number of atoms in a cyclic substituent containing one or more heteroatoms (for example, heteroaryl or heterocycloalkyl) is indicated by the prefix "x-y-membered", where x is the minimum and y is the maximum number of atoms forming the cyclic group of the substituent. Thus, for example, "4-6 membered heterocycloalkyl" refers to a heterocycloalkyl containing 4 to 6 atoms, including one to three heteroatoms, in the cyclic portion of the heterocycloalkyl. Similarly, the phrase "5-6 membered heteroaryl" refers to a heteroaryl containing 5 to 6 atoms, and "5-10 membered heteroaryl" refers to a heteroaryl containing 5 to 10 atoms, each of which includes one or more heteroatoms. , in the cyclic heteroaryl group. In addition, the phrases "5-membered heteroaryl" and "6-membered heteroaryl" refer to a five-membered heteroaromatic ring system and a six-membered heteroaromatic ring system, respectively. The heteroatoms present in these ring systems are selected from N, O and S.

The term "hydroxy" or "hydroxyl" refers to -OH. When used in conjunction with another term, the prefix "hydroxy" indicates that the substituent to which the prefix is attached has been substituted with one or more hydroxy substituents. Compounds bearing a carbon having one or more hydroxy substituents include, for example, alcohols, enols, and phenol. The terms cyano and nitrile refer to the -CN group. The term "oxo" means an oxygen that is attached to a carbon atom by a double bond (ie, when R ⁴ is oxo, then R ⁴ together with the carbon to which it is attached is a C=O group).

The term "halo" or "halogen" refers to fluorine (which can be shown as -F), chlorine (which can be shown as -Cl), bromine (which can be shown as -Br), or iodine (which can be shown as like -I).

The term "heterocycloalkyl" refers to a substituent obtained by removal of hydrogen from a saturated or partially saturated ring structure containing a total of the specified number of atoms, for example 4 to 6 ring atoms or 4 to 12 atoms, where at least one of ring atoms is a heteroatom (ie, oxygen, nitrogen, or sulfur), with the remaining ring atoms being independently selected from the group consisting of carbon, oxygen, nitrogen, and sulfur. Sulfur may be oxidized [i.e. S(O) or S(O) ₂ ] or not. In a group that has a heterocycloalkyl substituent, the ring atom of the heterocycloalkyl substituent that is bonded to the group may be a nitrogen heteroatom or may be a ring carbon atom. Similarly, if the heterocycloalkyl substituent is in turn substituted with a group or substituent, the group or substituent may be bonded to a nitrogen heteroatom or may be bonded to a carbon atom of the ring. It should be understood that a heterocyclic group may be monocyclic, bicyclic, polycyclic or spirocyclic.

The term "heteroaryl" refers to an aromatic ring structure containing the specified number of ring atoms, in which at least one of the ring atoms is a heteroatom (i.e. oxygen, nitrogen or sulfur) and the remaining ring atoms are independently selected from the group, composed of carbon, oxygen, nitrogen and sulfur. Examples of heteroaryl substituents include 6-membered heteroaryl substituents such as pyridyl, pyrazyl, pyrimidinyl and pyridazinyl; and 5-membered heteroaryl substituents such as triazolyl, imidazolyl, furanyl, thiophenyl, pyrazolyl, pyrrolyl, oxazolyl, isoxazolyl, thiazolyl, 1,2,3-, 1,2,4-, 1,2,5-, or 1 ,3,4-oxadiazolyl and isothiazolyl. The heteroaryl group may also be a bicyclic heteroaromatic group such as indolyl, benzofuranyl, benzothienyl, benzimidazolyl, benzothiazolyl, benzoxazolyl, benzisoxazolyl, oxazolopyridinyl, imidazopyridinyl, imidazopyrimidinyl and the like. In a group that has a heteroaryl substituent, the ring atom of the heteroaryl substituent that is bonded to the group may be one of the heteroatoms or may be a carbon atom of the ring. Similarly, if the heteroaryl substituent is in turn substituted with a group or substituent, the group or substituent may be bonded to one of the heteroatoms, or they may be bonded to a carbon atom of the ring. The term "heteroaryl" also includes pyridyl N -oxides and groups containing a pyridine N -oxide ring. In addition, the heteroaryl group may contain an oxo group, such as the group present in the pyridone group. Additional examples include furyl, thienyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, pyridine-2( 1H )-ont, pyridazin-2(1 H )-onyl, pyrimidin-2(1 H )-onyl, pyrazin-2(1 H )-onyl, imidazo[1,2- a ]pyridinyl and pyrazolo[1,5- a ]pyridinyl. Heteroaryl may be further substituted as defined herein.

Examples of single ring heteroaryls and heterocycloalkyls include furanyl, dihydrofuranyl, tetrahydrofuranyl, thiophenyl, dihydrothiophenyl, tetrahydrothiophenyl, pyrrolyl, isopyrrolyl, pyrrolinyl, pyrrolidinyl, imidazolyl, isoimidazolyl, imidazolinyl, imidazolidinyl, pyrazolyl, pyrazolinyl, pyrazolidinyl, triazolyl, tetrazolyl, oxathiolyl oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, thiazolinyl, isothiazolinyl, thiazolidinyl, isothiazolidinyl, thiaoxadiazolyl, oxathiazolyl, oxadiazolyl (including oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl), pyranyl (including 1,2-pyranyl or 1,4-pyranyl), dihydropyranyl, pyridinyl, piperidinyl, diazinyl (including pyridazinyl, pyrimidinyl, piperazinyl, triazinyl (including sec-triazinyl, as-triazinyl and v-triazinyl), oxazinyl (in including 2H-1,2- oxazinyl , 6H-1,3- oxazinyl or 2H-1,4- oxazinyl ), isoxazinyl (including o- isoxazinyl or p -isoxazinyl), oxazolidinyl, isoxazolidinyl, oxathiazinyl ( including 1,2,5 -oxathiazinyl or 1,2,6- oxathiazinyl ), oxadiazinyl (including 2H-1,2,4-oxadiazinyl or 2H-1,2,5-oxadiazinyl), and morpholinyl.

The term "heteroaryl" may also include, if defined as such, ring systems having two rings, where such rings may be fused, and where one ring is aromatic and the other ring is not wholly part of a conjugated aromatic system (i.e. a heteroaromatic ring may be fused to a cycloalkyl or heterocycloalkyl ring). Non-limiting examples of such ring systems include 5,6,7,8-tetrahydroisoquinolinyl, 5,6,7,8-tetrahydroquinolinyl, 6,7-dihydro- 5H -cyclopenta[ b ]pyridinyl, 6,7-dihydro- 5H -cyclopenta[ c ]pyridinyl, 1,4,5,6-tetrahydrocyclopenta[ c ]pyrazolyl, 2,4,5,6-tetrahydrocyclopenta[ c ]pyrazolyl, 5,6-dihydro- 4H -pyrrolo[1,2- b ]pyrazolyl, 6,7-dihydro- 5H -pyrrolo[1,2- b ][1,2,4]triazolyl, 5,6,7,8-tetrahydro-[1,2,4]triazolo[1 ,5- a ]pyridinyl, 4,5,6,7-tetrahydropyrazolo[1,5- a ]pyridinyl, 4,5,6,7-tetrahydro-1 H -indazolyl and 4,5,6,7-tetrahydro- 2 H -indazolyl. It should be understood that if a carbocyclic or heterocyclic group can be bonded or otherwise attached to the designated substrate through various ring atoms without specifying a specific point of attachment, then all possible points are meant, whether through a carbon atom or, for example, a trivalent nitrogen atom. For example, the term "pyridyl" means 2-, 3-, or 4-pyridyl, the term "thienyl" means 2- or 3-thienyl, and so on.

If substituents are described as "independent" having more than one variable, each substituent is selected independently from the others from the list of available variables. Therefore, each substituent may be identical to or different from another substituent.

Where substituents are described as "independently selected" from a group, each substituent is selected independently of the others. Therefore, each substituent may be identical to or different from another substituent.

As used herein, the terms "Formula I ", " Formula I ' " or " Formula I''" may hereinafter be referred to as "the compound of the invention", "the present invention" and "the compound of formula I, I' or I'' " . Such terms are also defined to include all forms of formula I , I' and I'', including their hydrates, solvates, isomers, crystalline and non-crystalline forms, isomorphs, polymorphs and their metabolites. For example, the compounds of the invention, or pharmaceutically acceptable salts thereof, may exist in unsolvated and solvated forms. When the solvent or water is tightly bound, the complex will have a well-defined stoichiometry independent of humidity. However, when the solvent or water is weakly bound, as in channel solvates and hygroscopic compounds, the water/solvent content will depend on humidity and drying conditions. In such cases, non-stoichiometry will be the norm.

The compounds of the invention may exist as clathrates or other complexes. Included within the scope of the invention are complexes such as clathrates, drug-host inclusion complexes, wherein the drug and host are present in stoichiometric or non-stoichiometric amounts. Also included are complexes of the invention containing two or more organic and/or inorganic components, which may be in stoichiometric or non-stoichiometric amounts. The resulting complexes may be ionized, partially ionized, or non-ionized. An overview of such complexes is presented in J. Pharm. Sci., 64 (8), 1269-1288 by Haleblian (August 1975).

), сплошного клина (

) или пунктирного клина (

). Использование сплошной линии для изображения связей с асимметричными атомами углерода предназначено для обозначения того, что включены все возможные стереоизомеры (например, определенные энантиомеры, рацемические смеси и т.д.) на этом атоме углерода. Использование сплошного или пунктирного клина для изображения связей с асимметричными атомами углерода означает, что предполагается включение только показанного стереоизомера. Возможно, что соединения формулы I, I' и I'' могут содержать более одного асимметричного атома углерода. В этих соединениях, использование сплошной линии для изображения связей с асимметричными атомами углерода предназначено для обозначения того, что все возможные стереоизомеры должны быть включены. Например, если не указано иное, предполагается, что соединения формулы I, I' и I'' могут существовать в виде энантиомеров и диастереомеров или в виде рацематов и их смесей. Использование сплошной линии для изображения связей с одним или несколькими асимметричными атомами углерода в соединении формулы I, I' и I'' и использование сплошного или пунктирного клина для изображения связей с другими асимметричными атомами углерода в том же соединении означает, что присутствует смесь диастереомеров.The compounds of the invention have asymmetric carbon atoms. The carbon-carbon compounds of the invention may be depicted herein using a solid line (

), solid wedge (

) or dotted wedge (

). The use of a solid line to depict bonds to asymmetric carbons is intended to indicate that all possible stereoisomers (eg, certain enantiomers, racemic mixtures, etc.) on that carbon are included. The use of a solid or dotted wedge to depict bonds to asymmetric carbons means that only the stereoisomer shown is intended to be included. It is possible that the compounds of formula I , I' and I'' may contain more than one asymmetric carbon atom. In these compounds, the use of a solid line to depict bonds to asymmetric carbons is intended to indicate that all possible stereoisomers are to be included. For example, unless otherwise indicated, it is contemplated that the compounds of formula I , I' and I'' may exist as enantiomers and diastereomers or as racemates and mixtures thereof. The use of a solid line to depict bonds to one or more asymmetric carbons in a compound of formula I , I' and I'' and the use of a solid or dotted wedge to represent bonds to other asymmetric carbons in the same compound means that a mixture of diastereomers is present.

Stereoisomers of formula I , I' and I'' include cis and trans isomers, optical isomers such as R and S enantiomers, diastereomers, geometric isomers, rotational isomers, conformational isomers and tautomers of the compounds of the present invention, including compounds exhibiting more one type of isomerism; and mixtures thereof (such as racemates and diastereomeric pairs). Also included are acid addition or base addition salts where the counterion is optically active, eg D-lactate or L-lysine, or racemic, eg DL-tartrate or DL-arginine.

During the crystallization of any racemate, two different types of crystals are possible. The first type is the racemic compound (true racemate) mentioned above, where a single homogeneous crystal form is formed containing both enantiomers in equimolar amounts. The second type is a racemic mixture or conglomerate, where the two crystal forms form in equimolar amounts, each containing a single enantiomer.

Compounds of the present invention, such as compounds of formula I , I' and I'' , may exhibit the phenomenon of tautomerism; such tautomers are also considered to be compounds of the invention. All such tautomeric forms and mixtures thereof are included within the scope of the compounds of formula I , I' and I'' . Tautomers exist as mixtures of tautomers in solution. In solid form, one tautomer usually predominates. Even though a single tautomer may be described, the present invention includes all tautomers of the compounds of formula I , I' and I'' and their salts.

As used herein, the phrase "pharmaceutically acceptable salts", unless otherwise indicated, includes salts of acidic or basic groups that may be present in the compounds described herein. The compounds used in the methods of the present invention, which are basic in nature, are capable of forming a wide range of salts with various inorganic and organic acids. Acids that can be used to prepare pharmaceutically acceptable acid addition salts of such basic compounds are those that form non-toxic acid addition salts, i.e. salts containing pharmacologically acceptable anions such as acetate, benzenesulfonate, benzoate, bicarbonate, bisulfate, bitartrate , borate, bromide, calcium edetate, camsylate, carbonate, chloride, clavulanate, citrate, dihydrochloride, edetate, edislate, estolate, esylate, ethyl succinate, fumarate, glucemate, gluconate, glutamate, hexylresorcinate, hydrabamine, hydrobromide, hydrochloride, iodide, isethionate, lactate, lactobionate, laurate, malate, maleate, almond, mesylate, methyl sulfate, mucate, napsilate, nitrate, oleate, oxalate, pamoate (embonate), palmitate, pantothenate, phosphate/diphosphate, polygalacturonate, salicylate, stearate, subacetate, succinate, tannate , tartrate, theoclate, tosylate, triethiododine and valerate.

With regard to the compounds of the invention used in the methods of the invention, if the compounds also exist in tautomeric forms, then the present invention relates to these tautomers and the use of all such tautomers and mixtures thereof.

The present invention also includes compounds and methods for treating coronavirus infections such as COVID-19 and methods for inhibiting SARS-CoV-2 using isotopically labeled compounds that are identical to those listed herein, but where one or more atoms are replaced by atoms having an atomic a mass or mass number other than the atomic mass or mass number normally found in nature. Examples of isotopes that can be included in the compounds of the present invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine and chlorine, such as ² H, ³ H, ¹³ C, ¹⁴ C, ¹⁵ N, ¹⁸ O, ¹⁷ O, ³¹ P, ³² P, ³⁵ S, ¹⁸ F, and ³⁶ Cl, respectively. The compounds of the present invention, their prodrugs and pharmaceutically acceptable salts of said compounds or said prodrugs which contain the above isotopes and/or isotopes of other atoms are within the scope of the present invention. Certain isotopically labeled compounds of the present invention, for example those incorporating radioactive isotopes such as ³ H and ¹⁴ C, can be used in drug and/or substrate tissue distribution assays. Isotopes of tritium, i.e. ³ H, and carbon-14, i.e. ¹⁴ C are particularly preferred due to their ease of preparation and detection. In addition, substitution by heavier isotopes such as deuterium, i.e. ² H may confer certain therapeutic benefits due to greater metabolic stability, such as increased in vivo half-life or reduced dosage requirements, and therefore may be preferred in some circumstances. The isotopically labeled compounds used in the methods of the present invention and their prodrugs can generally be prepared by following the compounding procedures described in the art by replacing a readily available isotopically labeled reagent with a non-isotopically labeled reagent.

This invention also includes methods using pharmaceutical compositions and methods for treating coronavirus infections, such as COVID-19 infections, by administering prodrugs of the compounds of the invention. Compounds containing free amino, amido or hydroxy groups can be converted into prodrugs. Prodrugs include compounds in which an amino acid residue or polypeptide chain of two or more (eg, two, three or four) amino acid residues is covalently linked through an ester bond to the hydroxy compounds used in the methods of the present invention. Amino acid residues include, but are not limited to, the 20 naturally occurring amino acids, usually denoted by three letter symbols, and also include 4-hydroxyproline, hydroxylysine, desmosine, isodesmosine, 3-methylhistidine, norvaline, beta-alanine, gamma-aminobutyric acid, citrulline, homocysteine, homoserine, ornithine and methionine sulfone. Additional types of prodrugs are also included. For example, free hydroxy groups can be derivatized using groups including, but not limited to, hemisuccinates, phosphate esters, dimethylaminoacetates, and phosphoryloxymethyloxycarbonyls, as outlined in Advanced Drug Delivery Reviews , 1996, 19, 115. Carbamate prodrugs of hydroxy and amino groups are also included, as well as carbonate prodrugs, sulfonate esters and sulfate esters of hydroxy groups. Derivatization of hydroxy groups as (acyloxy)methyl and (acyloxy)ethyl esters, where the acyl group may be an alkyl ether optionally substituted with groups including, but not limited to, ether, amine, and carboxylic acid functional groups, or where the acyl group is a complex an amino acid ester as described above is also covered. Prodrugs of this type are described in J. Med. Chem ., 1996, 29, 10. Free amines can also be formed as amides, sulfonamides or phosphonamides. All of these prodrugs may include groups including, but not limited to, ether, amine, and carboxylic acid functional groups.

The compounds of the present invention can be used in the methods of the present invention in combination with other drugs. For example, administering to a SARS-CoV-2 coronavirus infected patient (i.e., a COVID-19 patient) a SARS-CoV-2 coronavirus 3CL protease inhibitor of the invention and an interferon such as interferon alfa or a pegylated interferon such as PEG- Intron or Pegasus may provide greater clinical benefit than dosing either interferon alone, pegylated interferon, or the SARS-CoV-2 coronavirus inhibitor alone. Other additional agents that can be used in the methods of the present invention include dexamethasone, azithromycin, and remdesivir. Examples of greater clinical benefits may include greater reduction in COVID-19 symptoms, faster time to symptom relief, reduced lung pathology, greater reduction in patient SARS-CoV-2 (viral load), and reduced mortality.

SARS-CoV-2 coronavirus infects cells expressing P-glycoprotein. Some of the SARS-CoV-2 coronavirus 3CL protease inhibitors of the invention are P-glycoprotein substrates. Compounds that inhibit the SARS-CoV-2 coronavirus, which are also P-glycoprotein substrates, can be dosed with a P-glycoprotein inhibitor. Examples of P-glycoprotein inhibitors are verapamil, vinblastine, ketoconazole, nelfinavir, ritonavir or cyclosporine. The P-glycoprotein inhibitors act by inhibiting the outflow of the inhibitors of the SARS-CoV-2 coronavirus of the invention from the cell. Inhibition of P-glycoprotein-based efflux will prevent a decrease in intracellular concentrations of SARS-CoV-2 coronavirus inhibitor due to P-glycoprotein efflux. Inhibition of P-glycoprotein efflux will result in higher intracellular concentrations of SARS-CoV-2 coronavirus inhibitors. Dosing of a SARS-CoV-2 coronavirus infected patient with the SARS-CoV-2 coronavirus 3CL protease inhibitors of the invention and a P-glycoprotein inhibitor can reduce the amount of SARS-CoV-2 coronavirus 3CL protease inhibitor needed to achieve an effective dose by increasing intracellular concentration 3CL protease inhibitor of SARS-CoV-2 coronavirus.

Among the agents that can be used to increase the exposure of a compound of the present invention to a mammal are those that can act as inhibitors of at least one isoform of cytochrome P450 (CYP450) enzymes. Isoforms of CYP450 that can be effectively inhibited include, but are not limited to, CYP1A2, CYP2D6, CYP2C9, CYP2C19, and CYP3A4. Compounds useful in the methods of the present invention include compounds that may be CYP3A4 substrates and are metabolized by CYP3A4. Dosing to a patient infected with SARS-CoV-2 coronavirus an inhibitor of SARS-CoV-2 coronavirus that is a CYP3A4 substrate, such as an inhibitor of SARS-CoV-2 coronavirus 3CL protease, and a CYP3A4 inhibitor, such as ritonavir, nelfinavir, or delavirdine, will decrease metabolism inhibitor of SARS-CoV-2 coronavirus using CYP3A4. This will lead to a decrease in the clearance of the SARS-CoV-2 coronavirus inhibitor and an increase in the plasma concentration of the SARS-CoV-2 coronavirus inhibitor. Reduced clearance and higher plasma concentrations may result in a lower effective dose of the SARS-CoV-2 coronavirus inhibitor.

Additional therapeutic agents that can be used in combination with SARS-CoV-2 inhibitors in the methods of the present invention include the following:

PLpro inhibitors, apilomod, EIDD-2801, ribavirin, valganciclovir, β -thymidine, aspartame, oxprenolol, doxycycline, acetophenazine, iopromide, riboflavin, reproterol, 2,2'-cyclocytidine, chloramphenicol, chlorphenesin carbamate, levodropropizine, cefamandol, tigecycline , pemetrexed, L(+)-ascorbic acid, glutathione, hesperetin, ademethionine, mazoprocol, isotretinoin, dantrolene, antibacterial sulfasalazine, silybin, nicardipine, sildenafil, platicodin, chrysin, neohesperidin, baicalin, 9 sugetriol-3,9-diacetate, ( -)-epigallocatechin gallate, phytanthrine D, 2-(3,4-dihydroxyphenyl)-2-[[2-(3,4-dihydroxyphenyl)-3,4-dihydro-5,7-dihydroxy-2 H -1- benzopyran-3-yl]oxy]-3,4-dihydro-2H-1-benzopyran- 3,4,5,7 -tetrol, 2,2-di(3-indolyl)-3-indolone, ( S ) -(1 S ,2 R ,4a S ,5 R ,8a S )-1-formamido-1,4a-dimethyl-6-methylene-5-(( E )-2-(2-oxo-2,5- dihydrofuran-3-yl)ethenyl)decahydronaphthalene-2-yl-2-amino-3-phenylpropanoate, piceatannol, rosmarinic acid and magnolol.

3CLpro inhibitors, lymecycline, chlorhexidine, alfuzosin, cilastatin, famotidine, almitrin, progabid, nepafenac, carvedilol, amprenavir, tigecycline, montelukast, carminic acid, mimosin, flavin, lutein, cefpyramide, feneticillin, candoxatril, nicardipine, estradiol, piglyvalerate, conivaptan, telmisartan, doxycycline, oxytetracycline, (1 S ,2 R ,4a S ,5 R ,8a S )-1-formamido-1,4a-dimethyl-6-methylene-5-(( E )-2-(2 -oxo-2,5-dihydrofuran-3-yl)ethenyl)decahydronaphthalene-2-yl 5-(( R )-1,2-dithiolan-3-yl)pentanoate, betulonal, chrysin-7- O - β -glucuronide , andrographizide, (1 S ,2 R ,4a S ,5 R ,8a S )-1-formamido-1,4a-dimethyl-6-methylene-5-(( E )-2-(2-oxo-2, 5-dihydrofuran-3-yl)ethenyl)decahydronaphthalene-2- yl2 -nitrobenzoate, 2β-hydroxy-3,4-secofridelolactone-27th acid ( S )-(1 S ,2 R ,4a S ,5 R , 8a S )-1-formamido-1,4a-dimethyl-6-methylene-5-(( E )-2-(2-oxo-2,5-dihydrofuran-3-yl)ethenyl)decahydronaphthalene-2-yl- 2-amino-3-phenylpropanoate, isodecortinol, cerevisterol, hesperidin, neohesperidin, an droprapanine, 2-((1 R ,5 R ,6 R ,8a S )-6-hydroxy-5-(hydroxymethyl)-5,8a-dimethyl-2-methylenedecahydronaphthalen-1-yl)ethylbenzoate, cosmosiin, cleistocalton A, 2,2-di(3-indolyl)-3-indolone, biorobin, gnidicine, phyllaemblinol, theaflavin 3,3'-di- O -gallate, rosmarinic acid, kuichenside I, oleanolic acid, stigmast-5- ene -3- ol, deacetylcentapicrin and berghemol.

RdRp inhibitors, valganciclovir, chlorhexidine, ceftibuten, fenoterol, fludarabine, itraconazole, cefuroxime, atovaquone, chenodeoxycholic acid, cromolyn, pancuronium bromide, cortisone, tibolone, novobiocin, silubin , idarubicin ,30 β -dihydroxy-3,4-secofridelolactone-27-lactone, 14-deoxy-11,12-didehydroandrographolide, gnidithrin, theaflavin 3,3'-di- O -gallate, ( R )-((1 R ,5a S ,6 R ,9a S )-1,5a-dimethyl-7-methylene-3-oxo-6-(( E )-2-(2-oxo-2,5-dihydrofuran-3-yl)ethenyl)decahydro - 1H -benzo[ c ]azepin-1-yl)methyl2-amino-3-phenylpropanoate, 2β-Hydroxy-3,4- secofridelolactone -27-oic acid, 2-(3,4-dihydroxyphenyl)-2-[ [2-(3,4-dihydroxyphenyl)-3,4-dihydro-5,7-dihydroxy-2H-1- benzopyran -3-yl]oxy]-3,4-dihydro-2H-1- benzopyran- 3,4,5,7-tetrol, phyllaemblicin B, 14-hydroxycyperotundone, andrographizide, 2-((1 R ,5 R ,6 R ,8a S )-6-hydroxy-5-(hydroxymethyl)-5,8a- dimethyl-2-methylenedecahydronaphthalene-1-yl)ethylbenzoate, and rografolide, sugetriol-3,9-diacetate, Baicalin, (1 S ,2 R ,4a S ,5 R ,8a S )-1-formamido-1,4a-dimethyl-6-methylene-5-(( E )- 2-(2-oxo-2,5-dihydrofuran-3-yl)ethenyl)decahydronaphthalene-2-yl 5-(( R )-1,2-dithiolan-3-yl)pentanoate, 1,7-dihydroxy-3 -methoxexanthone, 1,2,6-trimethoxy-8-[(6- O - β -D-xylopyranosyl- β -D-glucopyranosyl)oxy]-9 H -xanthen-9-one and 1,8-dihydroxy-6 -methoxy-2-[(6- O - β -D-xylopyranosyl- β -D-glucopyranosyl)oxy]-9 H -xanthen-9-one, 8-( β -D-glucopyranosyloxy)-1,3,5 -trihydroxy -9H-xanthen-9-one.

Additional therapeutic agents that can be used in the methods of the present invention include diosmin, hesperidin, MK-3207, venetoclax, dihydroergocristine, bolazine, R428, ditherkalinium, etoposide, teniposide, UK-432097, irinotecan, lumacaftor, velpatasvir, eluxadoline, ledipasvir, lopinavir/ritonavir+ribavirin, alferon and prednisone. Other additional agents used in the methods of the present invention include dexamethasone, azithromycin, and remdesivir, as well as boceprevir, umifenovir, and favipiravir.

Other additional agents that can be used in the methods of the present invention include compoundsα—ketoamides, designated as 11r, 13a and 13b, shown below, as described in Zhang, L.; Lin, D.; Sun, X.; Rox, K.; Hilgenfeld, R.; X-ray Structure of Main Protease of the Novel Coronavirus SARS-CoV-2 Enables Design ofα-Ketoamide Inhibitors; bioRxiv preprint doi: https://doi.org/10.1101/2020.02.17.952879

Additional agents that may be used in the methods of the present invention include RIG 1 pathway activators such as those described in US Pat. No. 9,884,876.

Other additional therapeutic agents include protease inhibitors such as those described in Dai W, Zhang B, Jiang XM, et al. Structure-based design of antiviral drug candidates targeting the SARS-CoV-2 main protease. science . 2020;368(6497):1331-1335, including compounds such as those shown below and compounds designated as DC402234

.

.

Another embodiment of the present invention is a method of treating COVID-19 in a patient, wherein in addition to administering a compound of the present invention (i.e., a compound of formula I, I' or I'' or a solvate or hydrate thereof or a pharmaceutically acceptable salt thereof, a compound or solvate or hydrate) an additional agent is administered and this additional agent is selected from antivirals such as remdesivir, galidesivir, favilavir/avifavir, molnupiravir (MK-4482/EIDD 2801), AT-527, AT-301, BLD-2660, favipiravir , camostat, SLV213 emtrictabine/tenofivir, clevudine, dalcetrapib, boceprevir and ABX464, glucocorticoids such as dexamethasone and hydrocortisone, convalescent plasma, recombinant human plasma such as gelsolin (Rhu-p65N), monoclonal antibodies such as regdanvimab (Regkirova), ravulizumab (Ultomiris), VIR-7831/VIR-7832, BRII-196/BRII-198, COVI-AMG/COVI DROPS (STI-2020), bamlanivimab (LY-CoV555), mavrilimab, leronlimab (PRO140), AZD7442, lenzilumab , inf liximab, adalimumab, JS 016, STI-1499 (COVIGUARD), lanadelumab (Takhzyro), canakinumab (Ilaris), gimsilumab and otilimab, antibody cocktails such as casirivimab/imdevimab (REGN-Cov2), recombinant fusion protein such as MK- 7110 (CD24Fc/SACCOVID), anticoagulants such as heparin and apixaban, IL-6 receptor agonists such as tocilizumab (Actemra) and sarilumab (Kevzara), PIKfyve inhibitors such as apilimod dimesylate, RIPK1 inhibitors such as DNL758, DC10462234, VIP receptor agonists such as PB1046 SGLT2 inhibitors such as dapaglifosine TYK inhibitors such as abivertinib Kinase inhibitors such as ATR-002 , such as niclosamide, furin inhibitors, such as diminaschen.

The term "SARS-CoV-2 inhibitory agent" means any SARS-CoV-2 coronavirus-related 3C-like protease inhibitor compound described herein, or a pharmaceutically acceptable salt, hydrate, prodrug, active metabolite, or solvate thereof, or a compound that inhibits SARS-CoV-2 replication by any means.

The term "intervention or prevention" of viral replication of the SARS-CoV-2-related coronavirus ("SARS-CoV-2") in a cell means the reduction of SARS-CoV-2 replication or the production of SARS-CoV-2 components necessary for the progeny of the virus in the cell treated with the compound of the present invention compared to a cell not treated with the compound of the present invention. Simple and convenient assays for determining a reduction in SARS-CoV-2 virus replication include an ELISA assay for the presence, absence, or reduced presence of anti-SARS-CoV-2 antibodies in a subject's blood (Nasoff, et al., PNAS 88:5462-5466, 1991 ), RT-PCR (Yu, et al., in Viral Hepatitis and Liver Disease 574-577, Nishioka, Suzuki and Mishiro (Eds.); Springer-Verlag, Tokyo, 1994). Such methods are well known to those skilled in the art. Alternatively, total RNA from transduced and infected "control" cells can be isolated and analyzed by dot blot or Northern blot and probed for SARS-CoV-2 specific DNA to determine if SARS-CoV-2 replication is reduced. . Alternatively, decreased SARS-CoV-2 protein expression can also be used as an indicator of SARS-CoV-2 replication inhibition. A reduction in SARS-CoV-2 replication of more than fifty percent compared to control cells usually quantifies the prevention of SARS-CoV-2 replication.

If the SARS-CoV-2 inhibitor compound used in the method of the invention is a base, the desired salt can be obtained by any suitable method known in the art, including treatment of the free base with an inorganic acid (such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like) or with an organic acid (such as acetic acid, maleic acid, succinic acid, mandelic acid, fumaric acid, malonic acid, pyruvic acid, oxalic acid, glycolic acid, salicylic acid, pyranosidyl acid (such as glucuronic acid or galacturonic acid), alpha hydroxy acid (such as citric acid or tartaric acid), amino acid (such as aspartic acid or glutamic acid), aromatic acid (such as benzoic acid or cinnamic acid), sulfonic acid (such as p- toluenesulfonic acid or ethanesulfonic acid lot) and the like.

If the SARS-CoV-2 inhibitor compound used in the method of the invention is an acid, the desired salt may be prepared by any suitable method known in the art, including treatment of the free acid with an inorganic or organic base [such as an amine (primary, secondary, or tertiary )], alkali metal hydroxide or alkaline earth metal hydroxide. Illustrative examples of suitable salts include organic salts derived from amino acids (such as glycine and arginine), ammonia, primary amines, secondary amines, tertiary amines, and cyclic amines (such as piperidine, morpholine, and piperazine), as well as inorganic salts derived from sodium. , calcium, potassium, magnesium, manganese, iron, copper, zinc, aluminum and lithium.

In the case of SARS-CoV-2 inhibitor compounds, prodrugs, salts, or solvates that are solids, it will be understood by those skilled in the art that the compound, prodrugs, salts, and solvates used in the method of the present invention may exist in various polymorphs or crystalline forms, all of which are within the scope of the present invention and the stated formulas. In addition, the compound, salts, prodrugs and solvates used in the method of the present invention may exist as tautomers, all of which are intended to be included within the broad scope of the present invention.

Solubilizing agents can also be used with the compounds of the invention to increase the aqueous solubility of the compounds in physiologically acceptable solutions. These solubilizing agents include cyclodextrins, propylene glycol, diethylacetamide, polyethylene glycol, Tween, ethanol, and micelle forming agents. Suggested solubilizing agents are cyclodextrins, especially beta-cyclodextrins and in particular hydroxypropyl beta-cyclodextrin and sulfobutyl ether beta-cyclodextrin.

In some cases, SARS-CoV-2 inhibitor compounds, salts, prodrugs and solvates used in the method of the present invention may have chiral centers. When chiral centers are present, the compound, salts, prodrugs, and solvates may exist as single stereoisomers, racemates, and/or mixtures of enantiomers and/or diastereomers. All such individual stereoisomers, racemates, and mixtures thereof are intended to be included within the broad scope of the present invention.

As generally understood by those skilled in the art, an optically pure compound is one that is enantiomerically pure. As used herein, the term "optically pure" is intended to refer to a compound having at least sufficient activity. Preferably, an optically pure amount of one enantiomer to obtain a compound having the desired pharmacologically pure compound of the present invention comprises at least 90% of one isomer (80% enantiomeric excess), more preferably at least 95% (90% ee) , even more preferably at least 97.5% (95% ee), and most preferably at least 99% (98% ee).

The term "treatment" as used herein, unless otherwise indicated, means the treatment, alleviation, inhibition of development, or prevention of the disorder or condition to which the term is applied, or one or more symptoms of such condition or disorder. The term "treatment" as used herein, unless otherwise indicated, refers to the act of treating in the sense that "treating" is defined immediately above. In a preferred embodiment of the present invention, "treat" or "treatment" means at least alleviating a disease state in a human that is alleviated by inhibiting the activity of SARS-COV-2 3C-like protease, which is the main protease of SARS-COV-2 , the causative agent of COVID-19. For patients suffering from COVID-19, fever, fatigue, and dry cough are the main manifestations of the disease, while nasal congestion, runny nose, and other upper respiratory symptoms are rare. The Beijing Centers for Diseases Control and Prevention indicates that a typical case of COVID-19 causes a progressive exacerbation. COVID-19 can be classified into mild, normal, severe, and critical types based on the severity of the disease. National Health Commission of the People 's Republic of China. Diagnosis and Treatment of Pneumonia Caused by 2019-nCoV (Trial Version 4). Available online: http://www.nhc.gov.cn/jkj/s3577/202002/573340613ab243b3a7f61df260551dd4/files/c791e5a7ea5149f680fdcb34dac0f54e.pdf: (1) Mild cases - clinical symptoms were mild and no inflammation of the lungs was detected on CT scan ( CT) chest; (2) normal cases - fever, respiratory symptoms, and found that patients had visualized manifestations of pneumonia; (3) severe cases - one of the following three conditions: respiratory distress, respiratory rate ≥30 times / min (at rest, refer to oxygen saturation ≤93%), partial oxygen pressure (PaO2) / oxygen absorption concentration (FiO2 ) ≤300mmHg (1 mm Hg = 0133 kPa); (4) critical cases - one of the following three conditions: respiratory failure and the need for mechanical ventilation, shock, or associated failure of other organs requiring intensive care. Current clinical data show that the majority of deaths occurred among elderly patients. However, severe cases have been reported in young people who have unique factors, in particular chronic diseases such as diabetes or hepatitis B. Those who have taken hormones or immunosuppressants for a long time and have reduced immune functions are likely to be highly infected. .

Methods of treatment for alleviating a disease state associated with a coronavirus, such as COVID-19, include the use of one or more compounds of the invention in any conventional manner. According to some preferred embodiments of the invention, the compound or compounds used in the methods of the present invention is administered to a mammal, such as a human, if necessary. Preferably, the mammal in need thereof is infected with a coronavirus such as the causative agent of COVID-19, namely SARS-CoV-2.

The present invention also includes prophylactic methods comprising administering an effective amount of the SARS-CoV-2 inhibitor of the invention, or a pharmaceutically acceptable salt, prodrug, pharmaceutically active metabolite, or solvate thereof, to a mammal, such as a human, at risk for SARS-CoV-2 infection. In some preferred embodiments, an effective amount of one or more compounds of the present invention, or a pharmaceutically acceptable salt, prodrug, pharmaceutically active metabolite, or solvate thereof, is administered to a human subject at risk for SARS-CoV-2 infection, the causative agent of COVID-19. Prophylactic methods of the invention include the use of one or more compounds of the invention in any conventional manner.

Certain compounds used in the methods of the invention, such as dexamethasone, azithromycin and remdesivir, are known and can be prepared by methods known in the art.

Recent data indicate that the novel coronavirus SARS-CoV-2 is the causative agent of COVID-19. The nucleotide sequence of the SARS-CoV-2 coronavirus, as well as the recently identified L and S subtypes, have recently been identified and made available to the public.

The activity of inhibitor compounds as inhibitors of SARS-CoV-2 viral activity can be measured by any of the suitable methods available in the art, including in vivo and in vitro assays . The activity of the compounds of the present invention as inhibitors of coronavirus 3C-like protease activity (such as SARS-CoV-2 coronavirus 3C-like protease) can be measured by any of the suitable methods known to those skilled in the art, including in vivo assays and in vitro . Examples of suitable assays for measuring activity include the antiviral cell culture assays described herein as well as the antiprotease assays described herein, such as those described in the experimental section.

Administration of the SARS-CoV-2 inhibitor compounds and their pharmaceutically acceptable prodrugs, salts, active metabolites and solvates may be by any of the conventional routes of administration available to those skilled in the art. Illustrative examples of suitable routes of administration include oral, nasal, pulmonary, parenteral, topical, intravenous, injection, transdermal, and rectal. Oral, intravenous, subcutaneous and nasal delivery is preferred.

The SARS-CoV-2 inhibitory agent may be administered as a pharmaceutical composition in any suitable pharmaceutical form. Suitable pharmaceutical forms include solid, semi-solid, liquid or lyophilized formulations such as tablets, powders, capsules, suppositories, suspensions, liposomes and aerosols. The SARS-CoV-2 inhibitory agent may be formulated into a solution using any of a variety of techniques. For example, a SARS-CoV-2 inhibitory agent can be dissolved in an acid (e.g., 1 M HCl) and diluted with a sufficient volume of a 5% dextrose in water (D5W) solution to produce the desired final concentration of the SARS-CoV-2 inhibitory agent ( for example, about 15 mM). Alternatively, a D5W solution containing about 15 mM HCl can be used to prepare a solution of the SARS-CoV-2 inhibitory agent at the appropriate concentration. In addition, the SARS-CoV-2 inhibitory agent can be prepared as a suspension using, for example, a 1% solution of carboxymethyl cellulose (CMC).

Acceptable methods for the preparation of suitable pharmaceutical forms of pharmaceutical compositions are known or can be determined in the usual way by experts in this field of technology. For example, pharmaceutical preparations may be prepared according to conventional pharmaceutical chemistry techniques, including steps such as mixing, granulating and compressing, when necessary, for tablet forms, or mixing, filling, and dissolving ingredients, if necessary, to obtain the desired intravenous products. , oral, parenteral, topical, intravaginal, intranasal, intrabronchial, intraocular, intraaural and/or rectal administration.

Typically, the compound of the present invention is administered in an amount effective to treat the condition described herein. The compounds of the invention are administered by any suitable route in the form of a pharmaceutical composition adapted to such route and at a dose effective for the intended treatment. Therapeutically effective doses of the compounds required to treat the progression of a medical condition are readily determined by one of ordinary skill in the art using preclinical and clinical approaches familiar to those of skill in the medical arts.

The compounds of the invention may be administered orally. Oral administration may include swallowing, so that the compound enters the gastrointestinal tract, or buccal or sublingual administration may be used, in which the compound enters the bloodstream directly from the mouth.

In another embodiment, the compounds of the invention can also be administered directly to the bloodstream, to a muscle, or to an internal organ. Suitable methods of parenteral administration include intravenous, intraarterial, intraperitoneal, intrathecal, intraventricular, intraurethral, intrasternal, intracranial, intramuscular and subcutaneous. Suitable devices for parenteral administration include needle (including microneedle) injectors, needleless injectors and infusion techniques.

In another embodiment, the compounds of the invention can also be administered topically to the skin or mucosa, ie dermally or transdermally. In another embodiment, the compounds of the invention can also be administered intranasally or by inhalation. In another embodiment, the compounds of the invention may be administered rectally or vaginally. In another embodiment, the compounds of the invention can also be administered directly to the eye or ear.

The dosing schedule for compounds and/or compositions containing compounds is based on a variety of factors, including the type, age, weight, sex, and medical condition of the patient; the severity of the condition; method of administration; and the activity of the particular compound used. Thus, the dosing regimen can vary widely. Dosage levels from about 0.01 mg to about 100 mg per kilogram of body weight per day are useful in the treatment of the above conditions. In one embodiment, the total daily dose of a compound of the present invention (administered as a single or divided dose) is typically from about 0.01 mg/kg to about 100 mg/kg. In another embodiment, the total daily dose of the compound of the present invention is from about 0.1 to about 50 mg/kg, and in another embodiment, from about 0.5 to about 30 mg/kg (i.e. mg of the compound of the present invention per kg of body weight). In one embodiment, the dosage is from 0.01 to 10 mg/kg/day. In another embodiment, the dosage is 0.1 to 1.0 mg/kg/day. Unit dosage compositions may contain such amounts, or multiples thereof, to constitute a daily dose. In many cases, the administration of the compounds will be repeated multiple times per day (typically no more than 4 times). If desired, multiple doses per day can be used to increase the total daily dose.

For oral administration, the compositions may be presented in the form of tablets containing from about 0.01 mg to about 500 mg of the active ingredient, or, in another embodiment, from about 1 mg to about 100 mg of the active ingredient. When administered intravenously, doses may vary from about 0.1 to about 10 mg/kg/min at a constant infusion rate.

Suitable patients according to the present invention include mammalian patients. Mammals of the present invention include, but are not limited to, dogs, cats, cattle, goats, horses, sheep, pigs, rodents, lagomorphs, primates, and the like, and include mammals in the womb. In one embodiment, suitable patients are humans. Human patients can be of any gender and at any stage of development.

In another embodiment, the invention includes the use of one or more compounds of the invention for the preparation of a medicament for the treatment of the conditions listed herein.

For the treatment of the conditions mentioned above, the compound of the present invention may be administered as a compound per se. Alternatively, pharmaceutically acceptable salts are suitable for medical use due to their greater aqueous solubility compared to the parent compound.

In another embodiment, the present invention includes pharmaceutical compositions. Such pharmaceutical compositions contain a compound of the invention, presented with a pharmaceutically acceptable carrier. A pharmaceutically acceptable carrier includes any suitable dosage form suitable for administration to a patient. The carrier may be a solid, liquid, or both, and may be formulated with the compound in a single dose formulation, eg, a tablet, which may contain from 0.05% to 95% by weight of the active compounds. The compound of the present invention may be associated with suitable polymers as targeted drug carriers. Other pharmacologically active substances may also be present.

The compounds of the present invention may be administered by any suitable route, preferably in the form of a pharmaceutical composition adapted to such route and at a dose effective for the intended treatment. Active compounds and compositions, for example, can be administered orally, rectally, parenterally or topically.

Oral administration of a solid dosage form may, for example, be presented in discrete units such as hard or soft capsules, pills, cachets, lozenges or tablets, each containing a predetermined amount of at least one compound of the present invention. In another embodiment, oral administration may be in the form of a powder or granules. In another embodiment, the oral dosage form is sublingual, such as, for example, a lozenge. In such solid dosage form, the compounds of the present invention are usually combined with one or more adjuvants. Such capsules or tablets may contain a controlled release formulation. In the case of capsules, tablets and pills, the dosage form may also contain buffering agents or may be formulated with enteric coatings.

In another embodiment, oral administration may be in liquid dosage form. Liquid dosage forms for oral administration include, for example, pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs containing inert diluents commonly used in the art (eg, water). Such compositions may also contain adjuvants such as wetting, emulsifying, suspending, flavoring (eg sweetening) and/or flavoring agents.

In another embodiment, the present invention includes a parenteral dosage form. "Parenteral administration" includes, for example, subcutaneous injections, intravenous injections, intraperitoneal injections, intramuscular injections, intrasternal injections, and infusions. Injectable preparations (eg, sterile injectable aqueous or oily suspensions) can be formulated according to the known art using suitable dispersing, wetting and/or suspending agents.

In another embodiment, the present invention includes a topical dosage form. "Topical administration" includes, for example, transdermal administration, such as transdermal patches or iontophoresis devices, intraocular administration, or intranasal or inhalation administration. Topical compositions also include, for example, topical gels, sprays, ointments and creams. The topical formulation may include a compound that enhances the absorption or penetration of the active ingredient through the skin or other affected areas. When the compounds of the present invention are administered via a transdermal device, administration will be by either a reservoir-type patch, a porous membrane, or a solid matrix. Typical formulations for this purpose include gels, hydrogels, lotions, solutions, creams, ointments, powders, dressings, foams, films, skin patches, wafers, implants, sponges, fibers, dressings, and microemulsions. You can also use liposomes. Typical carriers include alcohol, water, mineral oil, liquid petrolatum, white petrolatum, glycerin, polyethylene glycol, and propylene glycol. Penetration enhancers may be included; see, for example, J. Pharm. Sci., 88 (10), 955-958, by Finnin and Morgan (October 1999).

Compositions suitable for topical administration to the eye include, for example, eye drops wherein the compound of the present invention is dissolved or suspended in a suitable vehicle. Typical formulations suitable for ophthalmic or otic administration may be in the form of drops of a micronized suspension or solution in isotonic pH-adjusted sterile saline. Other formulations suitable for ophthalmic and otic administration include ointments, biodegradable (eg, absorbable gel sponges, collagen) and non-biodegradable (eg, silicone) implants, cachets, lenses, and particle or vesicle systems such as niosomes or liposomes. A polymer such as cross-linked polyacrylic acid, polyvinyl alcohol, hyaluronic acid, a cellulose polymer such as hydroxypropyl methylcellulose, hydroxyethylcellulose or methylcellulose, or a heteropolysaccharide polymer such as gelan gum may be included along with a preservative such as benzalkonium chloride. Such drugs can also be delivered by iontophoresis.

For intranasal or inhalation administration, the active compounds of the present invention are conveniently delivered in the form of a solution or suspension from a pump nebulizer that is compressed or pumped by the patient, or as an aerosol spray from a pressurized container or nebulizer using a suitable propellant. Formulations suitable for intranasal administration are usually administered in dry powder form (either alone, as a mixture, eg dry-mixed with lactose, or as a mixed complex particle, eg mixed with phospholipids such as phosphatidylcholine) from a dry powder inhaler or as an aerosol spray from a pressurized container, pump, spray, nebulizer (preferably an EHD nebulizer to produce a fine mist), or nebulizer, with or without the use of a suitable propellant such as 1,1,1,2-tetrafluoroethane or 1,1,1,2,3,3,3-heptafluoropropane. For intranasal use, the powder may contain a bioadhesive agent such as chitosan or cyclodextrin.

In another embodiment, the present invention includes a rectal dosage form. Such a rectal dosage form may be in the form of, for example, a suppository. Cocoa butter is the traditional base for suppositories, but various alternatives can be used if necessary.

Other carrier materials and routes of administration known in the pharmaceutical art may also be used. Pharmaceutical compositions of the present invention can be prepared using any of the well-known pharmaceutical techniques, such as efficient formulation techniques and administration procedures. The above considerations for efficient drafting and insertion techniques are well known in the art and are described in standard textbooks. Drug formulation is discussed, for example, in Hoover, John E., Remington 's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania, 1975; Liberman et al., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, NY, 1980; and ^Kibbe et al., Eds., Handbook of Pharmaceutical Excipients (3rd Ed.), American Pharmaceutical Association, Washington, 1999.

The compounds of the present invention can be used alone or in combination with other therapeutic agents in the treatment of various conditions or disease states. The compound of the present invention and the other therapeutic agent may be administered simultaneously (either in the same dosage form or in separate dosage forms) or sequentially. Two or more compounds can be administered simultaneously, in parallel or sequentially. In addition, simultaneous administration can be carried out by mixing the compounds before administration, or by administering the compounds at the same time but at different anatomical sites, or using different routes of administration. The phrases "parallel administration", "co-administration", "simultaneous administration" and "administer simultaneously" mean that the compounds are administered in combination.

The present invention includes the use of a combination of a compound of the invention and one or more additional therapeutic agents. If a combination of active agents is administered, they may be administered sequentially or simultaneously, in a separate dosage form, or in combination in a single dosage form. Accordingly, the present invention also includes pharmaceutical compositions containing an amount of: (a) a first agent containing a compound of the invention or a pharmaceutically acceptable salt of the compound; (b) a second therapeutic agent; and (c) a pharmaceutically acceptable carrier. Pharmaceutical compositions of the invention may also include suitable excipients, diluents, carriers and carriers, as well as other pharmaceutically active agents, depending on the intended use. Pharmaceutical compositions may use solid or liquid pharmaceutically acceptable carriers, diluents, carriers or excipients. Illustrative solid carriers include starch, lactose, calcium sulfate dihydrate, calcium sulfate, sucrose, talc, gelatin, pectin, gum arabic, magnesium stearate, and stearic acid. Exemplary liquid carriers include syrup, peanut butter, olive oil, saline, and water. The carrier or diluent may include a suitable sustained release material such as glyceryl monostearate or glyceryl distearate alone or with a wax. When a liquid carrier is used, the formulation may be in the form of a syrup, elixir, emulsion, soft gelatin capsule, sterile injectable liquid (eg, solution), or non-aqueous or aqueous liquid suspension.

The dose of the pharmaceutical composition may contain at least a therapeutically effective amount of an agent that inhibits SARS-CoV-2, and preferably consists of one or more pharmaceutical dosage units. The selected dose may be administered to a mammal, e.g., a human patient, in need of treatment mediated by inhibition of SARS-CoV-2 related coronavirus activity by any known or suitable dosing route, including topical, e.g., as an ointment or cream; oral, rectal, for example, in the form of a suppository; parenteral, by injection; intravenous; or continuous by intravaginal, intranasal, intrabronchial, intraaural or intraocular infusion.

The phrases "therapeutically effective amount" and "effective amount" are intended to mean an amount of an agent of the invention which, when administered to a mammal in need of treatment, is sufficient to treat injuries or disease states ameliorated by inhibition of SARS-CoV-2 virus replication. The amount of a given SARS-CoV-2 inhibitory agent used in the method of the present invention that will be therapeutically effective will vary depending on factors such as the particular SARS-CoV-2 inhibitory agent, disease state and severity, personality and the characteristics of the mammal in need thereof, the amount of which can be determined in the usual manner by those skilled in the art.

It should be understood that the actual dosages of the SARS-CoV-2 inhibitory agents used in the pharmaceutical compositions of the present invention will be selected according to the properties of the particular agent employed, the particular composition formulated, the manner and site of administration, and the host being treated and the condition being treated. Optimal dosages for a given set of conditions can be determined by those skilled in the art using conventional dosage determination tests. For oral administration, for example, the dose that can be used is from about 0.01 to about 1000 mg/kg of body weight, preferably from about 0.1 to about 500 mg/kg of body weight, and even more preferably from about 1 to about 500 mg/kg of body weight with repeated courses of treatment at appropriate intervals. For intravenous administration, a dose of up to 5 grams per day can be used. Intravenous administration may occur intermittently throughout the day or continuously over a 24 hour period.

The terms "cytochrome P450 inhibitory amount" and "cytochrome P450 enzyme activity inhibitory amount" as used herein refer to the amount of a compound required to reduce the activity of cytochrome P450 enzymes or certain cytochrome P450 enzyme isoforms in the presence of such a compound. Whether a particular compound reduces the activity of the cytochrome P450 enzyme, and the amount of such compound required to do so, can be determined by methods known to those skilled in the art and by the methods described herein.

The protein functions required for the replication and transcription of the coronavirus are encoded by the so-called "replicase" gene. Two overlapping polyproteins are translated from this gene and are extensively processed by viral proteases. The C-proximal region is processed at eleven conserved cross-domain junctions by a major or "3C-like" coronavirus protease. The name "3C-like" protease comes from a certain similarity between the coronavirus enzyme and the well-known picornavirus 3C proteases. These include substrate preferences, the use of cysteine as the nucleophile at the active site in catalysis, and similarities in their putative common polypeptide folds. Comparison of the amino acid sequence of the SARS-CoV-2-associated coronavirus 3C-like protease with the amino acid sequence of other known coronaviruses such as SARS-CoV shows that the amino acid sequences share approximately 96% overall homology.

The substrate amino acids at the protease cleavage site are numbered from N to the C-terminus as follows: -P3-P2-P1-P1 ' -P2 ' -P3 ' , with the cleavage occurring between residues P1 and P1' (Schechter & Berger, 1967). Substrate specificity is largely determined by the P2, P1 and P1' positions. Cleavage site specificity for the major coronavirus protease is highly conserved with a requirement for glutamine at P1 and a small amino acid at P1' [ Journal of General Virology , 83, pp. 595-599 (2002)].

The compounds of the present invention can be prepared according to the methods set forth in Reaction Schemes 1-3 below.

The following schemes further illustrate and exemplify a compound of the present invention and methods for preparing such compounds. It should be understood that the scope of the present invention is in no way limited by the scope of the following examples and preparations. In the following examples, molecules with a single chiral center may exist as a single enantiomer or racemic mixture. These molecules with two or more chiral centers may exist as a single enantiomer, a racemic or other mixture of two enantiomers, or as various mixtures of diastereomers. Such enantiomers, racemates and diastereomers may be prepared and/or resolved by methods known to those skilled in the art. One of skill in the art will appreciate that certain synthetic manipulations may epimerize or racemize the stereocenter, and synthesis conditions may be chosen to promote or inhibit such epimerization or racemization.

Scheme 1 illustrates the synthesis sequence to obtain compounds of formula I as shown, where the N-BOC methyl ester of formula 1 (WO 2005/113580) is converted into a primary amide of formula 3 (N-BOC is N- tert -butoxycarbonyl). This can be done directly, for example by treatment with ammonia (NH ₃ ) in a sealed vessel, in a solvent such as methanol or ethanol, for example optionally in the presence of additives such as calcium chloride (CaCl ₂ ) or magnesium dimethoxide, Mg(OMe) ₂ .

Scheme 1

The conversion of a compound of formula 1 to a compound of formula 3 can also be carried out by preconversion to a carboxylic acid of formula 2 (WO 2005/113580). In this case, the compound of formula 2 can be converted to a compound of formula 3 using methods well known to those skilled in the art. For example, a compound of Formula 2 can be treated with a reagent such as O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU), isobutyl chloroformate, 1-[3-(dimethylamino) hydrochloride )propyl]-3-ethylcarbodiimide ( EDCI ) and hydroxybenzotriazole (HOBt) or 1,1'-carbonyldiimidazole (CDI), optionally in the presence of a base such as N,N-diisopropylethylamine (DIEA), 4-methylmorpholine (NMM) or triethylamine (TEA), followed by treatment with NH ₃ introduced as a gas or solution in a solvent compatible with the reaction, or an NH ₃ salt such as ammonium acetate or ammonium chloride in the presence of a base such as N,N-diisopropylethylamine, 4-methylmorpholine or triethylamine. suitable solvents include, but are not limited to, dichloromethane (CH ₂ Cl ₂ ), N,N-dimethylformamide (DMF), tetrahydrofuran (THF), or acetonitrile (CH ₃ CN).

The compound of formula 3 may be deprotected from the N -protection to give the amine of formula 4 using methods well known to one of skill in the art for such deprotections. Acidic reagents are often used, such as hydrochloride, methanesulfonic acid, or trifluoroacetic acid, usually in a solvent compatible with the reaction, such as CH ₂ Cl ₂ , 1,4-dioxane, 1,2-dichloroethane, or CH ₃ CN. One skilled in the art will appreciate that a compound of formula 4 is often prepared as an acid addition salt. The compound of formula 4 can then be converted to a compound of formula 6 by treating the compound with the N -protected amino acid of formula 5 under suitable conditions. such methods are well known to the person skilled in the art, and in general standard peptide coupling conditions can be chosen.

The compound of formula 6 may be deprotected from the N -protection to give the amine of formula 7 using methods well known to those skilled in the art for such deprotections. Acidic reagents are often used, such as hydrochloride, methanesulfonic acid, or trifluoroacetic acid, usually in a solvent compatible with the reaction, such as CH ₂ Cl ₂ , 1,4-dioxane, 1,2-dichloroethane, or CH ₃ CN. One skilled in the art will appreciate that a compound of Formula 7 is often prepared as an acid addition salt. The compound of formula 7 can then be converted to a compound of formula 9 by treatment with a carboxylic acid of formula 8 under suitable conditions. Such methods are well known to the person skilled in the art. For example, if X=chlorine atom, the carboxylic acid is known as an acid chloride, and the reaction is carried out in the presence of a base to consume the hydrohalide HX obtained as a by-product of the reaction. Examples of suitable bases include, but are not limited to, tertiary amines such as 4-methylmorpholine, 2,6-dimethylpyridine, or N,N-diisopropylethylamine, or inorganic bases such as magnesium oxide (MgO), sodium carbonate (Na ₂ CO ₃ ) or potassium bicarbonate (KHCO ₃ ). suitable solvents include, but are not limited to, CH ₂ Cl ₂ , DMF, THF, or CH ₃ CN. If X=OH, a reagent or combinations of reagents is usually used to promote the reaction of the carboxylic acid 8 . One skilled in the art may choose to use, for example, a carbodiimide reagent such as 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI) or N, N' -dicyclohexylcarbodiimide (DCC), optionally in the presence of an auxiliary a nucleophile such as hydroxybenzotriazole (HOBt) or 2-hydroxypyridine-N-oxide (HOPO). Then, if X=OH, the person skilled in the art can choose to use reagents that are suitable for the formation of mixed carboxylic/carboxylic anhydrides, such as CDI, isobutyl, or ethyl chloroformate, often in the presence of a base, such as described above. Suitable solvents include, but are not limited to, CH ₂ Cl ₂ , THF, or CH ₃ CN. Another approach, commonly used by those skilled in the art when X=OH, is to treat the carboxylic acid of formula 8 with a carboxylic acid chloride, such as Me ₃ CCOCl, for example, in the presence of a base, such as described above, to give a mixed carboxylic anhydride of formula R ₃ C(O)O(O)CCMe ₃ . Suitable solvents include, but are not limited to, CH ₂ Cl ₂ , THF, or CH ₃ CN. In many cases, it is possible to use the symmetrical anhydride of the desired carboxylic acid of formula 8 to carry out the reaction, optionally in the presence of a base such as described above, in which case X=O(O)CR ₃ and therefore the carboxylic acid of formula 8 is R ₃ C(O )O(O)CR ₃ . Suitable solvents include, but are not limited to, CH ₂ Cl ₂ , THF, or CH ₃ CN.

The compound of formula 9 can be converted to the compound of formula I by treatment under dehydration conditions well known to those skilled in the art. Often this dehydration step can be performed using an excess of trifluoroacetic anhydride or phosphorus oxychloride, usually in the presence of a base such as pyridine, N,N-diisopropylethylamine, 4-methylmorpholine or triethylamine.

The person skilled in the art will be aware that the N-BOC protected amino acids of Formula 5 are known in the chemical literature, are commercially available, and can be prepared from the corresponding known and commercially available amino acids by one of skill in the art using well known N-protected amino acid synthesis procedures. . Similarly, one of skill in the art will appreciate that carboxylic acid compounds of formula 8 may be known in the chemical literature and/or commercially available and/or may be prepared by published methods or by analogy with published methods.

One skilled in the art will appreciate that the bond formation steps in Scheme 1 may be performed in a different order with appropriate considerations, such as as shown in Scheme 2.

Scheme 2

In Scheme 2, a compound of formula 3 is converted to a compound of formula 10 by treatment under dehydration conditions well known to those skilled in the art. Often this dehydration step is carried out using an excess of trifluoroacetic anhydride or phosphorus oxychloride, usually in the presence of a base such as pyridine, N,N-diisopropylethylamine, 4-methylmorpholine or triethylamine. The compound of formula 10 is deprotected to form the amine of formula 11 using methods well known to those skilled in the art to carry out such deprotections. Often acidic reagents are used, such as hydrochloride, methanesulfonic acid, or trifluoroacetic acid, usually in a solvent compatible with the reaction, such as CH ₂ Cl ₂ , 1,4-dioxane, 1,2-dichloroethane, or CH ₃ CN. One skilled in the art will appreciate that a compound of formula 11 is often prepared as an acid addition salt. The compound of formula 11 can then be converted to a compound of formula I by treatment with a compound of formula 12 under suitable conditions. Such methods are well known to the person skilled in the art, and in general standard peptide coupling conditions can be chosen. Compounds of formula 12 are exceptionally well known in the chemical literature, and one skilled in the art may choose to prepare any given compound of formula 12 using methods similar to those described in the chemical literature.

One skilled in the art will appreciate that the bonding steps in Schemes 1 and 2 may be performed in a different order, taking into account relevant factors, such as shown in Scheme 3.

Scheme 3

In Scheme 3, a compound of formula 4 can then be converted to a compound of formula 9 by treatment with a compound of formula 12 under suitable conditions. Such methods are well known to those skilled in the art and generally standard peptide binding conditions can be selected. Compounds of formula 12 are exceptionally well known in the chemical literature, and one skilled in the art may choose to prepare any given compound of formula 12 using methods analogous to those described in the chemical literature. The compound of formula 9 is then converted to a compound of formula I by treatment under dehydration conditions well known to those skilled in the art. Often this dehydration step can be performed using an excess of trifluoroacetic anhydride or phosphorus oxychloride, usually in the presence of a base such as pyridine, N,N-diisopropylethylamine, 4-methylmorpholine or triethylamine.

One skilled in the art will appreciate that further permutations of the bonding steps and manipulation of functional groups in Schemes 1, 2, and 3 may be applied, taking into account appropriate factors. Such permutations in the choice of the order of steps are well known in the chemical literature, and if desired, a person skilled in the art may refer to the chemical literature for further guidance. One skilled in the art will appreciate that other choices of protecting groups and reagents can be made to effect the various transformations.

EXAMPLES

Experimental methods

The following illustrates the synthesis of various compounds of the present invention. Additional compounds within the scope of this invention may be prepared using the methods illustrated in these examples, either alone or in combination with methods generally known in the art. All starting materials in these Preparations and Examples are either commercially available or can be prepared by methods known in the art or described herein.

All reactions are carried out with continuous stirring under nitrogen or argon, unless otherwise indicated. If necessary, the reaction devices are dried under dynamic vacuum using a heat gun and anhydrous solvents (Sure-Seal™ products from Aldrich Chemical Company, Milwaukee, Wisconsin or DriSolv™ products from EMD Chemicals, Gibbstown, NJ). In some cases, commercial solvents are passed through columns packed with 4Å molecular sieves until the following quality control standards for water are achieved: a) <100 ppm. for dichloromethane, toluene, N,N -dimethylformamide and tetrahydrofuran; b) <180 ppm for methanol, ethanol, 1,4-dioxane and diisopropylamine. For very sensitive reactions, the solvents are further treated with sodium metal, calcium hydride or molecular sieves and distilled immediately before use. Other commercial solvents and reagents were used without further purification. For reference synthesis procedures in other examples or methods, the reaction conditions (reaction time and temperature) may vary. Typically, products are vacuum dried before being used in further reactions or submitted for biological testing.

If indicated, reactions are microwave heated using Biotage Initiator or Personal Chemistry Emrys Optimizer microwaves. The progress of the reaction is followed by thin layer chromatography (TLC), liquid chromatography-mass spectrometry (LCMS), high performance liquid chromatography (HPLC) and/or gas chromatography-mass spectrometry (GCMS). TLC is performed on pre-coated silica gel plates with a fluorescence indicator (excitation wavelength 254 nm) and visualized under UV light and/or with I ₂ , KMnO ₄ , CoCl ₂ , phosphomolybdic acid and/or cerium and ammonium molybdate stains. LCMS data were obtained on an Agilent 1100 Series instrument with a Leap Technologies autosampler, Gemini C18 columns, acetonitrile/water gradients, and modifiers trifluoroacetic acid, formic acid, or ammonium hydroxide. The column eluate is analyzed using a Waters ZQ mass spectrometer scanning in both positive and negative ion modes, from 100 to 1200 Da. Other similar tools are also used. HPLC data are typically obtained on an Agilent 1100 Series instrument using the indicated columns, acetonitrile/water gradients, and trifluoroacetic acid or ammonium hydroxide modifiers. GCMS data were obtained using a Hewlett Packard 6890 oven with HP 6890 injector, HP-1 column (12 m x 0.2 mm x 0.33 µm) and helium carrier gas. The sample is analyzed on a mass selective detector HP 5973 scanning from 50 to 550 Da using electron ionization. Purification is performed by medium efficiency liquid chromatography (MELC) using Isco CombiFlash Companion, AnaLogix IntelliFlash 280, Biotage SP1, or Biotage Isolera One instruments and prepackaged Isco RediSep or Biotage Snap silica cartridges. Chiral purification is performed by chiral supercritical liquid chromatography (SLC), usually using Berger or Thar instruments; columns such as ChiralPAK-AD, -AS, -IC, Chiralcel-OD, or -OJ columns; and mixtures of CO ₂ with methanol, ethanol, 2-propanol or acetonitrile alone or modified with trifluoroacetic acid or propane-2-amine. UV detection is used to trigger the collection of fractions. For reference synthesis procedures in other examples or methods, purifications may vary: in general, solvents and solvent ratios used for eluents/gradients are chosen to provide suitable R _f or retention times.

Mass spectrometry data is obtained from LCMS analysis. Mass spectrometry (MS) is carried out using atmospheric pressure chemical ionization (APCI), electrospray ionization (ESI), electron impact ionization (EI), or electron scattering ionization (ES) sources. Proton nuclear magnetic spectroscopy ( ¹ H NMR) chemical shifts are given in ppm downfield from tetramethylsilane and recorded on 300, 400, 500, or 600 MHz Varian, Bruker, or Jeol spectrometers. Chemical shifts are expressed in parts per million (ppm, δ) with reference to the residual peaks of the deuterated solvent (chloroform, 7.26 ppm; CD ₂ HOD, 3.31 ppm; acetonitrile- d ₂ , 1.94 ppm; dimethylsulfoxide- d ₅ , 2.50 ppm; DHO, 4.79 ppm). Peak shapes are described as follows: s, singlet; e, doublet; t, triplet; square, quartet; quint, quintet; m, multiplet; shs, wide singlet; view, visible. SLC analytical data are usually obtained on a Berger analytical instrument as described above. Optical rotation data were obtained on a PerkinElmer model 343 polarimeter using a 1 dm cell. Microanalyses are performed by Quantitative Technologies Inc. and the data obtained is within 0.4% of the calculated values.

Unless otherwise noted, chemical reactions are carried out at room temperature (about 23 degrees Celsius).

Unless otherwise indicated, all reagents are commercially available and used without further purification, or prepared by methods known in the literature.

The terms "concentrated", "evaporated" and "concentrated in vacuo " refer to the removal of the solvent under reduced pressure on a rotary evaporator with a bath temperature of less than 60°C. The abbreviations "min" and "h" stand for "minutes" and "hours" respectively. The term "TLC" refers to thin layer chromatography, "room temperature or ambient temperature" means a temperature of 18 to 25°C, "GCMS" refers to gas chromatography-mass spectrometry, "LCMS" refers to liquid chromatography-mass spectrometry, "UPLC" refers to ultra high performance liquid chromatography, "HPLC" refers to high performance liquid chromatography, and "SLC" refers to supercritical liquid chromatography.

Hydrogenation can be performed in a Parr shaker under hydrogen gas pressure or in a Thales-nano H-Cube hydrogenation flow apparatus at full hydrogen and a flow rate of 1-2 ml/min at a given temperature.

The retention times of HPLC, UPLC, LCMS, GCMS and SLC are measured using the methods indicated in the procedures.

In some examples, a chiral separation has been performed to separate the enantiomers or diastereomers of certain compounds of the invention (in some examples, the separated enantiomers are designated ENT-1 and ENT-2 according to their elution order; similarly, the separated diastereomers are designated DIAST-1 and DIAST- 2 according to their elution order). In some examples, the optical rotation of an enantiomer is measured using a polarimeter. According to the observed rotation data (or specific rotation data), the clockwise rotation enantiomer is designated as the (+)-enantiomer, and the counter-clockwise rotation enantiomer is designated as the (-)-enantiomer. Racemic compounds are indicated either by the absence of a stereochemistry drawn or described, or by the presence of a (+/-) adjacent structure; in this latter case, said stereochemistry is only one of the two enantiomers that make up the racemic mixture.

The compounds and intermediates described below have been named using the naming convention provided by ACD/ChemSketch 2019.1.1, File Version C05H41, Build 110712 (Advanced Chemistry Development, Inc., Toronto, Ontario, Canada). The naming convention provided by ACD/ChemSketch 2019.1.1 is well known to those skilled in the art and the naming convention provided by ACD/ChemSketch 2019.1.1 is generally considered to be consistent with the IUPAC (International Union for Pure and Applied Chemistry) recommendations in the rules of the Nomenclature of Organic Chemistry and the CAS Index.

Example 1

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 1 )

Stage 1. Synthesis of methyl (1R.2S,fiveS)-3-[N-(tert-butoxycarbonyl)-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate (C1).

0°C solutionN-(tert-butoxycarbonyl)-L-valine (69.7 g, 321 mmol) in a mixture of acetonitrile andN, N-dimethylformamide (10:1, 1.10 L) is treated with hexafluorophosphateO-(7-azabenzotriazol-1-yl)-N, N, N', N'-tetramethyluronium (GATU; 122 g, 321 mmol), thenN, N-diisopropylethylamine (127 ml, 729 mmol). then the reaction mixture was stirred for 5 minutes, methyl (1R.2S,fiveS)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate, hydrochloride (60.0 g, 292 mmol) and stirring continued at 0° C. for 1 hour. The reaction mixture was then diluted with aqueous citric acid (1 N; 50 ml) and water (100 ml), stirred for 2 minutes, and concentratedin a vacuum up to about half of the original volume. The resulting mixture was partitioned between ethyl acetate and water, and the aqueous layer was extracted three times with ethyl acetate. The combined organic layers are then washed three times with water and once with saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated.in a vacuum. The residue is stirred in a minimum amount of ethyl acetate and then filtered; the insoluble product is washed with ethyl acetate until it turns white. The combined filtrates are concentrated under reduced pressure and then subjected to silica gel chromatography (eluent: 1:1 ethyl acetate/heptane) to giveC1 as a yellow oil. Yield: 109 g, quantitative. HCMSm/z 369.3 [M+H]⁺.^oneH NMR (400 MHz, chloroform-d) δ 5.08 (d,J=9.6 Hz, 1H), 4.45 (s, 1H), 4.11 (dd,J=9.7, 7.8 Hz, 1H), 3.95 (d, half AB quartet,J=10.1 Hz, 1H), 3.86 (dd, ABX system component,J=10.2, 4.8 Hz, 1H), 3.74 (s, 3H), 2.04-1.93 (m, 1H), 1.50-1.41 (m, 2H), 1, 40 (s, 9H), 1.04 (s, 3H), 1.00 (d,J=6.8 Hz, 3H), 0.95 (d,J=6.8 Hz, 3H), 0.93 (s, 3H).

Stage 2. Synthesis of methyl (1R.2S,fiveS)-6,6-dimethyl-3-L-valyl-3-azabicyclo[3.1.0]hexane-2-caroxylate, hydrochloride (C2).

A solution of hydrochloride in 1,4-dioxane (4 M; 15 ml, 60 mmol) is added to a 0° C. solution of C1 (1.00 g, 2.71 mmol) in ethyl acetate (50 ml). The reaction mixture was stirred at 0°C for 2 hours, then additional hydrochloride in 1,4-dioxane solution (4 M; 10 ml, 40 mmol) was added and stirring was continued at 0°C for 3 hours, then at room temperature within 1 hour. The reaction mixture was then treated with a solution of hydrochloride in 1,4-dioxane (4 M; 10 ml, 40 mmol) and methanol (15 ml) and stirred overnight at room temperature. Concentration in vacuo gives C2 as a gum; this product is used for the next chemical treatment without further purification, and the reaction is evaluated as quantitative. LCMS m/z 269.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.24 (br s, 3H), 4.27 (s, 1H), 3.81-3.61 (m, 3H), 3.67 (s, 3H), 2.21-2.06 (m, 1H), 1.63-1.55 (m, 1H), 1.49 (d, component of the AB quartet, J = 7.6 Hz, 1H), 1 .09-0.88 (m, 12H).

Stage 3. Synthesis of methyl (1R.2S,fiveS)-6,6-dimethyl-3-[N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-caroxylate (C3).

Triethylamine (1.55 ml, 11.1 mmol) is added to a 0°C solution of C2 (1.0 g, 3.3 mmol) in dichloromethane (37 ml), then trifluoroacetic anhydride (0.57 ml, 4 .0 mmol) for 30 minutes. The reaction mixture was stirred at 0°C for 30 minutes, then it was diluted with dichloromethane (100 ml), washed successively with 10% aqueous potassium bisulfate (50 ml) and saturated aqueous sodium chloride (30 ml), dried over sodium sulfate, filtered , and concentrated in vacuo to give C3 as a light yellow oil. Yield: 1.2 g, 3.3 mmol, quantitative. LCMS m/z 365.2 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 7.04 (bd, J =8.8 Hz, 1H), 4.54 (dd, J =8.9, 6.3 Hz, 1H), 4, 46 (s, 1H), 3.91 (dd, J =10.1, 5.0 Hz, 1H), 3.80-3.73 (m, 1H), 3.76 (s, 3H), 2 .25-2.13 (m, 1H), 1.55-1.47 (m, 2H), 1.09-1.03 (m, 6H), 0.94 (d, J = 6.8 Hz , 3H), 0.92 (s, 3H).

Step 4. Synthesis of (1 R ,2 S ,5 S )-6,6-dimethyl-3-[ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxylic acids ( C4 ).

Concentrated hydrochloric acid (0.57 ml, 6.6 mmol) is added to a solution of C3 (1.25 g, 3.43 mmol) in a mixture of acetic acid (40.8 ml) and water (8.2 ml). The reaction mixture is heated at 55°C for 3 days, then it is divided between water (50 ml) and ethyl acetate (100 ml). The aqueous layer was extracted with ethyl acetate (2 x 50 ml) and the combined organic layers were washed with saturated aqueous sodium chloride (50 ml), dried over sodium sulfate, filtered, and concentrated in vacuo to give C4 as a white foam. Yield: 1.00 g, 2.85 mmol, 83%. LCMS m/z 351.2 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ), characteristic peaks: δ 4.56-4.44 (m, 2H), 2.24-2.12 (m, 1H), [1.66 (d, component AB quartet, J = 7.5 Hz) and 1.59-1.47 (m), total 2H], 1.10-1.01 (m, 6H), 0.96-0.91 (m, 6H ).

Step 5. Synthesis of tert -butyl {( 2S )-1-amino-1-oxo-3-[( 3S )-2-oxopyrrolidin-3-yl]propan-2-yl}carbamate ( C5 ).

A solution of ammonia in methanol (7.0 M; 150 ml, 1.0 mol) was added to a 0° C. solution of methyl N- ( tert -butoxycarbonyl)-3-[( 3S )-2-oxopyrrolidin-3-yl]- L-alaninate (5.00 g, 17.5 mmol) in methanol (25 ml). The reaction mixture is then stirred at room temperature for 3 days, it is concentrated in vacuo ; the residue was diluted and re-concentrated sequentially with a mixture of ethyl acetate and heptane (1:1, 4×50 ml) then heptane (50 ml) to give C5 as a solid (5.27 g, quantified) which contained residual solvent. Part of this product is used in the next step. LCMS m/z 216.2 [(M − 2-methylprop-1-ene)+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.16-3.96 (m, 1H), 3.40-3.27 (m, 2H, suggested; partially covered by solvent peak), 2.55- 2.42 (m, 1H), 2.35 (dddd, J =12.2, 8.6, 6.8, 3.3 Hz, 1H), 2.03 (ddd, J =14.0, 11 .0, 4.4 Hz, 1H), 1.93-1.81 (m, 1H), 1.74 (ddd, J = 14.2, 10.1, 4.3 Hz, 1H), 1, 45 (s, 9H).

Step 6. Synthesis of tert -butyl {(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}carbamate ( C6 ).

2,6-dimethylpyridine (2 ml, 17 mmol) and trifluoroacetic anhydride (0.94 ml, 6.6 mmol) are added to a 0°C solution of C5 (from the previous step; 1.0 g, ≤3.3 mmol) in dichloromethane (12 ml). The reaction mixture is stirred at room temperature for 1.5 hours, where it is treated with hydrochloric acid (1 M; 30 ml) and dichloromethane (60 ml). The organic layer was washed successively with saturated aqueous sodium chloride (30 ml) and saturated aqueous sodium bicarbonate (30 ml), dried over sodium sulfate, and concentrated in vacuo ; chromatography on silica gel (Gradient: 40%-100% ethyl acetate in heptane) gives C6 as a solid. Yield: 737 mg, 2.91 mmol, 88% over 2 steps. LCMS m/z 254.3 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.72 (dd, J =9.3, 6.8 Hz, 1H), 3.39-3.27 (m, 2H, suggested; partially covered by peak solvent), 2.57-2.46 (m, 1H), 2.36 (dddd, J = 12.2, 8.6, 6.3, 3.4 Hz, 1H), 2.21 (ddd, J = 13.8, 9.3, 5.6 Hz, 1H), 1.92-1.79 (m, 2H), 1.47 (s, 9H).

Step 7. Synthesis of ( 2S )-2-amino-3-[( 3S )-2-oxopyrrolidin-3-yl]propanenitrile, methanesulfonate ( C7 ).

To a solution of C6 (317 mg, 1.25 mmol) in 1,1,1,3,3,3-hexafluoropropan-2-ol (3 mL) was added methanesulfonic acid (81.2 μL, 1.25 mmol). The reaction mixture was then stirred at room temperature for 45 minutes, concentrated in vacuo , then placed several times in a mixture of solvents and re-concentrated: acetonitrile and ethyl acetate (1:1, 2x10 ml) then ethyl acetate and heptane (1:1, 2 ×10 ml). The resulting C7 was obtained as a glass (423 mg) which was free of the nitrile epimer through ¹ H and ¹³ C NMR analysis. A portion of this product was used in the following reactions without further purification. LCMS m/z 154.2 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.78 (t, J =7.3 Hz, 1H), 3.42-3.36 (m, 2H), 2.82-2.68 ( m, 1H), 2.70 (s, 3H), 2.50-2.39 (m, 1H), 2.20 (t, J =7.3 Hz, 1H), 2.07-1.80 (m, 2H).

Stage 8. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl 3-[ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 1 ).

A mixture of C7 (from the previous step; 98.8 mg, ≤0.292 mmol) and C4 (100 mg, 0.285 mmol) in acetonitrile (1.5 ml) was cooled to 0°C. Add O- (7-Azabenzotriazol-1-yl) -N,N,N',N'- tetramethyluronium hexafluorophosphate (GATU, 97%; 112 mg, 0.286 mmol) followed by a solution of 4-methylmorpholine (94.0 µl, 0.855 mmol) in acetonitrile (0.5 ml), and the reaction mixture was stirred at 0°C for approximately 2 hours. Saturated aqueous sodium bicarbonate solution (30 ml) was then added to the 0° C. reaction mixture, followed by dichloromethane (50 ml) and the organic layer was washed with hydrochloric acid (1 M; 30 ml). The combined aqueous layers were extracted with dichloromethane (60 ml), then the combined organic layers were dried over sodium sulfate, concentrated in vacuo , and subjected to silica gel chromatography (Gradient: 0% - 20% methanol in ethyl acetate). The resulting product was assessed by NMR and LCMS as being contaminated with the product epimer and then purified by reverse phase HPLC (Column: Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v). v/v Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% to 95% B over 8.54 minutes then 95% B over 1.46 minutes; Flow rate: 25 ml/minute) to give (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl }-6,6-dimethyl-3-[ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 1 ). Yield: 14.6 mg, 30.1 µmol, 11%. LCMS m/z 486.5 [M+H] ⁺ . Retention time: 2.33 minutes (Analytical conditions. Column: Waters Atlantis C18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% - 95% B over 4.0 minutes, then 95% B over 1.0 minutes Flow rate: 2 ml/ minute).

Alternative C4 synthesis

( 1R , 2S , 5S )-6,6-dimethyl-3-[ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxylic acid ( C4 )

Stage 1. Synthesis of (1 R ,2 S ,5 S )-3-[ N- ( tert -butoxycarbonyl)-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2 -carboxylic acid ( C8 ).

An aqueous solution of lithium hydroxide (2.0 M; 436 ml, 872 mmol) is added to a solution of C1 (107 g, 290 mmol) in tetrahydrofuran (730 ml). The resulting mixture was then stirred at room temperature for approximately 2 hours, diluted with water and ethyl acetate, then treated with 1 M aqueous sodium hydroxide. The aqueous layer was washed with ethyl acetate and the combined organic layers were extracted three times with 1M aqueous sodium hydroxide until LCMS analysis indicated that C8 had been completely removed from the organic layer. Acidification of the combined aqueous layers to pH 2 is carried out by adding concentrated hydrochloric acid, then the mixture is extracted three times with ethyl acetate. The combined organic layers are washed with saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated; trituration of the residue with heptane gives C8 as a white solid. Yield: 92.8 g, 262 mmol, 90%. LCMS m/z 355.3 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.32 (s, 1H), 4.05 (d, half AB quartet, J =10.5 Hz, 1H), 4.01 (d, J = 9.0 Hz, 1H), 3.88 (dd, ABX system component, J = 10.4, 5.3 Hz, 1H), 2.03-1.91 (m, 1H), 1.57 (dd , component of the ABX system, J = 7.5, 5.2 Hz, 1H), 1.50 (d, half of the AB quartet, J = 7.5 Hz, 1H), 1.41 (s, 9H), 1, 08 (s, 3H), 0.99 (d, J =6.8 Hz, 3H), 0.97-0.94 (m, 6H).

Step 2. Synthesis of (1R, 2S , 5S )-6,6-dimethyl-3-L-valyl-3-azabicyclo[ 3,1,0 ]hexane-2-carboxylic acid, hydrochloride ( C9 ).

To a solution of C8 (82.8 g, 234 mmol) in dichloromethane (230 ml) was added a solution of hydrochloride in 1,4-dioxane (4.0 M; 409 ml, 1.64 mol). The reaction mixture was stirred overnight at room temperature, then concentrated in vacuo to give C9 as a white foam. This product is used directly in the next step. LCMS m/z 255.3 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.42 (s, 1H), 4.05 (d, J = 4.8 Hz, 1H), 3.89 (dd, component of the ABX system, J = 10.5, 5.2 Hz, 1H), 3.74 (d, half AB quartet, J = 10.5 Hz, 1H), 2.36-2.25 (m, 1H), 1.62 (dd , component of the ABX system, J = 7.5, 5.1 Hz, 1H), 1.57 (d, half of the AB quartet, J = 7.6 Hz, 1H), 1.16 (d, J = 7.0 Hz, 3H), 1.10 (s, 3H), 1.04 (d, J =6.9 Hz, 3H), 1.01 (s, 3H).

Step 3. Synthesis of (1 R ,2 S ,5 S )-6,6-dimethyl-3-[ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxylic acids ( C4 ).

A solution of C9 (from the previous step; ≤234 mmol) in methanol (230 ml) was cooled to 0°C, treated with triethylamine (66.7 ml, 479 mmol) and stirred for 5 minutes, then ethyl trifluoroacetate (36.1 ml, 303 mmol). The reaction mixture is then stirred at room temperature for 90 minutes, it is concentrated in vacuo . The residue was diluted with water, 1 M aqueous sodium hydroxide and ethyl acetate, and the resulting organic layer was extracted twice with 1 M aqueous sodium hydroxide. The combined aqueous layers were acidified to pH 2 with 1 M hydrochloric acid, then extracted three times with ethyl acetate. The combined organic layers are washed with water and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo to give C4 as a white foam. Yield: 73.4 g, 210 mmol, 90% over 2 steps. LCMS m/z 351.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 12.65 (v br s, 1H), 9.82 (d, J = 7.7 Hz, 1H), 4.16 (dd, J = 9.9 , 7.9 Hz, 1H), 4.12 (s, 1H), 3.86 (d, half of the AB quartet, J = 10.4 Hz, 1H), 3.81 (dd, component of the ABX system, J = 10.5, 5.0 Hz, 1H), 2.18-2.05 (m, 1H), 1.54 (dd, ABX system component, J =7.7, 4.6 Hz, 1H), 1 .42 (d, half AB quartet, J = 7.5 Hz, 1H), 1.02 (s, 3H), 0.95 (d, J = 6.7 Hz, 3H), 0.89 (d, J = 6.6 Hz, 3H), 0.84 (s, 3H).

Alternative Synthesis of Example 1

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 1 )

Step 1. Synthesis of methyl 3-[(3S)-2-oxopyrrolidin-3-yl]-L- alaninate , methanesulfonate ( C10 ).

To a solution of methyl N- ( tert -butoxycarbonyl)-3-[(3S)-2-oxopyrrolidin-3-yl]-L- alaninate (10.1 g, 35.3 mmol) in 1,1,1,3 ,3,3-hexafluoropropan-2-ole (70 ml) methanesulfonic acid (2.30 ml, 35.4 mmol) is added. The reaction mixture was then stirred at room temperature for 70 minutes, LCMS analysis showed that the starting material had been converted to C10 : LCMS m/z 187.2 [M+H] ⁺ . The reaction mixture was concentrated in vacuo and the residue redissolved twice, then concentrated under reduced pressure in a mixture of acetonitrile and ethyl acetate (1:1, 2×20 ml). The product obtained is taken up in a mixture of acetonitrile and ethyl acetate (1:1, 30 ml), concentrated, then redissolved twice in ethyl acetate (2×40 ml) and concentrated. The residue was triturated with ethyl acetate (60 ml) to give C10 . Yield: 9.87 g, 35.0 mmol, 99%. ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.22 (dd, J =9.7, 3.6 Hz, 1H), 3.86 (s, 3H), 3.41-3.36 ( m, 2H), 2.84-2.74 (m, 1H), 2.70 (s, 3H), 2.41 (dddd, J =12.3, 8.6, 5.1, 3.6 Hz, 1H), 2.25 (ddd, J =15.1, 4.5, 3.6 Hz, 1H), 1.98 (ddd, J =15.1, 9.6, 9.6 Hz, 1H), 1.87 (dddd, J =12.6, 10.9, 9.2, 9.2 Hz, 1H).

Step 2. Synthesis of methyl N -({(1 R ,2 S ,5 S )-6,6-dimethyl-3-[ N -(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0] hexan-2-yl}carbonyl)-3-[(3S)-2-oxopyrrolidin-3-yl]-L- alaninate ( C11 ).

To a 0°C solution of C10 (2.76 g, 9.78 mmol) and C4 (3.43 g, 9.79 mmol) in acetonitrile (40 ml) was added 1-[3-(dimethylamino)propyl]-3 hydrochloride -ethylcarbodiimide (1.88 g, 9.81 mmol), then pyridine (2.37 ml, 29.3 mmol) is added dropwise. The reaction mixture was stirred at 0°C for 2.25 hours, then treated with hydrochloric acid (1 M; 50 ml) and extracted with ethyl acetate (150 ml). The organic layer was washed successively with saturated aqueous sodium chloride (50 ml), saturated aqueous sodium bicarbonate (50 ml) and saturated aqueous sodium chloride (50 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . The residue was taken up in methyl t- butyl ether (30 ml) and concentrated under reduced pressure, and the resulting glass was stirred with methyl t- butyl ether (50 ml) at room temperature overnight. After filtration, the filter cake was washed with methyl tert -butyl ether (3 x 6 ml) to give C11 as a solid which, according to ¹ H NMR analysis, contained significant residual methyl tert -butyl ether. Part of this product is used in the next step. Yield: 3.74 g; corrected for residual methyl t- butyl ether: 2.94 g, 5.67 mmol, 58%. LCMS m/z 519.5 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.55 (dd, J =12.0, 3.8 Hz, 1H), 4.34 (s, 1H), 4.29 (d, J = 9.6 Hz, 1H), 3.97 (d, J =3.1 Hz, 2H), 3.74 (s, 3H), 3.37-3.23 (m, 2H, assumed; partially covered by peak solvent), 2.73-2.62 (m, 1H), 2.32 (dddd, J =12.4, 8.8, 6.7, 2.4 Hz, 1H), 2.21-2, 10 (m, 2H), 1.86-1.74 (m, 2H), 1.60 (dt, component ABX ₂ system, J =7.7, 3.1 Hz, 1H), 1.49 (d , half of the AB quartet, J = 7.6 Hz, 1H), 1.09 (s, 3H), 1.02 (d, J = 6.9 Hz, 3H), 0.99-0.95 (m, 6H).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-[ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( C12 ).

A solution of ammonia in methanol (7.0 M; 5 ml, 40 mmol) is added to a solution of C11 (from the previous step: 205 mg, 0.311 mmol) in methanol (1 ml). The resulting solution was stirred at room temperature for 1.5 hours, then a solution of ammonia in methanol (7.0 M; 5 ml, 40 mmol) was added again and stirring was continued overnight. The reaction mixture is then treated a third time with the same amount of ammonia in methanol; after another 8 hours of reaction, it is concentrated in vacuo . The residue was diluted and re-concentrated successively with ethyl acetate (2×20 ml) and a mixture of ethyl acetate and heptane (1:1, 2×20 ml). The resulting product was dissolved in dichloromethane (50 ml), washed with hydrochloric acid (1 M; 30 ml) and saturated aqueous sodium chloride solution (30 ml), dried over sodium sulfate, filtered, and concentrated in vacuo to give C12 as a solid. Yield: 87 mg, 0.17 mmol, 55%. LCMS m/z 504.5 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 8.68 (d, J = 7.9 Hz, <1H, not completely replaced by solvent), 4.44 (ddd, J = 11.9, 7.9, 4.0 Hz, 1H), 4.37-4.26 (m, 2H), 4.01 (dd, ABX system component, J = 10.3, 5.1 Hz, 1H), 3.94 (d , half AB quartet, J = 10.2 Hz, 1H), 3.39-3.24 (m, 2H, assumed; mostly obscured by solvent peak), 2.72-2.62 (m, 1H), 2.38-2.28 (m, 1H), 2.21-2.08 (m, 2H), 1.90-1.72 (m, 2H), 1.58 (dd, ABX system component, J = 7.5, 5 Hz, 1H), 1.54 (d, half AB quartet, J = 7.7 Hz, 1H), 1.08 (s, 3H), 1.02 (d, J = 6, 7 Hz, 3H), 0.97 (d, J =6.7 Hz, 3H), 0.96 (s, 3H).

Stage 4. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl 3-[ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 1 ).

Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 88.4 mg, 0.371 mmol) was added to a solution of C12 (85.0 mg, 0.17 mmol) in dichloromethane (4.0 mL) and the reaction mixture was stirred at room temperature. After 3 hours, methyl again add N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 20 mg, 84 µmol); after 30 minutes, the reaction mixture was diluted with ethyl acetate (60 ml), washed successively with hydrochloric acid (1 M; 30 ml), saturated aqueous sodium bicarbonate solution (30 ml), and saturated aqueous sodium chloride solution (30 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . The residue is taken up in heptane and re-concentrated, followed by purification by silica gel chromatography (Gradient: 0% - 5% methanol in ethyl acetate). (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 1 ) is isolated as a solid. Yield: 35 mg, 72 µmol, 42%. LCMS m/z 486.5 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 5.04 (dd, J =10.7, 5.4 Hz, 1H), 4.28 (d, J =9.6 Hz, 1H), 4 .25 (s, 1H), 4.03-3.94 (m, 2H), 3.35-3.23 (m, 2H, inferred; mostly obscured by solvent peak), 2.72-2.62 (m, 1H), 2.37-2.26 (m, 2H), 2.19-2.08 (m, 1H), 1.93-1.75 (m, 2H), 1.64 (ddd , J = 7.6, 4.2, 2.1 Hz, 1H), 1.41 (d, J = 7.6 Hz, 1H), 1.09 (s, 3H), 1.02 (d, J = 6.8 Hz, 3H), 1.00-0.95 (m, 6H).

Example 2

N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -(pyrrolidin-1-ylacetyl)-L-leucinamide, trifluoroacetate ( 2 )

Step 1. Synthesis of benzyl 4-methyl-L-leucinate, p -toluenesulfonic acid salt ( C13 ).

Suspension of 4-methyl-L-leucine (9.5 g, 65 mmol), benzyl alcohol (28.3 g, 262 mmol), and p -toluenesulfonic acid monohydrate (14.9 g, 78.3 mmol) in toluene ( 200 ml) heated at the boil under reflux overnight; a Dean-Stark trap is used to azeotropically remove the produced water. The reaction mixture was then concentrated in vacuo , then the residue was diluted with diethyl ether (200 ml) and ethyl acetate (100 ml). The resulting suspension is stirred for 1.5 hours and filtered; the filter cake is washed with diethyl ether to give C13 as a white solid. Yield: 24.9 g, 61.1 mmol, 94%. LCMS m/z 236.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.30 (br s, 3H), 7.47 (d, J =8.1 Hz, 2H), 7.44-7.36 (m, 5H) , 7.11 (d, J = 7.8 Hz, 2H), 5.23 (AB quartet, J _{AB =} 12.3 Hz, δν _{AB =} 13.7 Hz, 2H), 4.02 (dd, J =7.3, 4.5 Hz, 1H), 2.29 (s, 3H), 1.81 (dd, J =14.5, 7.3 Hz, 1H), 1.57 (dd, J = 14.5, 4.6 Hz, 1H), 0.90 (s, 9H).

Step 2. Synthesis of benzyl 4-methyl- N- (pyrrolidin-1-ylacetyl)-L-leucinate ( C14 ).

A 0°C mixture of C13 (800 mg, 1.96 mmol) and pyrrolidin-1-ylacetic acid (254 mg, 1.97 mmol) in N,N -dimethylformamide (4 ml) was treated with O- (7-azabenzotriazole-1) hexafluorophosphate -yl) -N, N, N', N'- tetramethyluronium (GATU; 746 mg, 1.96 mmol), then with a solution of 4-methylmorpholine (0.496 ml, 4.51 mmol) in dichloromethane (1 ml). Then the reaction mixture was stirred at 0°C for 2 hours, saturated aqueous sodium bicarbonate solution (30 ml) was added at 0°C; the resulting mixture was extracted with ethyl acetate (2×60 ml) and the combined organic layers dried over sodium sulfate, filtered, and concentrated in vacuo . Purification by silica gel chromatography was carried out twice (Gradient: 0%-20% ethyl acetate in heptane followed by a second chromatographic purification using 0%-10% ethyl acetate in heptane) to give C14 as a gum (761 mg). This product is used directly in the next step. LCMS m/z 347.4 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.40-7.29 (m, 5H), 5.16 (AB quartet, J _AB= 12.2 Hz, δν _AB= 11.1 Hz, 2H ), 4.56 (dd, J =9.0, 3.1 Hz, 1H), 3.76 (AB quartet, J _AB= 15.6 Hz, δν _AB= 13.6 Hz, 2H), 3, 17-3.06 (m, 4H), 2.03-1.93 (m, 4H), 1.81 (dd, J =14.5, 3.1 Hz, 1H), 1.60 (dd, J = 14.5, 9.0 Hz, 1H), 0.95 (s, 9H).

Step 3. Synthesis of 4-methyl- N- (pyrrolidin-1-ylacetyl)-L-leucine ( C15 ).

To a solution of C14 (from the previous step; 760 mg, ≤1.96 mmol) in methanol (5 ml) was added palladium on charcoal (76.0 mg). The reaction mixture was stirred at room temperature under hydrogen (50 psi) overnight, then LCMS analysis showed conversion to C15 : LCMS m/z 257.4 [M+H] ⁺ . The reaction mixture is filtered twice through a 0.15 µm filter and the filtrate is concentrated in vacuo . The residue is dissolved twice in a mixture of ethyl acetate and heptane (1:1, 2×20 ml), then concentrated under reduced pressure; this gives C15 as a solid (646 mg). Portions of this material are used in subsequent chemical reactions without further purification. ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.46 (d, J = 8.3 Hz, 1H), 4.31 (ddd, J = 8.9, 8.6, 3.0 Hz, 1H), 3.74-3.60 (m, 2H), 3.00 br (s, 4H), 1.90-1.79 (m, 4H), 1.70 (dd, ABX system component, J =14.3, 3.0 Hz, 1H), 1.56 (dd, system component ABX, J =14.3, 9.2 Hz, 1H), 0.90 (s, 9H).

Stage 4. Synthesis of N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -(pyrrolidin-1-ylacetyl)- L-leucinamide, trifluoroacetate ( 2 ).

A mixture of C15 (from the previous step; 30 mg, ≤91 μmol) and C7 (from step 7 of example 1; 35.3 mg, ≤0.104 mmol) in N,N -dimethylformamide (1 ml) was cooled to 0°C and treated with hexafluorophosphate O - (7-azabenzotriazol-1-yl) - N, N, N', N' -tetramethyluronium (GATU, 97%; 39.9 mg, 0.102 mmol), then with a solution of 4-methylmorpholine (28.0 μl, 0.255 mmol) in dichloromethane (0.25 ml). Then the reaction mixture was stirred at 0°C for about 1.5 hours, it was diluted with saturated aqueous sodium bicarbonate (3 ml) at 0°C and extracted with dichloromethane (4×4 ml). The combined organic layers were concentrated in vacuo and purified by reverse phase HPLC (Column: Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% - 25% B over 8.5 minutes, then 25% - 95% acetonitrile over 0.5 minutes, then 95 % B for 1.0 min; Flow rate: 25 ml/min) to give N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}- 4-methyl- N ² -(pyrrolidin-1-ylacetyl)-L-leucinamide, trifluoroacetate ( 2 ) as gum. Yield: 8.1 mg, 16 µmol, 18% over 3 steps. LCMS m/z 392.6 [M+H] ⁺ . Retention time: 1.47 minutes (Analytical conditions. Column: Waters Atlantis C18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% - 95% B over 4.0 minutes, then 95% B over 1.0 minutes Flow rate: 2 ml/minute ).

Example 3

N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -(2,6-dichlorobenzoyl)-4-methyl-L-leucinamide ( 3 )

Step 1. Synthesis of 3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide, methanesulfonate ( C16 ).

To a solution of C5 (6.13 g, ≤19 mmol) in 1,1,1,3,3,3-hexafluoropropan-2-ol (40 mL) was added methanesulfonic acid (1.83 g, 19 mmol). The reaction mixture was stirred at room temperature for 1 hour, then it was concentrated in vacuo , taken up in a mixture of toluene and heptane, and concentrated once more to give a hydroscopic glass (7.47 g). A portion of this product (6.47 g) was diluted and re-concentrated sequentially with the following: a mixture of dichloromethane and ethanol (2:3, 2×50 ml); ethyl acetate and ethanol (2:3, 50 ml); ethyl acetate, heptane and dichloromethane (4:4:1, 2×50 ml). The resulting product was dissolved in a mixture of acetonitrile and water (1:1, 22 ml) and lyophilized for 2 days to obtain C16 as a glass. Yield: 3.23 g, 12.1 mmol, 73% over 2 steps. LCMS m/z 172.2 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.03 (dd, J =9.1, 4.6 Hz, 1H), 3.43-3.35 (m, 2H), 2.82- 2.72 (m, 1H), 2.71 (s, 3H), 2.49-2.38 (m, 1H), 2.12-1.96 (m, 2H), 1.94-1, 81 (m, 1H).

Step 2. Synthesis of N- ( tert -butoxycarbonyl)-4-methyl-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide ( C17 ).

0°C solution of C16 (1.34 g, 5.02 mmol) and N- ( tert -butoxycarbonyl)-4-methyl-L-leucine (1.28 g, 5.22 mmol) in N,N -dimethylformamide ( 7.0 ml) is treated with O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU, 97%; 2.04 g, 5.20 mmol), then with solution 4 -methylmorpholine (1.43 ml, 13.0 mmol) in dichloromethane (3 ml). The reaction mixture is then stirred at 0°C for 2.25 hours, it is quenched at 0°C by the addition of hydrochloric acid (1 M; 30 ml) and then diluted with dichloromethane (50 ml). The organic layer was washed with saturated aqueous sodium bicarbonate (30 ml) and the combined aqueous layers were extracted with dichloromethane (60 ml). The combined organic layers are dried over sodium sulfate, filtered, concentrated in vacuo , and suspended/concentrated with heptane (3×10 ml). Purification of the residue by chromatography on silica gel (Gradient: 0% - 20% methanol in ethyl acetate) gave C17 as a solid. Yield: 1.42 g, 3.56 mmol, 71%. LCMS m/z 399.4 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 6.83 (d, J =7.4 Hz, <1H, not completely replaced by solvent), 4.43 (dd, J =11.2, 4.2 Hz , 1H), 4.11-4.05 (m, 1H), 3.38-3.24 (m, 2H, inferred; partially covered by solvent peak), 2.52-2.41 (m, 1H), 2.40-2.30 (m, 1H), 2.13 (ddd, J =14.0, 11.2, 4.5 Hz, 1H), 1.91-1.75 (m, 2H), 1.71 (dd, ABX system component, J =14.4, 3.2 Hz, 1H), 1.51 (dd, ABX system component, J =14.4, 9.3 Hz, 1H), 1, 45 (s, 9H), 0.97 (s, 9H).

Step 3. Synthesis of 4-methyl-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide, methanesulfonate ( C18 ).

Methansulfonic acid (32.6 μl, 0.502 mmol) is added to a solution of C17 (200 mg, 0.502 mmol) in 1,1,1,3,3,3-hexafluoropropan-2-ol (1.5 ml). The reaction mixture was stirred at room temperature for 40 minutes, then concentrated in vacuo , dissolved in ethyl acetate and concentrated once more to give C18 as a solid (238 mg). Most of this product is used in the next step. LCMS m/z 299.4 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.53 (dd, J =10.3, 5.0 Hz, 1H), 3.91 (dd, J =7.6, 5.5 Hz, 1H), 3.41-3.27 (m, 2H, estimated; partially covered by solvent peak), 2.70 (s, 3H), 2.57-2.47 (m, 1H), 2.41 (dddd , J =12.0, 8.6, 7.0, 3.2 Hz, 1H), 2.15 (ddd, J =14.0, 10.3, 5.0 Hz, 1H), 2.01 (dd, J =14.4, 7.5 Hz, 1H), 1.96-1.85 (m, 1H), 1.78 (ddd, J =14.1, 9.1, 5.0 Hz , 1H), 1.59 (dd, J =14.3, 5.5 Hz, 1H), 1.01 (s, 9H).

Step 4. Synthesis of N- (2,6-dichlorobenzoyl)-4-methyl-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide ( C19 ).

A 0°C suspension of C18 (from the previous step: 234 mg, ≤0.49 mmol) in dichloromethane (2 ml) is treated with triethylamine (170 μl, 1.2 mmol), then a solution of 2,6-dichlorobenzoyl chloride (130 mg , 0.621 mmol) in dichloromethane (0.2 ml). The reaction mixture was stirred at room temperature for 1 hour, then it was diluted with dichloromethane (60 ml), then washed with hydrochloric acid (1 M; 30 ml), then with saturated aqueous sodium bicarbonate (30 ml). The organic layer is dried over sodium sulfate, filtered, concentrated in vacuo , and subjected to silica gel chromatography (Gradient: 0% - 30% methanol in ethyl acetate) to give C19 . Yield: 120 mg, 0.255 mmol, 52% over 2 steps. LCMS m/z 471.4 (dichlorine isotope pattern observed) [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 8.45 (d, J =7.9 Hz, <1H, not completely replaced by solvent), 7.45-7.35 (m, 3H), 4.59 (dd, J =7.8, 4.5 Hz, 1H), 4.52-4.44 (m, 1H), 3.37-3.24 (m, 2H, inferred; partly covered by solvent peak), 2.65-2.55 (m, 1H), 2.37 (dddd, J =12.5, 8.8, 6.6, 2.8 Hz, 1H), 2.19 (ddd, J =13 .9, 11.3, 4.5 Hz, 1H), 1.91-1.72 (m, 3H), 1.66 (dd, ABX system component, J = 14.4, 7.8 Hz, 1H ), 1.03 (s, 9H).

Step 5. Synthesis of N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -(2,6-dichlorobenzoyl)-4-methyl- L-leucinamide ( 3 ).

A solution of C19 (90 mg, 0.19 mmol) and 1 H -imidazole (33.8 mg, 0.496 mmol) in pyridine (1 ml) was cooled in an acetonitrile/dry ice bath (−35° C.). Phosphorus oxychloride (0.100 ml, 1.07 mmol) was added thereto and the reaction mixture was stirred at -30°C to -20°C. After 30 minutes, pyridine (2 ml) is added to facilitate mixing; after 1 hour, dichloromethane (2 ml) is added for the same reason. After 2 hours of reaction, phosphorus oxychloride (0.100 ml, 1.07 mmol) is added again and stirring is continued for 30 minutes at -30°C, then the reaction mixture is warmed to 0°C and stirred for another 40 minutes. It is then treated with hydrochloric acid (1 M; 30 ml) and extracted with dichloromethane (2×60 ml). The combined organic layers are dried over sodium sulfate, filtered, concentrated in vacuo , and subjected to silica gel chromatography (Gradient: 0% - 15% methanol in ethyl acetate) to give a solid (67 mg). This product is combined with the product (12 mg) from a similar reaction carried out using C19 (30 mg, 64 μmol) and taken up twice in ethyl acetate (2×3 ml), then concentrated under reduced pressure. The residue is stirred with a mixture of ethyl acetate and heptane (1:3, 4 ml) at room temperature for 40 minutes and filtered; the filter cake was washed with a mixture of ethyl acetate and heptane (1:3, 5×2 ml) to obtain N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -(2,6-dichlorobenzoyl)-4-methyl-L-leucinamide ( 3 ) as a solid. Combined yield: 70 mg, 0.15 mmol, 59%. LCMS m/z 453.3 (dichlorine isotope pattern observed) [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.45-7.34 (m, 3H), 5.05 (dd, J =10.7, 5.4 Hz, 1H), 4.56 ( dd, J = 7.0, 5.7 Hz, 1H), 3.37-3.23 (m, 2H, estimated; partially covered by solvent peak), 2.70-2.59 (m, 1H), 2 .42-2.29 (m, 2H), 1.95-1.77 (m, 3H), 1.67 (dd, ABX system component, J =14.4, 7.0 Hz, 1H), 1 .04 (s, 9H).

Example 4

N -[(2 S )-1-({(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-1 H -indole-2-carboxamide ( 4 )

Stage 1. Synthesis of methyl L-leucyl-3-[(3 S )-2-oxopyrrolidin-3-yl]-L-alaninate, hydrochloride ( C20 ).

Methyl N- ( tert -butoxycarbonyl)-L-leucyl-3-[(3S)-2-oxopyrrolidin-3-yl]-L- alaninate solution (see Prior, AM, et al., Bioorg. Med. Chem Lett. 2013 , 23 , 6317-6320; 2.0 g, 5.0 mmol) in a mixture of methanol (2 ml) and a solution of hydrochloride in ethyl acetate (4 M; 20 ml) was stirred at 25°C for 1 hour. Concentration in vacuo gives C20 as a white solid (1.92 g, quant.). ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 9.09-8.98 (m, 1H), 8.39 (br s, 3H), 7.69 (s, 1H), 4, 44-4.31 (m, 1H), 3.22-3.07 (m, 2H), 2.5-2.38 (m, 1H, estimated; partially covered by solvent peak), 2.24-2, 11 (m, 1H), 2.11-1.99 (m, 1H), 1.78-1.48 (m, 5H), 0.92 (d, J =6.5 Hz, 3H), 0 .89 (d, J = 6.5 Hz, 3H).

Step 2. Synthesis of methyl N- (4-methoxy- 1H -indole-2-carbonyl)-L-leucyl-3-[(3S)-2-oxopyrrolidin-3-yl]-L- alaninate ( C21 ).

Hexafluorophosphate O - (7-azabenzotriazol-1-yl) - N, N, N', N' -tetramethyluronium (GATU; 494 mg, 1.30 mmol) and N, N -diisopropylethylamine (388 mg, 3.00 mmol) added to a 0°C solution of C20 (from a smaller scale experiment similar to step 1; 336 mg, ≤0.840 mmol) and 4-methoxy- 1H -indole-2-carboxylic acid (159 mg, 0.832 mmol) in N,N -dimethylformamide (6 ml). The solution was stirred at 0°C for 1.5 hours, then poured into water/ice (10 ml) and extracted with ethyl acetate (3×10 ml). The combined organic layers are washed with saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (eluent: 10:1 dichloromethane/methanol) gave C21 as a yellow oil. Yield: 380 mg, 0.804 mmol, 97%. LCMS m/z 473.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 11.59-11.53 (m, 1H), 8.53 (d, J =8.0 Hz, 1H), 8.37 (d, J = 8.0 Hz, 1H), 7.65 (s, 1H), 7.37-7.33 (m, 1H), 7.09 (dd, J =8.8 Hz, 1H), 7.00 ( e, component of the AB quartet, J = 8.2 Hz, 1H), 6.50 (d, J = 7.6 Hz, 1H), 4.56-4.47 (m, 1H), 4.40-4 .31 (m, 1H), 3.88 (s, 3H), 3.62 (s, 3H), 3.18-3.05 (m, 2H), 2.41-2.29 (m, 1H ), 2.15-2.03 (m, 2H), 1.78-1.49 (m, 5H), 0.93 (d, J =6.3 Hz, 3H), 0.89 (d, J = 6.4 Hz, 3H).

Step 3. Synthesis of N- (4-methoxy- 1H -indole-2-carbonyl)-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanine ( C22 ).

To a stirred mixture of calcium chloride (0.887 g, 7.99 mmol) and sodium hydroxide (0.168 g, 4.20 mmol) in 2-propanol (7 mL) and water (3 mL) was added C21 (1.8 g, 3. 8 mmol). The reaction mixture was stirred at 20°C for 6 hours, then it was concentrated in vacuo , diluted with water (4 ml), adjusted to pH 4 with 1 M hydrochloric acid and extracted with ethyl acetate (3×10 ml). The combined organic layers are washed with saturated aqueous sodium chloride, dried over sodium sulfate, filtered and concentrated in vacuo . Chromatography on silica gel (eluent: 10:1:0.1 dichloromethane/methanol/acetic acid) gave C22 as a yellow solid. Yield: 1.76 g, 3.84 mmol, 100%. LCMS m/z 459.2 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ), characteristic peaks: δ 6.51-6.43 (m, 1H), 4.80-4.66 (m, 1H), 4.60-4.45 ( m, 1H), 3.92 (s, 3H), 3.36-3.18 (m, 2H), 2.59-2.44 (m, 1H).

Alternative Step 3 Synthesis of N- (4-methoxy- 1H -indole-2-carbonyl)-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanine ( C22 ).

A solution of C21 (20 mg, 42 μmol) in tetrahydrofuran (0.4 ml) is treated with an aqueous solution containing lithium hydroxide (14.2 mg, 0.593 mmol). The reaction mixture was then stirred at room temperature for 2.5 hours, diluted with ethyl acetate, and washed with 10% aqueous potassium bisulfate. The organic layer is then dried over sodium sulfate, filtered, and concentrated in vacuo to give C22 as a white solid. Yield: 20 mg, quantitative. LCMS m/z 459.2 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.27 (s, 1H), 7.14 (dd, ABX component, J =8.8 Hz, 1H), 7.02 (d, AB component quartet, J = 8.3 Hz, 1H), 6.50 (d, J = 7.7 Hz, 1H), 4.66 (dd, J = 9.0, 5.9 Hz, 1H), 4, 52 (dd, J = 11.7, 3.9 Hz, 1H), 3.92 (s, 3H), 3.30-3.18 (m, 2H), 2.65-2.52 (m, 1H), 2.38-2.26 (m, 1H), 2.21 (ddd, J = 14.0, 11.7, 4.1 Hz, 1H), 1.90-1.70 (m, 5H), 1.02 (d, J =6.3 Hz, 3H), 0.99 (d, J =6.3 Hz, 3H).

Step 4. Synthesis of N- (4-methoxy- 1H -indole-2-carbonyl)-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide ( C23 ).

To a 0°C solution of C22 (1.76 g, 3.84 mmol) and ammonium chloride (0.246 g, 4.60 mmol) in N,N -dimethylformamide (15 mL) was added hexafluorophosphate O- (7-azabenzotriazole-1- yl) -N,N,N',N'- tetramethyluronium (GATU; 1.90 g, 5.00 mmol) and N, N -diisopropylethylamine (1.49 g, 11.5 mmol). The reaction mixture is then stirred at 0°C for 1.5 hours, N,N -diisopropylethylamine (2.3 g, 18 mmol) is used to adjust the pH to 8. The reaction mixture is stirred for another 30 minutes, then it is poured into the mixture hydrochloric acid (1 M; 20 ml, 20 mmol) and ice. The resulting mixture is extracted with ethyl acetate (3×10 ml); the combined organic layers are washed successively with hydrochloric acid (1 M; 10 ml) and saturated aqueous sodium chloride solution (10 ml), dried over sodium sulfate, filtered, concentrated in vacuo , and purified by chromatography on silica gel (eluent: 10:1 dichloromethane/methanol ) to give C23 as a yellow solid. Yield: 1.09 g, 2.38 mmol, 62%. LCMS m/z 458.0 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 11.62-11.55 (m, 1H), 8.42 (d, J =7.9 Hz, 1H), 8.04 (d, J = 8.4 Hz, 1H), 7.60 (ws, 1H), 7.38-7.26 (m, 2H), 7.10 (dd, ABX system component, J =8.8 Hz, 1H), 7.06 (ws, 1H), 7.00 (d, AB component of the quartet, J = 8.2 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 4.54-4 .41 (m, 1H), 4.34-4.22 (m, 1H), 3.88 (s, 3H), 3.17-3.01 (m, 2H), 2.31-1.95 (m, 3H), 1.76-1.45 (m, 5H), 0.92 (d, J =6.1 Hz, 3H), 0.88 (d, J =6.3 Hz, 3H) .

Stage 5. Synthesis of N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl- 1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 4 ).

To a 0°C mixture of C23 (500 mg, 1.09 mmol) and N,N -diisopropylethylamine (565 mg, 4.37 mmol) in tetrahydrofuran (8 ml) was added ,2,4,6-trioxatrifosfinan 2,4,6-trioxide (50% w/w solution in ethyl acetate; 2.78 g, 4.37 mmol). The reaction mixture was then stirred at 50° C. for 3 hours, concentrated in vacuo , diluted with water (5 ml) and extracted with ethyl acetate (3×5 ml). The combined organic layers are washed with saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo ; purification by silica gel chromatography (eluent: 10:1 dichloromethane/methanol) then reverse phase HPLC (Column: YMC-Actus Triart C18, 50×250 mm, 7 µm; Mobile phase A: water containing 0.225% formic acid; Mobile phase B: acetonitrile Gradient: 18% - 58% B; Flow rate: 25 ml/minute) gives N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S ) -2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 4 ) as a yellow solid. Yield: 130 mg, 0.296 mmol, 27%. LCMS m/z 440.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 11.58 (br s, 1H), 8.90 (d, J =8.0 Hz, 1H), 8.47 (d, J =7.7 Hz , 1H), 7.71 (ws, 1H), 7.38-7.35 (m, 1H), 7.09 (dd, ABX system component, J = 8.8 Hz, 1H), 7.00 ( e, component of the AB quartet, J = 8.2 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 5.02-4.93 (m, 1H), 4.49-4 .40 (m, 1H), 3.88 (s, 3H), 3.19-3.05 (m, 2H), 2.41-2.29 (m, 1H), 2.20-2.06 (m, 2H), 1.85-1.62 (m, 4H), 1.58-1.47 (m, 1H), 0.94 (d, J =6.3 Hz, 3H), 0, 89 (d, J =6.3 Hz, 3H).

Alternative Synthesis of Example 4

N -[(2 S )-1-({(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-1 H -indole-2-carboxamide ( 4 )

Step 1. Synthesis of N -[(4-methoxy- 1H -indol-2-yl)carbonyl]-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide ( C23 ).

A solution of ammonia in methanol (7.0 M; 21 ml, 150 mmol) was added to a solution of C21 (500 mg, 1.06 mmol) in methanol (2.0 ml). Then the reaction mixture was stirred at room temperature for 6 hours, and a solution of ammonia in methanol (7.0 M; 7.0 ml, 49 mmol) was added again and stirring was continued overnight. Ammonia in methanol (7.0 M; 7.0 ml, 49 mmol) was added again and stirring was continued for 24 hours, followed by a final treatment with ammonia in methanol (7.0 M; 7.0 ml, 49 mmol). ). The reaction mixture is stirred for another day, then it is concentrated in vacuo . The residue is combined with the product of a similar reaction (350 mg from 512 mg isolated) using C21 (500 mg, 1.06 mmol) and the mixture is redissolved in ethyl acetate (5×10 ml) and concentrated under reduced pressure to give C23 (835 mg ). This product is used directly in the next step. LCMS m/z 458.4 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.29 (d, J = 0.9 Hz, 1H), 7.15 (dd, component of the ABX system, J = 8.8 Hz, 1H), 7 .03 (bd, AB component of the quartet, J = 8.3 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 4.59 (dd, J = 9.7, 5.0 Hz, 1H), 4.45 (dd, J =11.3, 4.2 Hz, 1H), 3.93 (s, 3H), 3.34-3.19 (m, 2H, assumed; partially closed solvent peak), 2.57-2.47 (m, 1H), 2.31 (dddd, J =12.6, 8.5, 6.8, 2.8 Hz, 1H), 2.15 (ddd , J =14.0, 11.4, 4.6 Hz, 1H), 1.88-1.67 (m, 5H), 1.02 (d, J =6.1 Hz, 3H), 0, 98 (d, J =6.1 Hz, 3H).

Stage 2. Synthesis of N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl- 1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 4 ).

A solution of C23 (from the previous step; 835 mg, ≤1.78 mmol) and 1 H -imidazole (323 mg, 4.74 mmol) in a mixture of pyridine (4 ml) and dichloromethane (4 ml) was cooled to -35°C with using an acetonitrile/dry ice bath, then phosphorus oxychloride (0.956 mL, 10.2 mmol) was added dropwise over 5 minutes. The reaction mixture is stirred at -30°C to -20°C for about 1.5 hours, then treated with hydrochloric acid (1 M; 50 ml) and stirred for 1 hour. After extraction with dichloromethane (3×60 ml), the resulting organic layers are combined, dried over sodium sulfate, filtered, and concentrated in vacuo . The residue was combined with purified 4 from another batch (75 mg, 0.17 mmol) and subjected to silica gel chromatography (Gradient: 0% - 5% methanol in ethyl acetate) to give 4 as a solid (800 mg). This product is combined with the product (80 mg) from a similar reaction carried out using C23 (161 mg, 0.352 mmol); the resulting product is stirred in diethyl ether (25 ml) for 3 days, then it is filtered. The filter cake is washed with a mixture of diethyl ether and heptane (1:1, 4×2 ml) to obtain N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )- 2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 4 ) as a solid. Combined yield: 519 mg, 1.18 mmol, approximately 50% over 2 steps. LCMS m/z 440.5 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 11.57 (d, J =2.3 Hz, 1H), 8.90 (d, J =8.1 Hz, 1H), 8.46 (d , J = 7.7 Hz, 1H), 7.70 (s, 1H), 7.37 (d, J = 2.3 Hz, 1H), 7.10 (dd, ABX system component, J = 8, 8 Hz, 1H), 7.00 (d, AB quartet component, J = 8.2 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 5.03-4.92 ( m, 1H), 4.51-4.39 (m, 1H), 3.88 (s, 3H), 3.19-3.05 (m, 2H), 2.42-2.30 (m, 1H), 2.20-2.06 (m, 2H), 1.80 (ddd, J = 13.2, 9.3, 6.7 Hz, 1H), 1.75-1.63 (m, 3H), 1.58-1.47 (m, 1H), 0.94 (d, J =6.2 Hz, 3H), 0.89 (d, J =6.2 Hz, 3H).

Examples 5 and 6

N -[(2 S )-1-({(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-3-(trifluoromethyl)-1 H -indole-2-carboxamide ( 5 ) and N -[(2 S )-1-({(1 S )-1-Cyano-2- [(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-4-methoxy-7-(trifluoromethyl)-1 H -indole-2-carboxamide ( 6 )

To a depressurized tube containing zinc(II) trifluoromethanesulfonate (98%, 2.44 mg, 7.21 µmol) is added successively a solution of 4 (0.79 mg, 1.8 µmol) in dimethyl sulfoxide (60 µl), trifluoroacetic acid (0.56 µl, 7.3 µmol) and tert -butyl hydroperoxide (70% in water; 1.25 µl, 9.03 µmol). The tube is capped and heated to 50°C overnight, then the reaction mixture is cooled and diluted with acetonitrile and 1% formic acid in water to a volume of approximately 2-3 ml. The final mixture of the solvent composition is such that the resulting mixture is clear, typically about 20%-30% acetonitrile. The whole mixture was subjected to reverse phase HPLC (Column: Phenomenex Luna C18.10×250 mm, 10 µm; Mobile phase A: 0.5% acetic acid in water; Mobile phase B: 9:1 acetonitrile/methanol; Gradient: 15% B over 5 minutes, then 15% - 70% B linear gradient over 84 minutes, then 70% - 95% B over 1 minute, then 95% B over 9 minutes; flow rate: 2 ml/min). The eluate is passed through a UV/VIS sensor and then split at approximately 15:1 between a fraction collector and an ion trap mass spectrometer. Fractions are collected every 20 seconds and those potentially containing products of interest are evaluated by UPLC-UV-HRMS before pooling. The two products elute at approximately 71 and 75 minutes. The product eluted first is 5 { N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)- 4-methyl-1-oxopentan-2-yl]-4-methoxy-3-(trifluoromethyl)-1 H -indole-2-carboxamide}, and the product eluted second is 6 { N -[(2 S )- 1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-4-methoxy -7-(trifluoromethyl)-1 H -indole-2-carboxamide}.

5 - Yield: 0.101 mg, 0.199 µmol, 11%. High resolution MS m/z 508.2171 [M+H] ⁺ ; calculated for C ₂₄ H ₂₉ F ₃ N ₅ O ₄ , 508.2172. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 12.22 (br s, 1H), 9.01 (d, J =7.6 Hz, 1H), 8.96 (d, J =7.9 Hz , 1H), 7.73 (s, 1H), 7.21 (dd, J = 8.8 Hz, 1H), 7.08 (d, J = 8.2 Hz, 1H), 6.69 (d , J = 7.8 Hz, 1H), 5.03-4.95 (m, 1H), 4.49-4.40 (m, 1H), 3.87 (s, 3H), 3.22- 3.08 (m, 2H), 2.43-2.34 (m, 1H), 2.23-2.10 (m, 2H), 1.82 (ddd, J =13.7, 9.3 , 6.8 Hz, 1H), 1.78-1.66 (m, 2H), 1.62 (ddd, J = 14.6, 9.7, 5.2 Hz, 1H), 1.49 ( ddd, J =13.8, 8.8, 5.5 Hz, 1H), 0.97-0.88 (m, 6H). Retention time: 8.43 minutes (Analytical conditions. Column: Phenomenex Kinetex XB-C18, 2.1×100 mm, 2.6 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile ; Gradient: 5% B over 0.5 minutes, then 5% - 70% B over 10.5 minutes, then 70% - 95% B over 2 minutes; Flow rate: 0.4 ml/min).

6 - Yield: 14.7 µg, 0.029 µmol, 1.6%. High resolution MS m/z 508.2178 [M+H] ⁺ ; calculated for C ₂₄ H ₂₉ F ₃ N ₅ O ₄ , 508.2172. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 11.47 (br s, 1H), 9.00 (d, J =7.9 Hz, 1H), 8.79 (d, J =7.8 Hz , 1H), 7.70 (s, 1H), 7.55 (d, J = 8.2 Hz, 1H), 7.35 (s, 1H), 6.72 (d, J = 8.3 Hz , 1H), 5.02-4.94 (m, 1H), 4.56-4.48 (m, 1H), 3.97 (s, 3H), 3.18-3.05 (m, 2H ), 2.39-2.30 (m, 1H), 2.18-2.08 (m, 2H), 1.86-1.77 (m, 1H), 1.75-1.64 (m , 3H), 1.61-1.52 (m, 1H), 0.95 (d, J =6.1 Hz, 3H), 0.90 (d, J =6.1 Hz, 3H). Retention time: 8.92 minutes (Analytical conditions identical to those used for 5 ).

Alternative Synthesis of Example 6

N -[(2 S )-1-({(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-7-(trifluoromethyl)-1 H -indole-2-carboxamide ( 6 )

Step 1. Synthesis of trifluoromethylated 4-methoxy- 1H -indole-2-carboxylic acid ( C24 ).

A mixture of 4-methoxy- 1H -indole-2-carboxylic acid (100 mg, 0.523 mmol) and zinc(II) trifluoromethanesulfinate (120 mg, 0.362 mmol) was treated with dimethyl sulfoxide (1.5 ml) followed by trifluoroacetic acid (56 μl, 0.727 mmol). After the reaction mixture was cooled to 0° C., tert- butyl hydroperoxide (70% in water; 143 μl, 1.03 mmol) was added and stirring was continued at 0° C. for 20 minutes, then at room temperature for 25 minutes. The reaction mixture is then heated at 52°C for 2 hours, then it is cooled to room temperature and treated dropwise with aqueous sodium bicarbonate until sparging stops. The resulting mixture was then partitioned between an aqueous sodium bicarbonate solution and ethyl acetate, the aqueous layer was extracted once with ethyl acetate, and the organic layers were discarded. The aqueous layer is then acidified to pH 7 with 1 M hydrochloric acid; ethyl acetate is added and the mixture is stirred until the pH is adjusted to 1 with 1 M hydrochloric acid. After stirring the biphasic mixture for 10 minutes, the organic layer was washed with saturated aqueous sodium chloride, dried over magnesium sulfate, filtered, and concentrated in vacuo . According to LCMS analysis, the residue (115 mg) contains a mixture of starting material and mono-trifluoromethylated products, as well as a small amount of di-trifluoromethylated product. The volume of this mixture is used in step 4. Yield: 115 mg, <0.4 mmol. LCMS m/z 189.8, 257.8, 325.8 (not significant) [M−H] ⁻ . ¹ H NMR (400 MHz, methanol- d ₄ ), characteristic peaks of their three main components: δ 7.07 (bd, J =8.4 Hz), 7.02 (bd, J =8.4 Hz), 6 .81 (d, J =7.8 Hz), 6.66 (d, J =7.8 Hz), 6.51 (d, J =7.7 Hz), 4.06 (s, -OMe) , 3.93 (s, -OMe), 3.92 (s, -OMe).

Step 2. Synthesis of N- ( tert -butoxycarbonyl)-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanine amide ( C25 ).

To a 0°C solution of methyl N- ( tert -butoxycarbonyl)-L-leucyl-3-[(3S)-2-oxopyrrolidin-3-yl]-L- alaninate (see Prior, AM, et al., Bioorg Med. Chem. Lett. 2013 , 23 , 6317-6320; 1.5 g, 3.8 mmol) in methanol (5 ml) was added a solution of ammonia in methanol (7 M; 43 ml, 300 mmol). After the reaction vessel is capped, the reaction mixture is stirred overnight at room temperature. A solution of ammonia in methanol (7 M; 10.7 ml, 74.9 mmol) is added again and the reaction is continued at room temperature for 3 days, then it is concentrated in vacuo . The residue is taken up twice in diethyl ether (40 ml) and concentrated under reduced pressure to give C25 as a white solid. Yield: 1.46 g, 3.80 mmol, quantitative. LCMS m/z 385.4 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 8.29-8.17 (m, 1H), 7.23 (br.s., 1H), 5.64 (br.s., 1H), 5.32 (br.s., 1H ), 5.02 (d, J = 6.1 Hz, 1H), 4.50-4.38 (m, 1H), 4.05 (ddd, J = 10.3, 6.3, 4.5 Hz, 1H), 3.44-3.32 (m, 2H), 2.51-2.35 (m, 2H), 2.16-1.98 (m, 2H), 1.97-1, 83 (m, 1H), 1.76-1.6 (m, 2H, estimated; partially covered by water peak), 1.49-1.39 (m, 1H), 1.45 (s, 9H), 0 .94 (d, J =6.4 Hz, 3H), 0.94 (d, J =6.3 Hz, 3H).

Step 3. Synthesis of L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide, methanesulfonate ( C26 ).

A solution of methanesulfonic acid (0.861 ml, 13.3 mmol) in 1,1,1,3,3,3-hexafluoropropan-2-ol (5 ml) was slowly added to a solution of C25 (5.1 g, 13 mmol) in 1 ,1,1,3,3,3-hexafluoropropan-2-ole (43 ml). After 30 minutes, LCMS analysis shows conversion to C26 : LCMS m/z 285.3 [M+H] ⁺ . The reaction mixture was concentrated in vacuo , then taken up in the following solvent mixture and re-concentrated: acetonitrile/ethyl acetate (1:1, 2×20 ml), then ethyl acetate/heptane (1:1, 2×20 ml). The resulting solid was azeotropically distilled twice with a mixture of acetonitrile and ethyl acetate, then twice with a mixture of ethyl acetate and heptane to give C26 as a white solid (6.05 g) which retained solvents by ¹ H NMR analysis. Yield: quantification. ¹ H NMR (600 MHz, methanol- d ₄ ) δ 4.50 (dd, J =10.7, 4.9 Hz, 1H), 3.91 (dd, J =8.6, 5.5 Hz, 1H), 3.39-3.28 (m, 2H, estimated; partially covered by solvent peak), 2.70 (s, 3H), 2.53-2.46 (m, 1H), 2.43-2 .36 (m, 1H), 2.14 (ddd, J =14.0, 10.7, 5.0 Hz, 1H), 1.95-1.86 (m, 1H), 1.82-1 .71 (m, 3H), 1.70-1.64 (m, 1H), 1.02 (d, J = 6.3 Hz, 3H), 1.01 (d, J = 6.1 Hz, 3H).

Step 4. Synthesis of N -{[4-methoxy-7-(trifluoromethyl)-1 H -indol-2-yl]carbonyl}-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl] -L-alanineamide ( C27 ).

A solution of C24 (from step 1; 101 mg, <0.35 mmol) and C26 (from previous step; 204 mg, ≤0.438 mmol) in acetonitrile (1.7 ml) and N,N -dimethylformamide (1 ml) was cooled to 0°C and treated with O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 163 mg, 0.429 mmol) followed by 4-methylmorpholine (0.129 mL, 1.17 mmol ). The reaction mixture was stirred at 0° C. for 40 minutes, then a 1:1 mixture of aqueous sodium bicarbonate solution and ice was slowly added until a cloudy precipitate formed. Ethyl acetate is then added and the biphasic mixture is stirred for 5 minutes. The aqueous layer is extracted once with ethyl acetate and the combined organic layers are washed with saturated aqueous sodium chloride, dried over magnesium sulfate, filtered, and concentrated in vacuo . Purification by silica gel chromatography was carried out twice (Gradient #1: 0% - 10% methanol in dichloromethane; Gradient #2: 5% - 10% methanol in dichloromethane) to give C27 . The regiochemistry of this product is confirmed by 2D NMR experiments. Yield: 19 mg, 36 µmol, approximately 10%. LCMS m/z 526.5 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.53 (brd, J =8.2 Hz, 1H), 7.41 (s, 1H), 6.68 (d, J =8.3 Hz , 1H), 4.60 (dd, J =9.5, 5.1 Hz, 1H), 4.45 (dd, J =11.4, 4.2 Hz, 1H), 4.01 (s, 3H), 3.3-3.21 (m, 2H, estimated; partially covered by solvent peak), 2.60-2.49 (m, 1H), 2.36-2.26 (m, 1H), 2 .15 (ddd, J = 14.1, 11.5, 4.6 Hz, 1H), 1.89-1.68 (m, 5H), 1.03 (d, J = 6.1 Hz, 3H ), 0.99 (d, J = 6.2 Hz, 3H).

Stage 5. Synthesis of N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl- 1-oxopentan-2-yl]-4-methoxy-7-(trifluoromethyl) -1H -indole-2-carboxamide ( 6 ).

Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 17.2 mg, 72.2 µmol) is added to a solution of C27 (19 mg, 36 µmol) in a mixture of dichloromethane (0.5 ml) and acetonitrile (0.2 ml). The reaction mixture was then stirred at room temperature for 1 hour, diluted with ethyl acetate, and washed with 1:1 aqueous sodium bicarbonate and ice. The aqueous layer was extracted with ethyl acetate, and the combined organic layers were washed with saturated aqueous sodium chloride, and passed through a solid phase extraction cartridge filled with magnesium sulfate. Concentration of the filtrate in vacuo gave a residue which was purified by reverse phase HPLC (Column: Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 25% - 65% B in 8.5 minutes, then 65% - 95% B in 0.5 minutes, then 95% B for 1.0 minutes; Flow rate: 25 ml/minute) to give N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine- 3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-4-methoxy-7-(trifluoromethyl) -1H -indole-2-carboxamide ( 6 ). Yield: 4.3 mg, 8.5 µmol, 24%. LCMS m/z 508.6 [M+H] ⁺ . Retention time: 2.83 minutes (Column: Waters Atlantis C18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v); Gradient: 5% - 95% B over 4.0 minutes, then 95% B over 1.0 minutes; Flow rate: 2 ml/minute).

Example 7

N -[(2 S )-1-({(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-3-methylimidazo[2,1- b ][1,3]thiazole-2-carboxamide ( 7 )

Stage 1. Synthesis of N ² -( tert -butoxycarbonyl)- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L- leucinamide ( C28 ).

A solution of C17 (560 mg, 1.41 mmol) and 1 H -imidazole (249 mg, 3.65 mmol) in a mixture of pyridine (3 ml) and dichloromethane (3 ml) was cooled to -35° C. using an acetonitrile/dry bath ice. Phosphorus oxychloride (0.74 ml, 7.94 mmol) is added dropwise over 4 minutes, then additional dichloromethane (2 ml) and stirring is continued at -30°C to -20°C. After 1 hour, the reaction mixture was diluted with dichloromethane (2 ml). After approximately 1.5 hours, hydrochloric acid (1 M; 30 ml) is added; the resulting mixture was stirred for 30 minutes and then extracted with dichloromethane (2×60 ml). The combined organic layers are dried over sodium sulfate, filtered, and concentrated in vacuo to give C28 as a solid. Yield: 492 mg, 1.29 mmol, 91%. LCMS m/z 381.4 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 5.03 (dd, J =10.4, 5.7 Hz, 1H), 4.09 (dd, J =8.7, 4.2 Hz, 1H), 3.39-3.25 (m, 2H, estimated; partially covered by solvent peak), 2.64-2.52 (m, 1H), 2.40-2.27 (m, 2H), 1 .97-1.78 (m, 2H), 1.70 (dd, ABX system component, J = 14.3, 4.1 Hz, 1H), 1.54 (dd, ABX system component, J = 14, 3, 8.7 Hz, 1H), 1.45 (s, 9H), 1.00 (s, 9H).

Stage 2. Synthesis of N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4- dimethyl-1-oxopentan-2-yl]-3-methylimidazo[2,1- b ][1,3]thiazole-2-carboxamide ( 7 ).

A solution of hydrochloride in 1,4-dioxane (4.0 M; 0.3 mL, 1.2 mmol) was added to a solution of C28 (100 mg, 0.263 mmol) in a mixture of acetonitrile (1.5 mL) and methanol (1.0 ml). The reaction mixture was stirred at room temperature for 30 minutes, then treated with 4-methylmorpholine (0.144 mL, 1.31 mmol). After removing the solvents in vacuo , the residue is taken up twice in a mixture of dichloromethane and heptane (1:1, 2×10 ml) and concentrated under reduced pressure. The residue is combined with 3-methylimidazo[2,1- b ][1,3]thiazole-2-carboxylic acid (47.9 mg, 0.263 mmol) in N,N -dimethylformamide (3.3 ml), cooled to 0° C, and treated with O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 99.9 mg, 0.263 mmol) followed by a solution of 4-methylmorpholine (72 µl, 0.655 mmol ) in dichloromethane (0.2 ml). The reaction mixture is then stirred at 0°C for approximately 2 hours, it is treated at 0°C with hydrochloric acid (1 M; 30 ml), and the resulting mixture is extracted with dichloromethane (2×60 ml). The aqueous layer was then basified to pH 9 with saturated aqueous sodium bicarbonate, then extracted with dichloromethane (3 x 60 ml). The combined organic layers are washed with saturated aqueous ammonium chloride (50 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . ¹ H NMR analysis of this product indicates the presence of a slight epimer, presumably arising from partial racemization at the center bearing the nitrile. The main product is isolated using silica gel chromatography (Gradient: 0% - 20% methanol in ethyl acetate) to give N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-3-methylimidazo[2,1- b ][1,3]thiazole-2-carboxamide ( 7 ) as a solid. Yield: 56 mg, 0.13 mmol, 49%. LCMS m/z 445.4 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.73 (d, J =1.6 Hz, 1H), 7.37 (d, J =1.6 Hz, 1H), 5.04 (dd , J = 10.3, 5.9 Hz, 1H), 4.53 (dd, J = 7.8, 5.0 Hz, 1H), 3.36-3.24 (m, 2H; assumed; partially closed by solvent peak), 2.70 (s, 3H), 2.67-2.57 (m, 1H), 2.38-2.27 (m, 2H), 1.93 (ddd, J =14, 0, 9.4, 6.0 Hz, 1H), 1.88-1.78 (m, 3H), 1.03 (s, 9H).

Examples 8 and 9

N-{1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ²-[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide, DIAST-1 (eight) andN-{1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ²-[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide, DIAST-2 (nine)

Step 1. Synthesis of N- {1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide ( C29 ).

To a solution of C28 (114 mg, 0.300 mmol) in a mixture of acetonitrile (1 ml) and methanol (1 ml) was added a solution of hydrochloride in 1,4-dioxane (4 M; 0.4 ml, 1.6 mmol). The reaction mixture was stirred at room temperature for 30 minutes, then 4-methylmorpholine (0.165 ml, 1.50 mmol) was added to adjust the pH to 7-8. The solvents are then removed in vacuo , the residue is taken up twice in ethyl acetate/heptane (1:1, 2 x 10 mL) and concentrated under reduced pressure to give C29 as a solid (269 mg); according to ¹ H NMR analysis, it consists of a mixture of epimers, presumably located on the center bearing the nitrile, in a ratio of 2-3 to 1. Part of this product is used in the next step. LCMS m/z 281.3 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ), characteristic peaks: δ [5.11 (dd, J =8.8, 7.3 Hz, fundamental) and 5.01 (dd, J =6.5, 6.5 Hz, minor), 1H total], [2.75-2.65 (m, minor) and 2.64-2.54 (m, major), 1H total], 2.48-2.38 (m, 1H), 2.30-2.20 (m, 1H), 2.06-1.83 (m, 3H), 1.64 (dd, J =14.1, 4.8 Hz, 1H ), [1.04 (s, major), 1.01 (s, minor), total 9H].

Stage 2. SynthesisN-{1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ²-[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide, DIAST-1 (eight) andN-{1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-N ²-[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide, DIAST-2 (nine).

To a 0°C solution of C29 (from the previous step; 83.4 mg, ≤93 µmol) and cyclohexyl(methoxy)acetic acid (17.2 mg, 99.9 µmol) in N,N -dimethylformamide (1 ml) was added hexafluorophosphate O - (7-azabenzotriazol-1-yl) - N, N, N', N' -tetramethyluronium (GATU; 38.0 mg, 0.100 mmol), then a solution of 4-methylmorpholine (30.8 μl, 0.280 mmol) in dichloromethane (0.2 ml). Then the reaction mixture was stirred at 0°C for about 2 hours, it was diluted with saturated aqueous sodium bicarbonate (3 ml) at 0°C, and extracted with dichloromethane (4×4 ml). The combined organic layers are concentrated in vacuo ; according to LCMS analysis, the residue consists of two components, presumably corresponding to two epimers on the center bearing the nitrile. These diastereomers are separated by reverse phase HPLC (Column: Waters XBridge C18, 19×100 mm, 5 µm; Mobile phase A: water; Mobile phase B: acetonitrile; Gradient: 5% - 95% B over 8.54 minutes, then 95 % B for 1.46 minutes Flow rate: 25 ml/minute). The first diastereomer eluted is designated as 8 ( N- {1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[cyclohexyl(methoxy)acetyl]-4-methyl-L- leucinamide, DIAST-1), and the second diastereomer eluted as 9 ( N- {1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -N ² -[cyclohexyl(methoxy)acetyl] -4-methyl-L-leucinamide, DIAST-2).

8 - Yield: 12.8 mg, 29.4 µmol, 32% over 2 steps. LCMS m/z 435.6 [M+H] ⁺ . Retention time: 2.63 minutes (Analytical conditions. Column: Waters Atlantis C18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% to 95% B over 4.0 minutes, then 95% B over 1.0 minutes Flow rate: 2 ml/ minute).

9 - Yield: 10 mg, 23.0 µmol, 25% over 2 steps. LCMS m/z 435.6 [M+H] ⁺ . Retention time: 2.72 minutes (Analytical conditions identical to those used for 8 ).

Example 10

N -[(2 S )-1-({(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-1 H -indole-2-carboxamide ( 10 )

Step 1. Synthesis of N -[(4-methoxy- 1H -indol-2-yl)carbonyl]-4-methyl-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L -alanineamide ( C30 ).

O- ( _ 7-azabenzotriazol-1-yl)- N, N, N', N' -tetramethyluronium (GATU; 175 mg, 0.460 mmol), then a solution of 4-methylmorpholine (0.127 ml, 1.16 mmol) in acetonitrile (0.2 ml). The reaction mixture was stirred at 0°C for 2.5 hours, then it was diluted with saturated aqueous sodium bicarbonate (30 ml) at 0°C, then extracted with dichloromethane (50 ml). The organic layer was washed with hydrochloric acid (1 M; 30 ml) and the aqueous layers were extracted with dichloromethane (60 ml). Then the combined organic layers are dried over sodium sulfate, filtered, and concentrated in vacuo , the residue is purified by silica gel chromatography (Gradient: 0% - 30% methanol in ethyl acetate) to give C30 as a solid. Yield: 148 mg, 0.314 mmol, 68% over 2 steps. LCMS m/z 472.4 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.25 (d, J =0.9 Hz, 1H), 7.15 (dd, J =8.8 Hz, 1H), 7.03 (br , component of the AB quartet, J = 8.3 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 4.65 (dd, J = 9.2, 3.4 Hz, 1H) , 4.44 (dd, J =11.2, 4.2 Hz, 1H), 3.93 (s, 3H), 3.29-3.15 (m, 2H), 2.54-2.44 (m, 1H), 2.29 (dddd, J =12.6, 8.6, 7.0, 2.7 Hz, 1H), 2.14 (ddd, J =14.0, 11.2, 4.6 Hz, 1H), 1.89 (dd, ABX system component, J = 14.5, 3.4 Hz, 1H), 1.85-1.74 (m, 3H), 1.02 (s , 9H).

Stage 2. Synthesis of N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4- dimethyl-1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 10 ).

A solution of C30 (143 mg, 0.303 mmol) and 1H -imidazole (53.7 mg, 0.789 mmol) in a mixture of pyridine (1 ml) and dichloromethane (1 ml) was cooled in an acetonitrile/dry ice bath (−35° C.). Phosphorus oxychloride (0.159 ml, 1.71 mmol) is added dropwise over 5 minutes and the reaction mixture is stirred at -30°C to -20°C for 2 hours, then it is treated with hydrochloric acid (1 M; 30 ml ), stirred for 20 minutes and extracted with dichloromethane (2×60 ml). The combined organic layers are dried over sodium sulfate, filtered and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 10% methanol in ethyl acetate) gives N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidine-3 -yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 10 ) as a solid. Yield: 68 mg, 0.15 mmol, 50%. LCMS m/z 454.5 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.24 (d, J =0.9 Hz, 1H), 7.14 (dd, J =8.8 Hz, 1H), 7.02 (br , component of the AB quartet, J = 8.3 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 5.03 (dd, J = 10.1, 6.0 Hz, 1H) , 4.64 (dd, J = 8.6, 4.3 Hz, 1H), 3.93 (s, 3H), 3.30-3.17 (m, 2H), 2.63-2.52 (m, 1H), 2.37-2.21 (m, 2H), 1.95-1.74 (m, 4H), 1.03 (s, 9H).

Example 11

N ² -[(4-Bromo-1-ethyl-3-methyl-1 H -pyrazol-5-yl)carbonyl]- N -{(1 S )-1-cyano-2-[(3 S )-2 -oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide ( 11 )

To 0°C suspension of C18 (43.4 mg, ≤0.10 mmol) and 4-bromo-1-ethyl-3-methyl- 1H -pyrazole-5-carboxylic acid (23.3 mg, 0.100 mmol) in acetonitrile (1.0 ml) add O - (7-azabenzotriazol-1-yl) - N, N, N', N' -tetramethyluronium hexafluorophosphate (GATU; 38.0 mg, 0.100 mmol), then a solution of 4-methylmorpholine ( 30 µl, 0.27 mmol) in acetonitrile (0.2 ml). The reaction mixture is then stirred at 0° C. for about 80 minutes, methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 71.5 mg, 0.300 mmol) are added and stirring is continued. After approximately 2.75 hours, methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 71.5 mg, 0.300 mmol) is added again and the reaction is carried out for 1.5 hours, then it is treated with saturated aqueous sodium bicarbonate (3 ml) at 0°C and extracted with dichloromethane (2×8 ml). The combined organic layers were concentrated in vacuo , then dissolved in acetonitrile (4 ml) and concentrated again using a Genevac evaporator to give the crude product (138 mg). A portion of this product (80 mg) was purified by reverse phase HPLC (Column: Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% - 95% B over 8.54 minutes then 95% B over 1.46 minutes Flow rate: 25 ml/ minute) to give N ² -[(4-bromo-1-ethyl-3-methyl-1 H -pyrazol-5-yl)carbonyl]- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide ( 11 ). Yield: 24.7 mg, 49.8 µmol, 86% over 2 steps. LCMS m/z 495.5 (bromine isotope pattern observed) [M+H] ⁺ . Retention time: 2.48 minutes (Analytical conditions. Column: Waters Atlantis C18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% - 95% B over 4.0 minutes, then 95% B over 1.0 minutes Flow rate: 2 ml/ minute).

Example 12

N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[(3,3-difluorocyclobutyl)acetyl]-4-methyl-L -leucinamide ( 12 )

Step 1. Synthesis of 4-methyl-L-leucyl-3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide, hydrochloride ( C18, HCl salt ).

A solution of hydrochloride in 1,4-dioxane (4 M; 1.7 mL, 6.8 mmol) was added to a solution of C17 (260 mg, 0.652 mmol) in acetonitrile (3 mL). The reaction mixture was stirred at room temperature for 1.5 hours, then it was concentrated in vacuo , then redissolved in a mixture of dichloromethane and heptane (1:1, 3×10 ml) and re-concentrated to give C18, HCl salt (242 mg ) in the form of glass. Part of this product is used in the next step. LCMS m/z 299.3 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.53 (dd, J =10.3, 5.0 Hz, 1H), 3.91 (dd, J =7.5, 5.4 Hz, 1H), 3.41-3.26 (m, 2H, estimated; partially covered by solvent peak), 2.57-2.47 (m, 1H), 2.41 (dddd, J =12.0, 8, 7, 7.0, 3.1 Hz, 1H), 2.15 (ddd, J =13.9, 10.3, 4.9 Hz, 1H), 2.05-1.97 (m, 1H) , 1.97-1.85 (m, 1H), 1.78 (ddd, J =14.1, 9.1, 5.0 Hz, 1H), 1.60 (dd, J =14.3, 5.4 Hz, 1H), 1.01 (s, 9H).

Stage 2. Synthesis of N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[(3,3-difluorocyclobutyl)acetyl]-4 -methyl-L-leucinamide ( 12 ).

A suspension of C18, HCl salt (from the previous step; 37.2 mg, ≤0.100 mmol) and (3,3-difluorocyclobutyl)acetic acid (15.8 mg, 0.105 mmol) in tetrahydrofuran (1.0 ml) was treated with trioxide 2, 2,4,6-tripropyl-1,3,5,2,4,6-trioxatrifosfinan 4,6-trioxide (50 wt% solution in ethyl acetate; 65.5 µl, 0.110 mmol) and 4-methylmorpholine (27, 5 µl, 0.250 mmol). Then the reaction mixture was stirred at room temperature overnight, it was heated at 50°C for 4.5 hours, then 2,4,6-trioxide 2,4,6-trioxide 2,4,6-tripropyl-1,3,5,2 was added again ,4,6-trioxatrifosfinan (50% by weight solution in ethyl acetate; 2.2 equivalents) and 4-methylmorpholine (5 equivalents). The reaction mixture was then stirred at 50°C for 3 additional days, treated with saturated aqueous sodium bicarbonate (3 ml) and extracted with dichloromethane (4×4 ml). The combined organic layers were concentrated in vacuo and purified by reverse phase HPLC (Column: Waters XBridge C18, 19×100 mm, 5 µm; Mobile phase A: water; Mobile phase B: acetonitrile; Gradient: 20% - 40% B over 8 .5 minutes, then 40%-95% B over 0.5 minutes, then 95% B over 1.0 minutes; Flow rate: 25 ml/minute) to give N -{( 1S )-1-cyano -2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[(3,3-difluorocyclobutyl)acetyl]-4-methyl-L-leucinamide ( 12 ). Yield: 10.1 mg, 24.5 µmol, 24% over 2 steps. LCMS m/z 413.5 [M+H] ⁺ . Retention time: 1.96 minutes (Analytical conditions. Column: Waters Atlantis C18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5% - 95% B over 4.0 minutes, then 95% B over 1.0 minutes Flow rate: 2 ml/ minute).

Example 13

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 )

Step 1. Synthesis of methyl ( 1R , 2S , 5S )-3-[ N- ( tert -butoxycarbonyl)-3-methyl-L-valyl]-6,6-dimethyl-3-azabicyclo[3,1, 0]hexane-2-carboxylate ( C31 ).

O- (7-Azabenzotriazol-1-yl) -N,N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 7.92 g, 20.8 mmol) is added to a 0°C mixture of N- ( tert -butoxycarbonyl) -3-methyl-L-valine (4.38 g, 18.9 mmol) and methyl (1 R ,2 S ,5 S )-6,6-dimethyl-3-azabicyclo[3.1.0]hexane- 2-caroxylate, hydrochloride (3.9 g, 19 mmol) in N,N -dimethylformamide (95 ml). The reaction mixture is then stirred for 5 minutes, N,N -diisopropylethylamine (8.25 ml, 47.4 mmol) is added; stirring is continued at 0° C. for 2 hours, then an aqueous solution of citric acid (1 N, 20 ml) and water (40 ml) is added. The resulting mixture was stirred for 2 minutes and then diluted with ethyl acetate (250 ml). The organic layer was washed with water (3×150 ml) and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . Purification by chromatography on silica gel (Gradient: 0% - 100% ethyl acetate in heptane) gave C31 as an oil. Yield: 3.60 g, 9.41 mmol, 50%. ¹ H NMR (400 MHz, methanol- d ₄ ) δ 6.42 (d, J = 9.7 Hz, <1H; not completely replaced by solvent), 4.35 (s, 1H), 4.21 (d, J = 9.7 Hz, 1H), 4.02 (d, half AB quartet, J = 10.4 Hz, 1H), 3.91 (dd, component of the ABX system, J = 10.3, 5.3 Hz, 1H), 3.73 (s, 3H), 1.57 (dd, component of the ABX system, J = 7.5, 5.1 Hz, 1H), 1.47 (d, half of the AB quartet, J = 7, 5 Hz, 1H), 1.41 (s, 9H), 1.07 (s, 3H), 1.02 (s, 9H), 0.93 (s, 3H).

Step 2. Synthesis of ( 1R , 2S , 5S )-3-[ N- ( tert -butoxycarbonyl)-3-methyl-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0 ]hexane-2-carboxylic acid ( C32 ).

An aqueous solution of lithium hydroxide (1.0 M; 14.7 mmol, 14.7 ml) was added dropwise to a 0°C solution of C31 (3.60 g, 9.41 mmol) in a mixture of tetrahydrofuran and methanol (1:1, 30 ml). The reaction mixture is then stirred at 0° C. for 1 hour, warmed to room temperature and stirred for 1 hour, then LCMS analysis shows conversion to C32 : LCMS m/z 367.3 [M−H] ⁻ . Adjustment to pH 3 is carried out by adding 1 M hydrochloric acid, after which the mixture is diluted with water (30 ml). The aqueous layer was extracted with ethyl acetate (2 x 75 ml) and the combined organic layers were dried over sodium sulfate, filtered, and concentrated under reduced pressure to give C32 as an off-white solid. Yield: 3.10 g, 8.41 mmol, 89%. ¹ H NMR (400 MHz, methanol- d ₄ ) δ 6.39 (d, J = 9.7 Hz, approx. 0.5H; not completely replaced by solvent), 4.33 (s, 1H), [4.21 ( d, J=9.6 Hz) and 4.21 (s), total 1H], 4.01 (d, half AB quartet, J =10.5 Hz, 1H), 3.91 (dd, component of the ABX system , J = 10.4, 5.2 Hz, 1H), 1.56 (dd, component of the ABX system, J = 7.5, 5.0 Hz, 1H), 1.50 (d, half of the AB quartet, J =7.6 Hz, 1H), 1.42 (s, 9H), 1.07 (s, 3H), 1.02 (s, 9H), 0.93 (s, 3H).

Stage 3. Synthesis of tert -butyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(1 S )-1-cyano-2-[(3 S )-2- oxopyrrolidin-3-yl]ethyl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3,3-dimethyl-1-oxobutan-2-yl}carbamate ( C33 ).

0°C mixture of C7 (31.9 mg, ≤94 µmol) and C32 (34 mg, 92 µmol) in acetonitrile (1 ml) is treated with O- (7-azabenzotriazol-1-yl) -N, N,N' hexafluorophosphate ,N'- tetramethyluronium (GATU, 97%; 36.2 mg, 92.3 µmol) followed by a solution of 4-methylmorpholine (25 µl, 0.23 mmol) in acetonitrile (0.25 ml). Then the reaction mixture was stirred at 0°C for approximately 1 hour, it was diluted with saturated aqueous sodium bicarbonate (3 ml) at 0°C and extracted with dichloromethane (4×4 ml). The combined organic layers were concentrated in vacuo to give C33 as a gum (48 mg). Most of this product is used in the next step. LCMS m/z 504.6 [M+H] ⁺ .

Stage 4. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ).

Prepare a stock solution of methanesulfonic acid (60 µl) in 1,1,1,3,3,3-hexafluoropropan-2-ol (940 µl). To a solution of C33 (from the previous step; 47 mg, ≤90 µmol) in 1,1,1,3,3,3-hexafluoropropan-2-ol (1 mL) was added a portion of the stock methanesulfonic acid solution (0.1 mL; 100 µmol). The reaction mixture was then stirred at room temperature for 1 hour, concentrated in vacuo , then taken up in the following solvent mixture and re-concentrated: a mixture of acetonitrile and ethyl acetate (1:1, 2×10 ml), and then a mixture of ethyl acetate and heptane (1 :1, 2×10 ml). The residue was dissolved in dichloromethane (1 ml) and treated with 4-methylmorpholine (30.8 μl, 0.280 mmol) followed by trifluoroacetic anhydride (0.143 ml, 1.01 mmol). The reaction mixture was stirred at room temperature for 40 minutes, then treated with 4-methylmorpholine (30.8 μl, 0.280 mmol) followed by trifluoroacetic anhydride (0.143 ml, 1.01 mmol); 30 minutes later, 4-methylmorpholine (30.8 μl, 0.280 mmol) was added again, followed by trifluoroacetic anhydride (0.143 ml, 1.01 mmol). After another 15 minutes of stirring, the reaction mixture is treated with hydrochloric acid (1 M; 3 ml), and the resulting mixture is extracted with dichloromethane (3×4 ml); the combined organic layers were concentrated in vacuo and purified using reverse phase HPLC (Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 20% - 60% B over 8.5 minutes, then 60% - 95% B over 0.5 minutes, then 95% B for 1 minute; Flow rate: 25 ml/minute) to give (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine -3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ) . Yield: 7.5 mg, 15 µmol, 17% over 2 steps. LCMS m/z 500.5 [M+H] ⁺ . Retention time: 2.66 minutes (Analytical conditions. Column: Waters Atlantis dC18, 4.6×50 mm, 5 µm; Mobile phase A: 0.05% trifluoroacetic acid in water (v/v); Mobile phase B : 0.05% trifluoroacetic acid in acetonitrile (v/v) Gradient: 5.0% - 95% B over 4.0 minutes, then 95% B over 1.0 minutes Flow rate: 2 ml /minute).

Alternative Synthesis of Example 13, Methyl tert -butyl ether solvate; creation 13, methyl tert -butyl ether solvate, Solid form 2

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate (13, methyl tert -butyl ether solvate), Solid form 2

Step 1. Synthesis of tert -butyl {( 2S )-1-amino-1-oxo-3-[( 3S )-2-oxopyrrolidin-3-yl]propan-2-yl}carbamate ( C5 ).

This experiment is carried out in 2 parallel batches. A solution of ammonia in methanol (7 M; 2.4 L, 17 mol) was added to methyl N- ( tert -butoxycarbonyl)-3-[(3S)-2-oxopyrrolidin-3-yl]-L- alaninate (600 g , 2.10 mol) and the reaction mixture was stirred at 25°C for 40 hours. Concentration in vacuo and pooling of 2 batches gives C5 as a yellow solid. Combined yield: 1.10 kg, 4.05 mol, 96%. ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 7.63 (br s, 1H), 7.29 (br br, 1H), 7.01 (br br, 1H), 6.89 (d, J =8, 5 Hz, 1H), 3.96-3.85 (m, 1H), 3.22-3.06 (m, 2H, estimated; partially covered by water peak), 2.28-2.08 (m, 2H ), 1.89 (ddd, J =14.6, 10.8, 4.0 Hz, 1H), 1.74-1.60 (m, 1H), 1.56-1.43 (m, 1H ), 1.36 (s, 9H).

Step 2. Synthesis of 3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide, hydrochloride ( C16, HCl salt ).

This experiment is carried out in 3 parallel batches. To a 0°C solution of C5 (840 g, 3.10 mol) in dichloromethane (2.0 L) was added a solution of hydrochloride in 1,4-dioxane (4 M; 2 L, 8 mol). The reaction mixture is stirred at 25°C for 2 hours, then it is concentrated in vacuo ; combining 3 batches gives C16, the HCl salt as a white solid. Combined yield: 1.20 kg, 5.78 mol, 62%. MS m/z 172.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.52-8.35 (br s, 3H), 8.12 (s, 1H), 7.95 (s, 1H), 7.57 (s, 1H), 3.88-3.76 (m, 1H), 3.24-3.10 (m, 2H), 2.59-2.5 (m, 1H, inferred; partially covered by solvent peak), 2 .35-2.24 (m, 1H), 2.01 (ddd, J =14.9, 9.2, 6.1 Hz, 1H), 1.80-1.68 (m, 2H).

Sample C16, HCl salts triturated in 2-propanol for 1.5 hours, then collected by filtration and washed with 2-propanol. The combined solid is dried overnight under high vacuum to provide a sample for X-ray powder diffraction analysis. Powder x-ray diffraction pattern for this product is given in figure 10; characteristic peaks are listed in Table Q.

Acquisition of X-ray powder diffraction data

Powder x-ray diffraction analysis was performed using a Bruker AXS D4 Endeavor diffractometer equipped with a Cu radiation source. The divergence slit is set to 0.6 mm while the secondary optics uses variable slits. Diffracted radiation is recorded by the PSD-Lynx Eye sensor. The voltage and current of the X-ray tube are set to 40 kV and 40 mA, respectively. Data is collected in a Theta-2Theta goniometer at a Cu wavelength of 3.0 to 40.0 degrees 2-Theta using a step size of 0.020 degrees and a step time of 0.3 seconds. Samples are prepared by placing them in a silicon low background sample holder and rotating during collection.

Powder x-ray diffraction analysis is carried out using a Bruker AXS D8 Advance diffractometer equipped with a Cu radiation source. Diffracted radiation is recorded by a LYNXEYE_EX sensor with motorized slits. Both primary and secondary are equipped with 2.5-inch slots. The voltage and current of the X-ray tube are set to 40 kV and 40 mA, respectively. Data is collected on a Theta-Theta goniometer with a closed-pair scan at Cu wavelength K-alpha (average) from 3.0 to 40.0 degrees 2-Theta in 0.02 degree increments, using a scan speed of 0.5 seconds per step. Samples are obtained by placing them in a silicon low background sample holder.

Data is collected with both instruments using Bruker DIFFRAC Plus software and analysis is performed with EVA DIFFRAC plus software. The PXRD data file is not processed until the peak search. Using the peak search algorithm in the EVA software, peaks selected with a threshold of 1 are used for peak preassignment. To ensure reliability, adjustments are made manually; the output of the automatic assignments is checked visually and the peak positions are adjusted to the maximum peak. Usually choose peaks with a relative intensity ≥3%. Typically, peaks that are not resolved or correspond to noise are not selected. The typical error associated with PXRD peak position is specified in the USP up to +/- 0.2° 2-Theta (USP-941).

Table Q. Selected X-ray Powder Diffraction Peaks for C16, HCl Salt Angle (°2 theta) Rel. intensity Angle (°2 theta) Rel. intensity 9.97 3 29.24 13 11.67 one 30.98 6 14.17 one 31.78 2 16.08 one 32.32 23 16.35 one 32.79 10 17.10 fourteen 33.10 one 17.27 3 33.50 6 18.23 24 33.70 4 19.21 4 33.90 3 20.83 twenty 35.27 3 22.20 58 36.20 3 22.97 12 36.42 6 23.35 34 36.75 6 23.79 2 36.95 7 24.62 3 37.83 3 25.10 100 38.58 2 26.85 eleven 39.44 7 28.39 fourteen 39.75 one

Alternative synthesis of 3-[( 3S )-2-oxopyrrolidin-3-yl]-L-alanineamide, hydrochloride, C16 HCl salt

An alternative preparation of Compound C16, the HCl salt is depicted in the reaction scheme below.

Methyl (S)-2-amino-3-((S)-2-oxopyrrolidin-3-yl)propanoate 4-methylbenzenesulfonate (20 g, 55.8 mmol) and magnesium sulfate (6.7 g, 55.8 mmol) at room temperature. After the reaction mixture was stirred for 7 hours at room temperature, nitrogen was bubbled into the reaction mixture for 1 hour to purge excess ammonia. The reaction mixture is then filtered through a pad of Celite® and then concentratedin a vacuum and the resulting 4-methylbenzenesulfonate (S)-2-amino-3-((S)-2-oxopyrrolidin-3-yl)propanamide was used directly in the next step without further purification. A portion of 4-methylbenzenesulfonate is added to a solution of dimethylformamide (50 ml, 647 mmol). (S)-2-amino-3-((S)-2-oxopyrrolidin-3-yl)propanamide (10 g, 25.9 mmol) and hydrochloride solution in 1,4-dioxane (4.0 M; 19, 4 ml, 77.7 mmol). After stirring for 12 hours at room temperature, the suspension is filtered and washed with dimethylformamide (15 ml, 190 mmol). The resulting solid was dried in a vacuum oven at 40° C. for 12 hours to give 3-[(3S)-2-oxopyrrolidin-3-yl]-L-alanineamide, hydrochloride,C16 HCl salts(2.7 g, 12.4 mmol) as a tan solid (48% overall yield).

Stage 3. Synthesis of methyl (1 R ,2 S ,5 S )-3-[ N- ( tert -butoxycarbonyl)-3-methyl-L-valyl]-6,6-dimethyl-3-azabicyclo[3,1, 0]hexane-2-carboxylate ( C31 ).

This experiment is carried out in 3 parallel batches. To a 0°C solution of methyl (1 R ,2 S ,5 S )-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate, hydrochloride (237 g, 1.15 mol) and N- ( tert -butoxycarbonyl)-3-methyl-L-valine (293 g, 1.27 mol) in a mixture of N, N -dimethylformamide (400 ml) and acetonitrile (3.6 L) is added hexafluorophosphate O - (7- azabenzotriazol-1-yl) -N,N,N',N'- tetramethyluronium (GATU; 481 g, 1.26 mol), then N,N -diisopropylethylamine (601 ml, 3.45 mol) was added dropwise. The reaction mixture is then heated to 25°C and stirred for 16 hours, then it is poured into a mixture of ice water (1 L) and hydrochloric acid (0.5 M; 1 L), pH approximately 5, and stirred for 6 minutes . The resulting mixture was extracted with ethyl acetate (2 L) and the organic layer was washed with water (2 L), dried over sodium sulfate, filtered, and concentrated in vacuo . The residue was purified using silica gel chromatography (Gradient: 0% - 50% ethyl acetate in petroleum ether) to give, after combining 3 batches, C31 as a colorless oil. Combined yield: 1.17 kg, 3.06 mol, 89%. LCMS m/z 383.3 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 5.10 (d, J =10.2 Hz, 1H), 4.46 (s, 1H), 4.20 (d, J =10.3 Hz, 1H), 3.98 (d, half AB quartet, J = 10.2 Hz, 1H), 3.89-3.82 (m, 1H), 3.74 (s, 3H), 1.48-1 .41 (m, 2H), 1.38 (s, 9H), 1.03 (s, 3H), 1.01 (s, 9H), 0.89 (s, 3H).

Step 4. Synthesis of ( 1R , 2S , 5S )-3-[ N- ( tert -butoxycarbonyl)-3-methyl-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0 ]hexane-2-carboxylic acid ( C32 ).

This experiment is carried out in 3 parallel batches. To a solution of C31 (668 g, 1.75 mol) in tetrahydrofuran (2.5 L) was added lithium hydroxide monohydrate (220 g, 5.24 mol) and water (500 ml). Then the reaction mixture is stirred at 25°C for 2 hours, it is concentrated in vacuo to remove most of the tetrahydrofuran; the residue is then brought to pH 2 by adding 1 M hydrochloric acid. The resulting mixture was extracted with ethyl acetate (2×500 ml) and the combined organic layers washed with saturated aqueous sodium chloride solution (500 ml), dried over sodium sulfate, filtered, and concentrated in vacuo to give C32 as a white solid (2.0 kg ) after combining 3 batches. This product is used directly in the next step. LCMS m/z 313.2 [(M − 2-methylprop-1-ene)+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 5.14 (d, J =10.2 Hz, 1H), 4.46 (s, 1H), 4.24 (d, J =10.2 Hz, 1H), 4.06 (d, half AB quartet, J = 10.5 Hz, 1H), 3.82 (dd, component of the ABX system, J = 10.5, 5.5 Hz, 1H), 1.75 (d, J =7.7 Hz, 1H), 1.49 (dd, J =7.7, 5.4 Hz, 1H), 1.40 (s, 9H), 1.06 (s, 3H) , 1.00 (s, 9H), 0.89 (s, 3H).

Stage 5. Synthesis of (1 R ,2 S ,5 S )-6,6-dimethyl-3-(3-methyl-L-valyl)-3-azabicyclo[3.1.0]hexane-2-carboxylic acid, hydrochloride ( C41 ).

This experiment is carried out in 2 parallel batches. A solution of hydrochloride in 1,4-dioxane (4 M; 4.0 L, 16 mol) is added to the solutionC32 (from the previous step; 1.00 kg, ≤2.62 mol) in dichloromethane (1.0 L) and the reaction mixture was stirred at 25° C. for 16 hours. Solvent removalin a vacuum at 50°C givesC41 as a white solid (1.8 kg) after combining 2 batches. This product is used directly in the next step.^oneH NMR (400 MHz, methanol-d ₄ ) δ 4.42 (s, 1H), 4.00 (s, 1H), 3.94 (dd, component of the ABX system,J=10.7, 5.4 Hz, 1H), 3.80 (d, half AB quartet,J=10.7 Hz, 1H), 1.62 (dd, ABX system component,J\u003d 7.7, 5.2 Hz, 1H), 1.56 (d, half AB quartet,J=7.6 Hz, 1H), 1.15 (s, 9H), 1.09 (s, 3H), 1.03 (s, 3H).

Step 6. Synthesis of (1 R ,2 S ,5 S )-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane -2-carboxylic acid ( C42 ).

This experiment is carried out in 3 parallel batches. To a 0°C solution of C41 (from the previous step; 600 g, ≤1.75 mol) in methanol (2 L) was added triethylamine (1.64 L, 11.8 mol) followed by ethyl trifluoroacetate (699 g, 4.92 mol ), then the reaction mixture is heated to 25°C, and stirred for 16 hours. It is then concentrated in vacuo at 50° C. and the residue is diluted with ethyl acetate (3 L) and adjusted to pH 3-4 with 2 M hydrochloric acid. After extracting the aqueous layer with ethyl acetate (1 L), the combined organic layers are washed with saturated aqueous sodium chloride (3 L), dried over sodium sulfate, filtered, and concentrated under reduced pressure. At this point, 3 parties are combined, treated with a mixture of petroleum ether and ethyl acetate (5:1, 3 l), and stirred at 25°C for 2 hours. Filtration gives C42 as a white solid. Combined yield: 1.90 kg, 5.21 mol, 99% over 3 steps. LCMS m/z 365.1 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 8.88 (d, J =8.8 Hz, <1H; not completely replaced), [4.60 (d, J =8.9 Hz) and 4 .59 (s), total 1H], 4.35 (s, 1H), 3.96 (dd, component of the ABX system, J = 10.5, 5.1 Hz, 1H), 3.90 (d, half AB quartet, J = 10.4 Hz, 1H), 1.58 (dd, component of the ABX system, J = 7.6, 4.9 Hz, 1H), 1.52 (d, half of the AB quartet, J = 7 .6 Hz, 1H), 1.08 (s, 12H), 0.92 (s, 3H).

Step 7. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( C43 ).

This experiment is carried out in 4 parallel batches. 2-Hydroxypyridine 1-oxide (33.9 g, 305 mmol) is added to a solution of C42 (445 g, 1.22 mol) and C16, HCl salt (256 g, 1.23 mol) in butan-2-one (2 .5 l), and the mixture is cooled to 0°C. N,N -diisopropylethylamine (638 ml, 3.66 mol) was then added followed by 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (351 g, 1.83 mol) added dropwise. The reaction mixture was stirred at 25°C for 16 hours, then it was diluted with ethyl acetate (1 L) and treated with a mixture of hydrochloric acid (1 M; 1.5 L, 1.5 mol) and saturated aqueous sodium chloride solution (1 L). The organic layer was washed with a mixture of aqueous sodium hydroxide (1 M; 1.5 L, 1.5 mol) and saturated aqueous sodium chloride (1 L), dried over sodium sulfate, filtered, and concentrated in vacuo . The combination of 4 batches gives C43 as a white solid (2.3 kg). Combined yield: 2.1 kg (corrected for residual ethyl acetate), 4.1 mol, 84%. LCMS m/z 518.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.41 (bd, J = 7.7 Hz, 1H), 8.30 (d, J = 8.8 Hz, 1H), 7.56 (s , 1H), 7.32(brs, 1H), 7.04(brs, 1H), 4.43(bd, J =7.3Hz, 1H), 4.35-4.25(m, 1H) , 4.28 (s, 1H), 3.89 (dd, J =10.3, 5.5 Hz, 1H), 3.67 (d, J =10.4 Hz, 1H), 3.17- 3.09 (m, 1H), 3.07-2.98 (m, 1H), 2.46-2.35 (m, 1H), 2.19-2.10 (m, 1H), 1, 99-1.89 (m, 1H), 1.70-1.58 (m, 1H), 1.55-1.44 (m, 2H), 1.38 (d, half AB quartet, J =7 .6 Hz, 1H), 1.01 (s, 3H), 0.98 (s, 9H), 0.84 (s, 3H).

Stage 8. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate ( 13, methyl tert - butyl ether solvate ether ), Solid form 2.

This experiment is carried out in 3 parallel batches. Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 552 g, 2.32 mol) was added to a solution of C43 (600 g, 1.16 mol) in ethyl acetate (3 L). The reaction mixture was then stirred at 25° C. for 3 hours, treated with additional methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 27.6 g, 116 mmol) and the reaction mixture stirred for 1 hour. It is then filtered; the filter cake was washed with ethyl acetate (2×500 mL) and the combined filtrates were washed successively with aqueous sodium bicarbonate (1 M; 2 L), saturated aqueous sodium chloride (2 L), hydrochloric acid (1 M; 2 L), and saturated aqueous sodium chloride solution (2 l). The organic layer is then dried over sodium sulfate, filtered and concentrated in vacuo . The residue is taken up in a mixture of ethyl acetate and methyl tert -butyl ether (1:10, 2.5 L) and heated to 50°C; after stirring for 1 hour at 50°C, it is cooled to 25°C and stirred for 2 hours. The solid is collected by filtration and 3 batches are combined in ethyl acetate (8 L) and filtered through silica gel (3.0 kg); silica gel is then washed with ethyl acetate (2×2 l). The combined eluates are then concentrated in vacuo and the residue taken up in ethyl acetate (900 ml) and methyl t- butyl ether (9 l). This mixture is heated to 50°C for 1 hour, cooled to 25°C and stirred for 2 hours. Filtration gives ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl- 3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate (13, methyl tert -butyl ether solvate) as a white solid. The powder x-ray diffraction pattern for this product, designated Solid Form 2, is given in Figure 1; characteristic peaks are listed in Table A. Combined yield: 1.41 kg, 2.82 mol, 81%. LCMS m/z 500.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.42 (d, J =8.4 Hz, 1H), 9.03 (d, J =8.6 Hz, 1H), 7.68 (s , 1H), 4.97 (ddd, J = 10.9, 8.5, 5.1 Hz, 1H), 4.41 (d, J = 8.5 Hz, 1H), 4.16 (s, 1H), 3.91 (dd, J =10.4, 5.5 Hz, 1H), 3.69 (d, J =10.4 Hz, 1H), 3.18-3.10 (m, 1H ), 3.08-2.99 (m, 1H), 2.46-2.34 (m, 1H), 2.20-2.03 (m, 2H), 1.78-1.65 (m , 2H), 1.57 (dd, J =7.6, 5.4 Hz, 1H), 1.32 (d, J =7.6 Hz, 1H), 1.03 (s, 3H), 0 .98 (s, 9H), 0.85 (s, 3H).

Acquisition of X-ray powder diffraction data

X-ray powder diffraction analysis was performed using a Bruker AXS D8 Endeavor diffractometer equipped with a Cu (average K- α ) radiation source. The discrepancy gap is set to constant illumination of 15 mm. The diffracted radiation is recorded by a PSD-Lynx Eye sensor with a PSD sensor opening of 2.99 degrees. The voltage and current of the X-ray tube are set to 40 kV and 40 mA, respectively. Data is collected on a Theta-Theta goniometer at a Cu wavelength of 3.0 to 40.0 degrees 2-Theta using a step size of 0.00998 degrees and a step time of 1.0 seconds. The anti-scatter shield is set at a fixed distance of 1.5 mm. During collection, the samples are rotated at a speed of 15/min. Samples are obtained by placing them in a silicon low background sample holder and rotating during collection. Data is collected with Bruker DIFFRAC Plus software and analysis is performed with EVA DIFFRAC Plus software. Using the peak search algorithm in the EVA software, peaks selected with a threshold value of 1 are used for peak pre-assignment. To ensure reliability, adjustments are made manually; the output of automatic deductions is checked visually and the positions of the peaks are adjusted to the maximum peak. Usually choose peaks with a relative intensity ≥3%. Peaks that are not resolved or correspond to noise are not selected. Typical error associated with USP PXRD peak position is up to +/- 0.2° 2-Theta (USP-941).

Table A. Selected X-ray powder diffraction peaks for 13, methyl tert-butyl ether solvate , Solid Form 2, from the alternative synthesis of Example 13, methyl tert-butyl ether solvate; Preparation 13, Methyl tert-butyl ether solvate, Solid form 2 Angle (°2-theta)
+/- 0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/- 0.2° 2-Theta Relative intensity (%) 7.1 78 22.7 nine 10.5 eight 22.9 10 11.3 fifteen 23.1 five 11.8 36 23.4 6 12.5 49 23.7 22 12.9 4 25.3 fourteen 14.2 34 27.3 3 15.7 10 27.9 6 16.0 24 28.3 nine 16.8 100 28.5 4 17.0 41 29.1 3 18.5 fifty 29.4 6 18.8 7 30.2 3 19.1 25 30.8 five 19.9 eleven 32.0 4 20.2 eight 33.3 7 20.8 fourteen 33.8 4 21.1 nine 35.4 7 21.4 4 36.4 6 21.7 4 38.1 3 22.2 24

Alternative synthesis of ( 1R , 2S , 5S )-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2 -carboxylic acid ( C42 ).

Second alternative synthesis of Example 13, Methyl t- butyl ether solvate; creation 13, methyl tert -butyl ether solvate, Solid form 2

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate (13, methyl tert -butyl ether solvate), Solid form 2

Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 392 g, 1.64 mol) was added to a solution of C43 (415 g, 802 mmol) in ethyl acetate (2.0 L). The reaction mixture was stirred at 25° C. for 3 hours, then methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 86.0 g, 361 mmol) was added again. The reaction mixture was then stirred for 1 hour, filtered, and the filtrate was washed successively with aqueous sodium bicarbonate (1 M; 1.5 L), saturated aqueous sodium chloride (1.5 L), hydrochloric acid (1 M; 1.5 L). 5 L) and saturated aqueous sodium chloride solution (1.5 L), dried over sodium sulfate, filtered, and concentrated in vacuo. The residue is taken up in a mixture of ethyl acetate and methyl t- butyl ether (1:10, 2.5 L) and heated to 50°C; after stirring for 1 hour at 50°C , it is cooled to 25°C and stirred for 2 hours. Collection of the solid by filtration gives ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-6.6 -dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate ( 13, methyl tert -butyl ether solvate butyl ether ) as a crystalline white solid. The powder x-ray diffraction pattern for this product, designated Solid Form 2, is given in Figure 2; characteristic peaks are listed in Table B. Yield: 338 g, 575 mmol, 72%. LCMS m/z 500.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.43 (d, J =8.4 Hz, 1H), 9.04 (d, J =8.6 Hz, 1H), 7.68 (s , 1H), 4.97 (ddd, J = 10.9, 8.5, 5.0 Hz, 1H), 4.41 (d, J = 8.5 Hz, 1H), 4.15 (s, 1H), 3.91 (dd, ABX system component, J = 10.4, 5.5 Hz, 1H), 3.69 (d, half AB quartet, J = 10.4 Hz, 1H), 3.18 -3.10 (m, 1H), 3.08-2.98 (m, 1H), 2.46-2.34 (m, 1H), 2.20-2.02 (m, 2H), 1 .77-1.65 (m, 2H), 1.57 (dd, J = 7.6, 5.4 Hz, 1H), 1.32 (d, half AB quartet, J = 7.6 Hz, 1H ), 1.02 (s, 3H), 0.98 (s, 9H), 0.85 (s, 3H); methyl t- butyl ether peaks: 3.07 (s, 3H), 1.10 (s, 9H).

A method for collecting X-ray powder diffraction data is described in Alternative Synthesis of Example 13, Methyl tert -butyl ether solvate, Step 8.

Table B. Selected X-ray powder diffraction peaks for 13, methyl tert-butyl ether solvate , Solid Form 2, from the alternative synthesis of Example 13, methyl tert-butyl ether solvate; Preparation 13, Methyl tert-butyl ether solvate, Solid form 2 Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) 7.2 66 20.0 nine 27.4 3 10.6 nine 20.3 6 28.0 6 11.4 12 20.8 6 28.4 7 11.9 32 20.9 12 29.5 4 12.6 49 21.2 7 30.3 3 13.0 4 21.5 4 30.9 five 14.3 37 21.8 3 32.1 3 15.8 eight 22.3 24 33.4 five 16.1 22 22.8 6 33.5 3 16.9 100 23.0 nine 35.5 6 17.2 46 23.2 five 36.5 3 18.6 42 23.5 6 38.2 3 18.9 6 23.8 17 19.3 23 25.4 10

Third Alternative Synthesis of Example 13

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 )

Stage 1. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate ( 13, methyl tert - butyl ether solvate ether )

0°C mixtureC42(90.5 wt%, 5.05 g, 12.5 mmol) andC16, HCl salt(98.9% wt., 3.12 g, 14.9 mmol) in acetonitrile (50 ml) is treated with 2,4,6-trioxide 2,4,6-tripropyl-1,3,5,2,4, 6-trioxatrifosfinan (50% by weight solution in acetonitrile; 17 ml, 24.3 mmol) for about 10 minutes. Then slowly add 1-methyl-1H-imidazole (4.0 ml, 50.2 mmol) for about 15 minutes, and the reaction mixture is stirred at 0°C for 3.5 hours, then it is heated to 25°C. 2,4,6-tripropyl-1,3,5,2,4,6-trioxatrifosfinan 2,4,6-trioxide (50% w/w solution in acetonitrile; 17 ml, 24.3 mmol) was added in one portion, and the reaction mixture is stirred at 45°C for 16 hours. It is cooled to 25° C. at this point, and then treated with aqueous sodium bicarbonate solution (1.14 M; 35 ml, 40 mmol) for 10 minutes. After adding ethyl acetate (25 ml) and sufficient water to dissolve the resulting solids, the organic layer is washed twice with aqueous sodium bicarbonate (1.14 M; 25 ml, 28 mmol). The organic layer is then washed with aqueous sodium chloride solution (14%, 2×20 ml), dried over sodium sulfate, filtered, and concentrated in vacuo. The residue is taken up in ethyl acetate (2.1 ml) and treated with methyltert-butyl ether (19 ml); the resulting suspension is heated with stirring at 50°C for 1 hour, cooled to 25°C for 1 hour, and maintained at 25°C for 1.5 hours. The solids are isolated by filtration, washed with methyltert-butyl ether (2 ml/g) and dried in a vacuum oven overnight at 50°C to obtain (1R.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl solvatetert-butyl ether (13, methyl solvate tert- butyl ether) as a crystalline white solid. The volume of this product progresses to the next stage. Yield: 3.71 g, 6.31 mmol, 50%.^oneH NMR (400 MHz, DMSO-d ₆) δ 9.40 (d,J=8.4 Hz, 1H), 9.02 (d,J=8.6Hz, 1H), 7.66(s, 1H), 4.97(ddd,J=10.7, 8.6, 5.1 Hz, 1H), 4.41 (d,J=8.4 Hz, 1H), 4.16 (s, 1H), 3.91 (dd, ABX system component,J=10.3, 5.5 Hz, 1H), 3.69 (d, half AB quartet,J=10.4 Hz, 1H), 3.18-3.10 (m, 1H), 3.09-2.99 (m, 1H), 2.46-2.35 (m, 1H), 2.20 -2.04 (m, 2H), 1.78-1.64 (m, 2H), 1.56 (dd,J=7.4, 5.6 Hz, 1H), 1.32 (d, half AB quartet,J=7.6 Hz, 1H), 1.03 (s, 3H), 0.98 (s, 9H), 0.85 (s, 3H); methyl peakstert-butyl ether: 3.07 (s, 3H), 1.10 (s, 9H).

Stage 2. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ).

A mixture of propan-2-ylacetate (17 ml) and heptane (17 ml) was added to13, methyl solvate tert- butyl ether(from the previous step; 3.41 g, 5.80 mmol) and stirring is carried out overnight at 20°C. Heptane (17 ml) is then added over 2 hours and the mixture is stirred overnight at room temperature. The resulting suspension is filtered and the collected solids are washed with a mixture of propan-2-ylacetate (1.36 ml) and heptane (3.73 ml) and dried at 50°C under vacuum to obtain (1R.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (13) as a crystalline solid. A portion of this batch was used as seed material for the recrystallization of Example 13; obtaining solid form 1 below. Yield: 2.73 g, 5.46 mmol, 94%.

Recrystallization of Example 13; obtaining a solid form 1

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ), Solid Form 1

Mixture13, methyl solvate tert- butyl ether(from alternative synthesis from example 13, methyl solvatetert-butyl ether; preparation 13, methyl solvatetert-butyl ether, Solid form 2; 60.1 g, 102 mmol) and propan-2-ylacetate (480 ml) heated to 60°C. Sample13 (seed product from the third alternative synthesis of example 13, Step 2; 1.2 g, 2.4 mmol) was added; after 10 minutes the seed product is still present in solid form. Heptane (360 ml) was slowly added to the stirred mixture over 12 hours. Additional heptane (360 ml) was added over 4 hours and the resulting mixture was stirred for 30 minutes. It is then cooled to 20°C at a rate of 0.1 degrees/minute, then it is stirred overnight. The solid is collected by filtration and washed with a mixture of propan-2-ylacetate (72 ml) and heptane (168 ml). Then it is then dried under vacuum at 50°C to obtain (1R.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (13) as a white crystalline solid. Powder X-ray diffraction pattern of this product, designated as Solid Form 1, is given in Figure 3; characteristic peaks are listed in Table C. Yield: 47.8 g, 95.7 mmol, 94%.

A method for collecting X-ray powder diffraction data is described in Alternative Synthesis of Example 13, Methyl tert -butyl ether solvate, Step 8.

Table C. Selected X-ray Powder Diffraction Peaks for 13 , Solid Form 1 Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) 7.6 16 18.9 eleven 24.7 eight 9.8 10 19.7 7 25.3 7 11.4 10 19.9 fourteen 27.0 3 11.9 13 20.5 36 27.2 6 12.7 100 21.0 fourteen 27.9 4 15.7 40 21.7 4 28.1 3 15.8 eighteen 22.2 23 29.5 7 17.3 10 22.5 3 32.6 6 17.8 12 23.1 6 35.7 4 18.3 55 23.6 10 37.0 3

Single Crystal X-Ray Structure Determination of Example 13, Solid Form 1

A sample of example 13 is subjected to crystallization by diffusion using ethyl acetate and hexane. The crystallization vessel is left at room temperature while the solvent evaporates; X-ray quality crystals appeared after 2.5 months. One of them is used to define the structure. The ORTEP chart for single crystal data is shown in Figure 4. Mercury software is used to calculate the powder structure based on the resolved crystal structure; comparison with the diffraction pattern from Recrystallization Example 13; obtaining solid form 1 identifies this product as Solid form 1 (see Figure 5). The peak characteristics for these calculated data are presented in Table D.

Table D. Powder diffraction data for 13, Solid Form 1, calculated from Single Crystal X-ray Structure Analysis of Example 13, Solid Form 1 Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) 7.6 22 21.0 28 30.1 4 9.8 21 21.6 fourteen 30.3 3 10.4 nine 21.7 fourteen 31.5 4 10.8 4 22.2 40 31.7 five 11.4 16 22.5 7 31.9 4 11.9 75 23.1 five 32.7 eight 12.7 89 23.6 fifteen 33.4 3 14.6 3 24.3 five 33.6 eight 15.7 100 24.8 fifteen 35.7 eight 15.9 thirty 25.4 10 36.6 3 17.4 34 26.4 3 36.6 3 17.9 24 27.0 nine 37.0 4 18.3 67 27.3 eight 37.3 4 18.9 12 27.9 3 38.3 3 19.7 fifteen 28.1 five 39.4 3 19.9 63 28.7 five 39.6 4 20.5 53 29.5 nine 20.8 nine 30.0 nine

X-ray analysis of single crystals

Data collection is carried out on a Bruker D8 Quest diffractometer at room temperature. Data collection consists of omega and phi scans.

The structure was resolved by internal phasing using the SHELX software package in the space group of the orthorhombic class P 2 ₁ 2 ₁ 2 ₁ . In the future, the structure is refined by the full-matrix least squares method. All non-hydrogen atoms are found and refined using anisotropic displacement parameters.

Hydrogen atoms located on nitrogen were found from the difference Fourier map and refined with limited distances. The remaining hydrogen atoms are placed in calculated positions and allowed to move on their carrier atoms. The final refinement includes isotropic displacement parameters for all hydrogen atoms.

Absolute structure analysis using probability methods (Hooft, 2008) is performed using PLATON (Spek). The results show that the absolute structure is assigned correctly. The method calculates that the probability that the structure is assigned correctly is 100%. Hooft's parameter is presented as -0.01 s os (estimated standard deviation) (3) and Parson's parameter is presented as -0.01 s s o (2).

The resulting R-index is 3.3%. The final difference Fourier map showed no missing or shifted electron density.

The corresponding crystal, data acquisition, and refinement information are summarized in Table E. Atomic coordinates, bond lengths, bond angles, and bias parameters are listed in Tables F-H.

Software and Links

SHELXTL , Version 5.1, Bruker AXS, 1997.

PLATON , A. L. Spek, J. Appl. Cryst. 2003 , 36 , 7-13.

MERCURY , C. F. Macrae, P. R. Edington, P. McCabe, E. Pidcock, G. P. Shields, R. Taylor, M. Towler, and J. van de Streek, J. Appl. Cryst. 2006, 39 , 453-457.

OLEX2 , OV Dolomanov, LJ Bourhis, RJ Gildea, JAK Howard, and H. Puschmann, J. Appl. Cryst. 2009, 42 , 339-341.

R. W. W. Hooft, L. H. Straver, and A. L. Spek, J. Appl. Cryst. 2008 , 41 , 96-103.

HD Flack, Acta Cryst. 1983 , A39 , 867-881.

Table E. Crystal data and structure refinements for Example 13, Solid Form 1 . Empirical formula
formula mass
Temperature
Wavelength
Crystal system
space group
Lattice constant
Volume
Z
Density (calculated)
Absorption coefficient
F (000)
Crystal size
Theta interval for data collection
Index ranges
Collected Reflections
Independent Reflections
Completeness to theta=67.679°
Absorption correction
Refinement method
Data/Restrictions/Parameters
Compliance with F ²
Finite indices R [ I>2σ(I) ]
R indices (all data)
Absolute Structure Parameter
Excitation factor
The largest diff. peaks and valleys C ₂₃ H ₃₂ F ₃ N ₅ O ₄
499.53
296(2) K
1.54178 Å
orthothrombic
P 2 ₁ 2 ₁ 2 ₁
a =9.6836(2) Å
b =15.0522(4) Å
c = 18.0272(5) Å
2627.64(11) Å ³
4
1.263 Mg/ ^m3
0.862 mm ^-1
1056
0.300×0.280×0.260 mm ³
3.826 to 80.042°
−12<= h <=12, −18<= k <=19, −22<= l <=23
79731
5628 [ _Rint = 0.0294 ]
99.3%
empirical
Least squares in the full-matrix approximation in F ²
5628/9/358
1.040
R 1=0.0326, wR 2=0.0906
R 1=0.0346, wR 2=0.0928
−0.01(3)
n/a
0.112 and -0.121 e.Å ^-3 α= 90°
β= 90°
γ =90°

Table F. Atomic coordinates (x 10 ⁴ ) and equivalent isotropic substitution parameters (Å ² ×10 ³ ) for example 13, Solid form 1 . U(equiv) is defined as one third of the trace of the orthogonalized tensor U ^ij .

Table G. Bond lengths [Å] and angles [°] for Example 13, Solid Form 1 .

Symmetric transformations are used to create equivalent atoms.

Table H. Anisotropic substitution parameters (Å ² ×10 ³ ) for Example 13, Solid Form 1 . The exponent of the anisotropic substitution coefficient takes the form: −2π ² [h ² a* ² U ¹¹ + ... + 2 hka* b* U ¹² ].

NMR analysis of a solid body of the compound from Example 13, Forms 1 and 4, is carried out on a CPMAS probe installed in a Bruker-BioSpin Avance III 500 MHz NMR spectrometer (frequency ¹ H). A rotation speed at a magic angle of 15.0 kHz is used. Form 1 spectra were collected at ambient temperature (not controlled temperature) and Form 4 spectra were collected at 15°C.

^.sup.13 C NMR spectra were obtained using a cross-polarization magic angle spinning experiment (CPMAS) with a decoupled proton. During the collection of spectra, a phase-modulated proton decoupling field of 80-100 kHz is used. The cross polarization contact time is set to 2 ms and the recirculation delay is set to 3.5 seconds for Form 1 and Form 4. The number of scans is adjusted to obtain an adequate signal to noise ratio. The ¹³ C chemical shift scale was compared using the ¹³ C CPMAS experiment on a crystalline adamantane external standard, setting its upfield resonance to 29.5 ppm.

The ¹⁹ F NMR spectra were obtained using a magic angle spinning (MAS) experiment with proton decoupling. During the collection of spectra, a phase-modulated proton decoupling field of 80-100 kHz is used. The spectra are collected with a recirculation delay of up to 6 seconds for Form 1 and 5.25 seconds for Form 4. The number of scans is adjusted to obtain an adequate signal to noise ratio. For the ¹⁹ F chemical shift scale, use the ¹⁹ F MAS experiment with an external standard of trifluoroacetic acid (50%/50% v/v in H ₂ O), setting its resonance to -76.54 ppm.

Automatic peak collection is performed using Bruker-BioSpin TopSpin software version 3.6. Typically, a threshold of 4% relative intensity is used for peak preselection. The output of the automatic peak collection is visually checked to ensure that it is valid, and manual adjustments are made, if necessary. Although specific solid state NMR peak values are given here, there is a range for these peak values due to differences in instruments, samples, and sample preparation. This is common practice in the field of solid state NMR due to differences in peak positions. Typical variability in ¹³ C chemical shift along the abscissa is on the order of plus or minus 0.2 ppm, unless otherwise noted, for a crystalline solid. The variability in the ¹⁹ F chemical shift value is on the order of plus or minus 0.1 ppm. The heights of the solid NMR peaks given here are relative intensities. The NMR intensity of a solid can vary depending on the actual setting of the experimental parameters and the thermal history of the sample.

¹³ C NMR of the solid of Example 13 Form 1 was prepared as described above and the following list of peaks of Example 13 Form 1 was determined. The variability for ¹³ C chemical shift values is ±0.2 ppm unless otherwise noted.

¹³ C chemical shift (ppm) Relative intensity (%) ¹³ C Chemical shift (ppm) Relative intensity (%) ¹³ C Chemical shift (ppm) Relative intensity (%) 178.9 24 62.7 24 33.0±0.1 35 172.3 21 58.6 27 31.0±0.1 31 172.1 25 47.2 26 27.9±0.1 100 169.6 21 40.3 28 26.3 58 156.7 fourteen 39.4 24 26.0 41 123.5 10 39.0 31 20.8±0.1 49 122.6±0.1 6 37.8±0.1 48 13.0±0.1 47 118.5 4 37.4±0.1 41 116.1 4 34.6±0.1 33

¹⁹ F NMR solid of the compound of Example 13 Form 1 was prepared and the ¹⁹ F NMR peak of the solid was determined at a chemical shift of -73.3 ± 0.1 ppm.

Characteristic peaks for the compound of example 13, form 1 are ¹⁹ F peak with a chemical shift of -73.3±0.1 ppm. in combination with ¹³ C peaks with chemical shifts of 31.0 ± 0.1 ppm, 27.9 ± 0.1 ppm. and 178.9±0.2 ppm

Alternative recrystallization of example 13; obtaining a solid form 4

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ), Solid Form 4

Stage 1. Recrystallization (1R.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (13) from propan-2-yl acetate and heptane.

A mixture of propan-2-ylacetate (50 ml) and heptane (50 ml) was added to13, methyl solvate tert- butyl ether, Solid form 2 (from the second alternative synthesis from example 13, solvate methyltert-butyl ether; 10.02 g, 17.0 mmol) and the mixture is stirred at 20° C. and 3500 rpm. during the night. Heptane (50 ml) is then added slowly and stirring is continued for 30 minutes, then the mixture is cooled to 10° C. over 30 minutes. After stirring for another 2 hours, the suspension is filtered; the filter cake was washed with a mixture of propan-2-yl acetate (4 ml) and heptane (16 ml) and then dried at 55°C under vacuum to obtain (1R.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (13) as a crystalline solid. Part of this product is used in the next recrystallization. Yield: 7.74 g, 15.5 mmol, 91%.

Stage 2. Recrystallization of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ) from water.

Suspension 13 (from the previous step; 1.0 g, 2.0 mmol) in water (12 ml) was stirred at 5°C for 21 days, then the solid was collected by filtration. It is then dried under vacuum for 10 minutes and air dried in a thin layer on paper for 20 minutes to give (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[ (3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0] hexane-2-carboxamide ( 13 ) as a white crystalline solid. Powder X-ray diffraction pattern of this product, designated as Solid form 4, is given in figure 6; characteristic peaks are listed in Table J. Yield: 755 mg, 1.51 mmol, 76%. A method for collecting powder X-ray diffraction data is described in Alternative Synthesis of Example 13, Methyl tert -butyl ether solvate, Step 8.

Table J. Selected X-ray Powder Diffraction Peaks for Example 13 , Solid Form 4 Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) 7.6 fifteen 20.4 43 30.6 3 9.8 fourteen 20.7 44 30.8 five 10.8 7 21.1 42 31.3 3 11.2 4 21.6 nine 31.8 6 11.4 4 21.8 16 32.5 6 11.4 eight 22.3 53 32.8 4 11.7 five 23.1 fourteen 33.2 4 12.0 6 23.4 eleven 34.4 nine 12.3 82 24.2 nine 35.5 12 12.7 61 24.9 12 35.6 7 13.7 4 25.2 eight 35.6 7 14.9 3 26.1 4 36.0 3 15.1 five 27.0 five 36.4 3 15.9 100 27.2 fifteen 37.1 6 17.5 47 28.1 17 38.7 3 18.0 five 28.9 4 39.4 3 18.2 57 29.4 nine 39.5 3 18.5 21 29.5 4 39.8 4 18.8 37 29.8 five 20.0 12 30.0 21

Single crystal X-ray structural analysis of example 13, solid form 4

A sample of sample 13 is subjected to crystallization by diffusion at room temperature using ethyl acetate and pentane; one of the resulting crystals is used for single crystal x-ray structural analysis. The ORTEP chart for single crystal data is shown in Figure 7. Mercury software is used to calculate the powder diffraction pattern based on the resolved crystal structure; comparison with the diffraction pattern from the alternative recrystallization of example 13; obtaining solid form 4 identifies this product as solid form 4 (see figure 8). The characteristic peaks for these calculated data are shown in Table K.

Table K. Powder diffraction data for 13, solid form 1 calculated from single crystal X-ray diffraction analysis of example 13, solid form 4 Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) Angle (°2-theta)
+/-0.2° 2-Theta Relative intensity (%) 7.6 22 20.1 6 29.9 3 9.7 nineteen 20.4 40 30.0 13 10.8 10 20.7 31 30.7 3 11.2 4 21.1 32 30.9 3 11.3 7 21.6 eight 31.3 3 11.7 6 21.8 12 31.9 3 12.2 100 22.3 35 32.6 4 12.6 49 22.5 3 33.2 five 13.7 4 23.1 13 34.5 7 14.9 3 23.5 7 35.5 6 15.1 4 24.2 eight 35.8 6 15.7 fourteen 24.8 6 37.2 3 15.9 82 24.9 eight 39.9 3 17.5 35 25.2 7 18.2 54 26.2 4 18.5 fourteen 27.0 4 18.7 31 27.2 7 19.9 3 28.1 eleven

X-ray analysis of single crystals

Data collection is carried out on a Bruker D8 Venture diffractometer at -100°C. Data collection consists of omega and phi scans.

The structure was resolved by internal phasing using the SHELX software package in the space group of the orthorhombic class P 2 ₁ 2 ₁ 2 ₁ . In the future, the structure is refined by the full-matrix least squares method. All non-hydrogen atoms are found and refined using anisotropic displacement parameters.

Hydrogen atoms located on nitrogen were found from the difference Fourier map and refined with limited distances. The remaining hydrogen atoms are placed in calculated positions and allowed to move on their carrier atoms. The final refinement includes isotropic displacement parameters for all hydrogen atoms.

Absolute structure analysis using probability methods (Hooft, 2008) is performed using PLATON (Spek). Absolute stereochemistry not determined due to out-of-spec Hooft/Parsons/Flac parameters and standard deviations.

The final R-index is 6.3%. The final difference Fourier map showed no missing or shifted electron density.

The corresponding crystal, data acquisition, and refinement information are summarized in Table L. Atomic coordinates, bond lengths, bond angles, and bias parameters are listed in Tables M-P.

Software and Links

SHELXTL , Version 5.1, Bruker AXS, 1997.

PLATON , A. L. Spek, J. Appl. Cryst. 2003 , 36 , 7-13.

MERCURY , C. F. Macrae, P. R. Edington, P. McCabe, E. Pidcock, G. P. Shields, R. Taylor, M. Towler, and J. van de Streek, J. Appl. Cryst. 2006, 39 , 453-457.

OLEX2 , OV Dolomanov, LJ Bourhis, RJ Gildea, JAK Howard, and H. Puschmann, J. Appl. Cryst. 2009, 42 , 339-341.

R. W. W. Hooft, L. H. Straver, and A. L. Spek, J. Appl. Cryst. 2008 , 41 , 96-103.

HD Flack, Acta Cryst. 1983 , A39 , 867-881.

Table L. Crystal data and structure refinements for Example 13, Solid Form 4 . Empirical formula
formula mass
Temperature
Wavelength
Crystal system
space group
Lattice constant
Volume
Z
Density (calculated)
Absorption coefficient
F (000)
Crystal size
Theta interval for data collection
Index ranges
Collected Reflections
Independent Reflections
Fullness to theta=54.284°
Absorption correction
Refinement method
Data/Restrictions/Parameters
Compliance with F ²
Finite indices R [ I>2σ(I) ]
R indices (all data)
Absolute Structure Parameter
Excitation factor
The largest diff. peaks and valleys C ₂₃ H ₃₂ F ₃ N ₅ O ₄
499.53
173(2) K
1.54178 Å
orthothrombic
P 2 ₁ 2 ₁ 2 ₁
a =9.2114(9) Å
b = 15.1607(16) Å
c = 18.191(2) Å
2540.5(5) Å ³
4
1.306 Mg/ ^m3
0.892 mm ^-1
1056
0.100×0.060×0.040 mm ³
3.795-54.284°
-9<= h <=9, -15<= k <=15, -18<= l <=18
15896
3070 [ _Rint = 0.1260 ]
99.1%
empirical
Least squares in the full-matrix approximation in F ²
3070/9/349
1.067
R 1=0.0625, wR 2=0.1515
R 1=0.0696, wR 2=0.1578
0.14(14)
n/a
0.280 and -0.220 e.Å ^-3 α= 90°
β= 90°
γ =90°

Table M. Atomic coordinates (x 10 ⁴ ) and equivalent isotropic substitution parameters (Å ² ×10 ³ ) for example 13, Solid form 4 . U(eq) is defined as one third of the trace of the orthogonalized tensor U ^ij .

Table N. Bond lengths [Å] and angles [°] for example 13, solid form 4 .

Symmetric transformations are used to create equivalent atoms.

Table P. Anisotropic substitution parameters (Å ² ×10 ³ ) for example 13, solid form 4 . The exponent of the anisotropic substitution factor takes the form: −2π ² [h ² a* ² U ¹¹ + ... + 2 hka* b* U ¹² ].

¹³ C NMR Peak List Solid State PF-07321332-00 Form 4 Variability for ¹³ C chemical shift values is ±0.2 ppm unless otherwise noted.

¹³ C Chemical shift (ppm) Relative intensity (%) ¹³ C Chemical shift (ppm) Relative intensity (%) ¹³ C Chemical shift (ppm) Relative intensity (%) 179.3 16 62.0 17 32.9 16 178.8 eleven 58.5 26 31.9 23 172.3 twenty 47.1 26 31.3±0.1 16 169.6 eight 41.5 17 29.7 eighteen 168.6 12 40.2 12 27.9±0.1 100 156.7 12 38.8 22 26.9±0.1 36 123.7 4 38.3 27 26.2 28 120.1 6 37.9 43 25.9 eighteen 119.1 4 37.5 39 21.6±0.1 31 118.5 4 37.0 25 20.8±0.1 22 62.4 16 34.4 13 12.9±0.1 46

¹⁹ F NMR solid of Example 13 Form 4 was obtained and a peak at -73.6 ± 0.1 with a relative intensity of 100% was determined.

For the compound of example 13, form 4, six characteristic peaks were identified: ¹⁹ F chemical shift at -73.6±0.1 ppm. and ¹³ C chemical shifts at 26.9±0.1 ppm, 21.6±0.1 ppm, 41.5±0.2 ppm, 27.9±0 .1 ppm, and 12.9±0.1. ¹⁹ F peak with a chemical shift of -73.6±0.1 ppm is characteristic of the compound of example 13, form 4. ¹³ C peaks at 26.9±0.1 ppm, 21.6±0.1 ppm and 41.5±0.1 ppm each is a characteristic peak of Example 13, Form 4. ¹³ C peaks at 27.9 ppm. and 12.9 ppm each is characteristic of Example 13 Form 4 when taken in combination with one or more of the peaks selected from the ¹³ C peaks at 21.6 ppm, 26.9 ppm and 41.5 ppm and ¹⁹ F peak at -73.6 ppm.

Fourth Alternative Synthesis of Example 13, Methyl t -Butyl Ether Solvate

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate (13, methyl tert -butyl ether solvate)

To 0°C suspensionC43 (5.0 g, 9.7 mmol) in propan-2-yl acetate (7.6 ml/g, 38 ml) loaded with 4-methylmorpholine (4.5 equivalents; 4.8 ml, 44 mmol). The resulting suspension was charged with trifluoroacetic anhydride (2.25 equivalents; 3.1 ml, 22 mmol) over 1 hour via a metering pump. After stirring at 0°C for at least 1 hour, the reaction mixture is heated to about 20°C, quenched with water (8 ml/g, 40 ml) and stirred for at least 10 minutes. After decanting, the lower (aqueous) layer was discarded and water (8 ml/g, 40 ml) was added to the organic layer. After stirring for at least 10 minutes, the layers are separated and the bottom (water) layer is discarded. The organic layer is concentrated under reduced pressure to about 4 ml/g (about 20 ml), then the vacuum is broken using nitrogen and the solution is heated to about 50°C. Methyltert-butyl ether (12 ml/g, 60 ml) is added slowly over at least 4 hours via an addition funnel and the reaction mixture is kept at 50°C for at least 1 hour, then cooled to 25°C within 1 hour. The resulting suspension is kept at 25°C overnight, then filtered, washed successively with a mixture of propan-2-ylacetate and methyltert-butyl ether [1:3 (v/v); 2 ml/g] and methyltert-butyl ether (2 ml/g, 10 ml) and filter dried for at least 30 minutes. The solids are then transferred to a vacuum oven at 50°C and dried for at least 8 hours to obtain (1R.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl solvatetert-butyl ether (13, methyl solvate tert- butyl ether) as an off-white solid. Yield: 3.4 g, 5.8 mmol, 60%.

Fifth Alternative Synthesis of Example 13, Methyl t -Butyl Ether Solvate

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, methyl tert -butyl ether solvate (13, methyl tert -butyl ether solvate)

To a suspension of C43 (4.0 g, 7.7 mmol) in acetonitrile (10 mL/g, 40 mL) at room temperature was charged 1-methyl- 1H -imidazole (4.6 equivalents; 2.84 mL, 35. 6 mmol) and the resulting mixture is heated to approximately 30°C. A solution of 2,4,6-tripropyl-1,3,5,2,4,6-trioxatrifosfinan 2,4,6-trioxide (50% by weight solution in acetonitrile; 2 equivalents; 10.8 ml) was added over at least 6 hours using the pump. The reaction mixture is then stirred for at least 10 hours, cooled to 25° C. and carefully quenched by addition of saturated aqueous sodium bicarbonate (7 ml/g, 28 ml) (exothermic and no gas). Acetonitrile is then distilled off under reduced pressure; ethyl acetate (10 ml/g, 40 ml) and additional saturated aqueous sodium bicarbonate solution (5 ml/g, 20 ml) were added to the resulting mixture. After phase separation, the lower (aqueous) layer was discarded and the organic layer was washed with saturated aqueous sodium bicarbonate (3.5 ml/g; 14 ml). The aqueous layer was discarded again and the organic layer was concentrated under reduced pressure to about 1 ml/g (4 ml). Methyl tert -butyl ether (9 ml/g; 36 ml) was charged and the resulting solution was heated to 50° C. to rapidly form a suspension. This suspension is stored at 50°C for at least 30 minutes, then it is cooled to 25°C for 1 hour and stored at 25°C for at least 8 hours. The suspension is then filtered, washed with a mixture of ethyl acetate and methyl t- butyl ether [1:3 (v/v); 2 ml/g], and then washed with methyl tert -butyl ether (2 ml/g; 8 ml). The combined solid is filter dried for at least 30 minutes, transferred to a vacuum oven at about 50° C., and dried for at least 8 hours to give (1 R ,2 S ,5 S ) - N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl) -L-valyl]-3-azabicyclo[3,1,0]hexane-2-carboxamide, methyl tert -butyl ether solvate (13, methyl tert -butyl ether solvate) as an off-white solid. Yield: 2.9 g, 4.9 mmol, 64%.

Formulation Examples for Compound of Example 13

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide or a hydrate or solvate thereof or a pharmaceutically acceptable salt of the compound, hydrate or solvate is formulated to form a conventional tablet and immediate release film-coated for oral administration, where doses range from 100 to 250 mg. As an example, the immediate release formulation is described in the formulation table and contains the usual inactive excipients microcrystalline cellulose and lactose monohydrate (diluents), crospovidone (disintegrant), colloidal silicon dioxide (glidant) and sodium stearyl fumarate (lubricant). The immediate release tablets are film coated using commercially available film coating formulations including Opadry white and Opadry pink. All excipients in the film-coated tablet are commonly used and are present at known levels. The present formulation is an example of an immediate release tablet formulation and as such, one skilled in the art will be able to use readily available conventional formulation techniques using alternative formulation excipients to obtain suitable tablets and achieve the desired tablet quality characteristics.

Formulation Examples: Typical coated tablet formulations for ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl} -6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide.

Immediate Release Tablets _ _ _ dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (referred to below as API) is produced using conventional, standard batch processes for solids oral tablets with immediate release. Examples of standard batch processes that can be used to produce tablets ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide with immediate release include direct compression, dry granulation and wet granulation. Alternatively, a continuous manufacturing process may be used. After pressing the tablets, the tablet cores are coated with a film. Film coating of tablets can be carried out using continuous coating or using a conventional batch film coating process.

Tablet composition: tablet formulations 100 mg, 150 mg and 250 mg (1 R ,2 S , 5 S )- N yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide

Component Role 100 mg tablet (% w/w) 100 mg tablet (mg/tablet) 150 mg tablet (% w/w) 150 mg tablet (mg/tablet) 250 mg tablet (% w/w) 250 mg tablet (mg/tablet) API Active Ingredient 20.00 100.00 20.00 150.00 25.00 250.00 Microcrystalline cellulose Diluent 49.33 246.67 49.33 370.00 46.00 460.00 Lactose monohydrate Diluent 24.67 123.33 24.67 185.00 23.00 230.00 Crospovidone Baking powder 3.00 15.00 3.00 22.50 3.00 30.00 colloidal silicon dioxide Glidant 1.00 5.00 1.00 7.50 1.00 10.00 Magnesium stearyl fumarate (intragranulated) Lubricant 1.00 5.00 1.00 7.50 1.00 10.00 Magnesium stearyl fumarate (extragranulated) Lubricant 1.00 5.00 1.00 7.50 1.00 10.00 Whole core 100.00 500.00 100.00 750.00 100.00 1000.00 Opadry White
(YS-1-7027-SP) Coating 3.50 17.50 - - - - - - - - Opadry Pink
(058140011) Coating - - - 3.00 22.50 - - - -

Example 14

N -[(2 S )-1-({(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-5,5,5-trifluoro- 1-oxopentan-2-yl]-4-methoxy-1 H -indole-2-carboxamide ( 14 )

Step 1. Synthesis of 9H- fluoren -9-ylmethyl [( 2S )-1-amino-5,5,5-trifluoro-1-oxopentan-2-yl]carbamate ( C34 ).

Sodium bicarbonate (4.8 g, 57 mmol) is added to a solution of 5,5,5-trifluoro-L-norvalinamide, hydrochloride (it is synthesized using the method described for its enantiomer in JE Starrett, PCT Int. Appl., 2010107997 , September 23, 2010; 4.0 g, 19 mmol) and 9 H -fluoren-9-ylmethyl carbonochloridate (Fmoc chloride; 10.2 g, 39.4 mmol) in water (80 ml). The resulting suspension is stirred at 15°C - 25°C for 24 hours, then it is divided between water and dichloromethane. The organic layer is washed successively with water and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo to give C34 as a solid. Yield: 6.2 g, 16 mmol, 83%. LCMS m/z 393.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 7.9 (d, 2H), 7.7 (m, 2H), 7.5 (d, 1H), 7.4 (m, 5H), 7 .1 (ws, 1H), 4.3 (m, 3H), 4.0 (m, 1H), 2.2 (m, 2H), 1.9 (m, 1H), 1.7 (m, 1H).

Step 2. Synthesis of N -[(9H- fluoren -9-ylmethoxy)carbonyl]-5,5,5-trifluoro-L-norvaline ( C35 ).

To a solution of C34 (6.2 g, 16 mmol) in 1,4-dioxane (60 ml) was added hydrochloric acid (3 M; 10 ml, 30 mmol) and the reaction mixture was stirred at 80° C. for 16 hours. It is then partitioned between water and dichloromethane, and the organic layer is washed with water and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . The residue is triturated with petroleum ether to give C35 as a solid. Yield: 5.5 g, 14 mmol, 88%. LCMS m/z 392.1 [M−H] ⁻ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 12.83 (br s, 1H), 7.89 (d, J= 7.4 Hz, 2H), 7.77-7.67 (m, 3H) , 7.46-7.38(m, 2H), 7.36-7.28(m, 2H), 4.38-4.28(m, 2H), 4.26-4.19(m, 1H), 4.06 (ddd, J= 9, 9, 4.9 Hz, 1H), 2.43-2.15 (m, 2H), 2.01-1.89 (m, 1H), 1 .89-1.75 (m, 1H).

Step 3. Synthesis of benzyl N -[(9H- fluoren -9-ylmethoxy)carbonyl]-5,5,5-trifluoro-L-norvalinate ( C36 ).

A mixture of C35 (435 mg, 1.11 mmol), benzyl bromide (0.263 ml, 2.21 mmol) and sodium bicarbonate (464 mg, 5.52 mmol) in N,N -dimethylformamide (20 ml) was stirred for 15 hours at 25°C. The reaction mixture was then diluted with water (30 ml) and extracted with ethyl acetate (3 x 30 ml), the combined organic layers were washed successively with saturated aqueous sodium chloride and 5% aqueous lithium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (gradient: 0% - 100% ethyl acetate in petroleum ether) gives C36 as a white solid. Yield: 510 mg, 1.05 mmol, 95%. LCMS m/z 506.1 [M+Na ⁺ ].

Step 4. Synthesis of benzyl 5,5,5-trifluoro-L-norvalinate ( C37 ).

Diethylamine (10 ml) was added to a 0°C mixture of C36 (510 mg, 1.05 mmol) in acetonitrile (25 ml). Then the reaction mixture is stirred at 20°C for 2 hours, it is concentrated under reduced pressure; then chromatography on silica gel (gradient: 0% - 10% methanol in dichloromethane) gives C37 as a colorless oil. Yield: 250 mg, 0.957 mmol, 91%. LCMS m/z 302.9 [M+CH ₃ CN+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 7.42-7.32 (m, 5H), 5.17 (s, 2H), 3.50 (dd, J =8.4, 5.0 Hz , 1H), 2.32-2.13 (m, 2H), 2.01 (dddd, J = 13.7, 10.8, 5.2, 5.2 Hz, 1H), 1.76 (dddd , J =13.6, 10.8, 8.4, 5.3 Hz, 1H).

Step 5. Synthesis of benzyl 5,5,5-trifluoro- N -[(4-methoxy- 1H -indol-2-yl)carbonyl]-L-norvalinate ( C38 ).

To a 0°C solution of C37 (250 mg, 0.957 mmol) and 4-methoxy- 1H -indole-2-carboxylic acid (220 mg, 1.15 mmol) in N,N -dimethylformamide (10 ml) was added hexafluorophosphate O - (7-Azabenzotriazol-1-yl) -N,N,N',N'- tetramethyluronium (GATU; 437 mg, 1.15 mmol), then 4-methylmorpholine (194 mg, 1.92 mmol) was added dropwise. Stirring is continued at 0°C - 10°C for 1 hour, then the reaction mixture is diluted with water (20 ml) and aqueous citric acid solution (20 ml) and extracted with ethyl acetate (3×20 ml). The combined organic layers were washed successively with saturated aqueous sodium bicarbonate (30 ml), saturated aqueous sodium chloride, and aqueous lithium chloride (5%, 20 ml), then dried over sodium sulfate, filtered, and concentrated in vacuo . The residue was purified using silica gel chromatography (gradient: 0% - 100% ethyl acetate in petroleum ether) to give C38 as a white solid. Yield: 350 mg, 0.806 mmol, 84%. LCMS m/z 435.1 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 9.09 (br s, 1H), 7.42-7.33 (m, 5H), 7.23 (dd, J =8.8 Hz, 1H), 7.09 (d, J =2.3 Hz, 1H), 7.03 (d, J =8.3 Hz, 1H), 6.76 (br, J =7.6 Hz, 1H), 6, 53 (d, J = 7.8 Hz, 1H), 5.25 (AB quartet, J _{AB =} 12.1 Hz, δν _{AB =} 11.4 Hz, 2H), 4.94-4.87 (m, 1H), 3.96 (s, 3H), 2.35-2.14 (m, 2H), 2.14-1.96 (m, 2H).

Step 6. Synthesis of 5,5,5-trifluoro- N -[(4-methoxy- 1H -indol-2-yl)carbonyl]-L-norvaline ( C39 ).

A mixture of C38 (350 mg, 0.806 mmol) and palladium-carbon (10%, 85.7 mg, 80.5 µmol) in methanol (10 ml) was hydrogenated for 16 hours at 20° C. and 15 psi . The reaction mixture is then filtered and the filter cake washed with methanol (10 ml); the combined filtrates were concentrated in vacuo and chromatographed on silica gel (eluent: ethyl acetate) to give C39 as a white solid. Yield: 270 mg, 0.784 mmol, 97%. LCMS m/z 345.0 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 11.62 (br s, 1H), 8.61 (d, J =7.9 Hz, 1H), 7.32 (d, J =2.2 Hz , 1H), 7.11 (dd, J = 8.8 Hz, 1H), 7.01 (d, half AB quartet, J = 8.2 Hz, 1H), 6.51 (d, J = 7, 6 Hz, 1H), 4.47 (ddd, J = 8.5, 8.5, 4.8 Hz, 1H), 3.89 (s, 3H), 2.5-2.27 (m, 2H , putative; partially obscured by solvent peak), 2.12-1.92 (m, 2H).

Step 7. Synthesis of 5,5,5-trifluoro- N -[(4-methoxy- 1H -indol-2-yl)carbonyl]-L- norvalyl -3-[(3S)-2-oxopyrrolidine-3- yl]-L-alanineamide ( C40 ).

A 0°C mixture of C16 (58.2 mg, 0.218 mmol) and C39 (75.0 mg, 0.218 mmol) in N,N -dimethylformamide (4 mL) was treated with O- (7-azabenzotriazol-1-yl) -N hexafluorophosphate , N,N',N'- tetramethyluronium (GATU; 99.4 mg, 0.261 mmol) and 4-methylmorpholine (44.1 mg, 0.436 mmol). Then the reaction mixture was stirred at 0°C for 1 hour, it was diluted with water (20 ml) and aqueous citric acid solution (1 M; 20 ml), and extracted with ethyl acetate (3×30 ml). The combined organic layers were washed with saturated aqueous sodium bicarbonate (20 ml) and saturated aqueous sodium chloride (3×20 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (eluent: 10:1 ethyl acetate/methanol) gives C40 as a white solid. Yield: 72 mg, 0.145 mmol, 66%. LCMS m/z 498.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 11.60 (br s, 1H), 8.52 (d, J =7.7 Hz, 1H), 8.20 (d, J =8.3 Hz , 1H), 7.61 (s, 1H), 7.42-7.33 (m, 2H), 7.14-7.05 (m, 2H), 7.00 (d, half AB quartet, J = 8.2 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 4.58-4.46 (m, 1H), 4.32-4.22 (m, 1H) , 3.89 (s, 3H), 3.18-3.02 (m, 2H), 2.45-2.21 (m, 3H), 2.18-2.07 (m, 1H), 2 .06-1.88 (m, 3H), 1.73-1.59 (m, 1H), 1.59-1.48 (m, 1H).

Step 8. Synthesis of N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-5.5, 5-trifluoro-1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 14 ).

To a mixture of C40 (52 mg, 0.10 mmol) in dichloromethane (13 mL) was added methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 37 mg, 0.16 mmol). The reaction mixture was stirred at room temperature for 1.5 hours, then methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 37 mg, 0.16 mmol) was added again and stirring was continued for 16 hours. The final addition of methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 24.9 mg, 0.105 mmol) was then stirred for 2 hours, then the reaction mixture was diluted with water (20 ml) and extracted with dichloromethane (3×10 ml). The combined organic layers are washed with saturated aqueous sodium chloride (2×20 ml), dried over sodium sulfate, filtered, and concentrated in vacuo ; preparative thin layer chromatography (eluent: 20:1 ethyl acetate/methanol) gives N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl ]ethyl}amino)-5,5,5-trifluoro-1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 14 ) as a white solid. Yield: 17.4 mg, 36.3 µmol, 36%. This product is combined with the purified products of two other syntheses 14 (3 mg and 4 mg) and subjected to supercritical liquid chromatography [Column: Chiral Technologies ChiralCel OD-H, 30×250 mm, 5 μm; Mobile phase: 7:3 carbon dioxide/(ethanol containing 0.1% ammonium hydroxide); Flow rate: 60 ml/minute] to give N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino )-5,5,5-trifluoro-1-oxopentan-2-yl]-4-methoxy- 1H -indole-2-carboxamide ( 14 ) as a solid. Yield: 11.3 mg, 23.6 µmol, 46% for supercritical liquid chromatography. LCMS m/z 480.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 11.61 (br s, 1H), 8.96 (d, J =8.0 Hz, 1H), 8.61 (d, J =7.7 Hz , 1H), 7.71 (s, 1H), 7.37 (d, J = 2.2 Hz, 1H), 7.11 (dd, J = 8.8 Hz, 1H), 7.01 (d , half AB quartet, J = 8.2 Hz, 1H), 6.51 (d, J = 7.7 Hz, 1H), 5.03-4.94 (m, 1H), 4.51-4, 43 (m, 1H), 3.89 (s, 3H), 3.19-3.07 (m, 2H), 2.43-2.28 (m, 3H), 2.20-2.08 ( m, 2H), 2.06-1.92 (m, 2H), 1.86-1.76 (m, 1H), 1.76-1.64 (m, 1H).

Examples 15, 16, 17, 18, and 19

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-7-fluoro-4-methoxy-1H-indole-2-carboxamide (fifteen), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-5-fluoro-4-methoxy-1H-indole-2-carboxamide (16), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-3-fluoro-4-methoxy-1H-indole-2-carboxamide (17), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-5,7-difluoro-4-methoxy-1H-indole-2-carboxamide (eighteen) and N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-3,5-difluoro-4-methoxy-1H-indole-2-carboxamide (nineteen)

Mixture4(10.0 mg, 22.8 µmol), decatungstate tetra-n-butylammonium (TBADT; 3.78 mg, 1.14 µmol) andN-fluorine-N-(phenylsulfonyl)benzenesulfonamide (8.61 mg, 27.3 µmol) is treated with acetonitrile (0.75 ml), water (0.5 ml) and trifluoroacetic acid (1.74 µl, 22.6 µmol) under argon. The reaction tube is then sealed, placed in an EvoluChem™ PhotoRedOx Box equipped with a fan and irradiated with black light (PAR20-18W LG 365 nm, 100-240 VAC) at 25°C for 16 hours. An aqueous solution of potassium phosphate (1 M, pH 7.45; 1 ml) was added to the reaction mixture, followed by alternating aliquots of water and acetonitrile to maintain the purified solution in a final volume of 18 ml. Aliquots (3 ml) of this mixture are applied to Biotage Isolute C18 solid phase extraction cartridges which are pretreated with methanol (3 ml) then water (3 ml). The cartridges were washed with water (3 ml) and 20% acetonitrile in 20 mM aqueous ammonium acetate (3 ml), then eluted with acetonitrile (3 ml). After evaporating the eluates in a vacuum centrifuge, the residues are reconstituted in a mixture of 1% aqueous formic acid and acetonitrile and combined to a total of 6 ml. This solution is divided in half and each half is subjected to reverse phase HPLC (Column: Phenomenex Luna C18.10×250 mm, 10 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile; gradient: 15% B for 5 minutes, then 15% - 70% B for 70 minutes, then 70% - 95% B for 15 minutes; flow rate: 2 ml/min). Fractions are collected every 20 seconds and similar fractions of interest from the two separations are pooled and concentrated. These fractions are then purified by reverse phase HPLC (Column: Agilent Polaris C18, 4.6×250 mm, 5 µm; Mobile phase A: water containing 10 mM ammonium acetate; Mobile phase B: acetonitrile; Gradient: 10% B over 5 minutes, then 10% - 35% B over 35 minutes, then 35% - 60% B over 15 minutes, then 60% - 95% B over 9 minutes; Flow rate: 0.8 ml/min). Fractions are collected every 20 seconds to obtainN-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-7-fluoro-4-methoxy-1H-indole-2-carboxamide (fifteen), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-5-fluoro-4-methoxy-1H-indole-2-carboxamide (16), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-3-fluoro-4-methoxy-1H-indole-2-carboxamide (17), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-5,7-difluoro-4-methoxy-1H-indole-2-carboxamide (eighteen), and N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan-2-yl]-3,5-difluoro-4-methoxy-1H-indole-2-carboxamide (nineteen).

Example No. Retention time, first HPLC cleanup (minutes) Retention time, second HPLC purge (minutes) fifteen 57.1 45.1 16 57.7 45.9 17 58.6 46.7 eighteen 59.9 47.9 nineteen 61.7 49.5

15 - First separation, fraction numbers 172-174; Second separation, fraction numbers 136-137. Yield: 58 µg, 0.13 µmol, 0.6%. High resolution MS m/z 458.2201 [M+H] ⁺ ; calculated for C ₂₃ H ₂₉ FN ₅ O ₄ , 458.2204. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 12.05 (br s, 1H), 8.93 (d, J =7.9 Hz, 1H), 8.51 (d, J =7.6 Hz , 1H), 7.70 (s, 1H), 7.37 (d, J = 2.3 Hz, 1H), 6.92 (dd, J = 10.8, 8.6 Hz, 1H), 6 .42 (dd, J = 8.4, 2.5 Hz, 1H), 5.01-4.94 (m, 1H), 4.50-4.43 (m, 1H), 3.87 (s , 3H), 3.18-3.07 (m, 2H), 2.40-2.31 (m, 1H), 2.19-2.08 (m, 2H), 1.85-1.76 (m, 1H), 1.76-1.64 (m, 3H), 1.58-1.49 (m, 1H), 0.94 (d, J =6.3 Hz, 3H), 0, 89 (d, J =6.3 Hz, 3H). Retention time: 7.90 minutes (Analytical conditions. Column: Phenomenex Kinetex XB-C18, 2.1×100 mm, 2.6 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile ; Gradient: 5% B over 0.5 minutes, then 5% - 70% B over 10.5 minutes, then 70% - 95% B over 2 minutes; Flow rate: 0.4 ml/min).

16 - First separation, fraction numbers 172-174; Second separation, fraction numbers 138-139. Yield: 153 µg, 0.33 µmol, 1.4%. High resolution MS m/z 458.2201 [M+H] ⁺ ; calculated for C ₂₃ H ₂₉ FN ₅ O ₄ , 458.2204. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 11.73 (br s, 1H), 8.95 (d, J =8.0 Hz, 1H), 8.61 (d, J =7.7 Hz , 1H), 7.70 (s, 1H), 7.49 (s, 1H), 7.09-7.03 (m, 2H), 5.01-4.94 (m, 1H), 4, 51-4.43 (m, 1H), 4.06 (s, 3H), 3.18-3.07 (m, 2H), 2.40-2.31 (m, 1H), 2.19- 2.08 (m, 2H), 1.80 (ddd, J =13.6, 9.2, 7.2 Hz, 1H), 1.76-1.65 (m, 3H), 1.58- 1.50 (m, 1H), 0.94 (d, J =6.3 Hz, 3H), 0.89 (d, J =6.3 Hz, 3H). Retention time: 7.94 minutes (Analytical conditions identical to those used for 1 5 ).

17 - First separation, fraction numbers 176-177; Second separation, fraction numbers 141-142. Yield: 22 µg, 0.048 µmol, 0.21%. High resolution MS m/z 458.2199 [M+H] ⁺ ; calculated for C ₂₃ H ₂₉ FN ₅ O ₄ , 458.2204. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 11.45 (s, 1H), 8.94 (d, J =7.8 Hz, 1H), 7.71 (s, 1H), 7.62 (bd, J =7.5 Hz, 1H), 7.16 (dd, J =8.8 Hz, 1H), 6.95 (bd, J =8.3 Hz, 1H), 6.54 (d , J = 7.8 Hz, 1H), 5.02-4.94 (m, 1H), 4.54-4.46 (m, 1H), 3.88 (s, 3H), 3.19- 3.07 (m, 2H), 2.41-2.31 (m, 1H), 2.20-2.08 (m, 2H), 1.85-1.77 (m, 1H), 1, 76-1.63 (m, 3H), 1.61-1.53 (m, 1H), 0.94 (d, J =6.3 Hz, 3H), 0.91 (d, J =6, 3 Hz, 3H). Retention time: 8.06 minutes (Analytical conditions identical to those used for 1 5 ).

18 - First separation, fraction numbers 180-181; Second separation, fraction number 145. Yield: 17 μg, 0.036 μmol, 0.16%. High resolution MS m/z 476.2100 [M+H] ⁺ ; calculated for C ₂₃ H ₂₈ F ₂ N ₅ O ₄ , 476.2109. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 12.23 (s, 1H), 8.96 (d, J =8.0 Hz, 1H), 8.62 (d, J =7.6 Hz , 1H), 7.70 (s, 1H), 7.52-7.48 (m, 1H), 7.13 (dd, J =11.11 Hz, 1H), 5.02-4.94 ( m, 1H), 4.53-4.44 (m, 1H), 4.01 (s, 3H), 3.18-3.07 (m, 2H), 2.38-2.30 (m, 1H), 2.19-2.08 (m, 2H), 1.81 (ddd, J =13.6, 9.1, 7.0 Hz, 1H), 1.76-1.65 (m, 3H), 1.59-1.51 (m, 1H), 0.95 (d, J =6.2 Hz, 3H), 0.90 (d, J =6.3 Hz, 3H). Retention time: 8.20 minutes (Analytical conditions identical to those used for 1 5 ).

19 - First separation, fraction numbers 185-187; Second separation, fraction numbers 150-151. Yield: 35 µg, 0.074 µmol, 0.32%. High resolution MS m/z 476.2107 [M+H] ⁺ ; calculated for C ₂₃ H ₂₈ F ₂ N ₅ O ₄ , 476.2109. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 11.64 (s, 1H), 8.94 (d, J =7.9 Hz, 1H), 7.80 (brd, J =7 Hz, 1H ), 7.71 (s, 1H), 7.16 (dd, component of the ABX system, J = 11.9, 9.1 Hz, 1H), 7.08 (sd, half of the AB quartet, J = 8.5 Hz, 1H), 5.02-4.94 (m, 1H), 4.55-4.47 (m, 1H), 3.99 (s, 3H), 3.19-3.08 (m, 2H), 2.41-2.32 (m, 1H), 2.19-2.10 (m, 2H), 1.81 (ddd, J =13.7, 9.0, 7.2 Hz, 1H), 1.77-1.63 (m, 3H), 1.57 (ddd, J =12.9, 8.4, 4.8 Hz, 1H), 0.94 (d, J =6, 4 Hz, 3H), 0.91 (d, J =6.4 Hz, 3H). Retention time: 8.44 minutes (Analytical conditions identical to those used for 1 5 ).

Examples 20, 21, 22, and 23

N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-7-fluoro-4-methoxy-1H-indole-2-carboxamide (twenty), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-5-fluoro-4-methoxy-1H-indole-2-carboxamide (21), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-3-fluoro-4-methoxy-1H-indole-2-carboxamide (22) and N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-5,7-difluoro-4-methoxy-1H-indole-2-carboxamide (23)

Mixture10(10.0 mg, 22.0 µmol), decatungstate tetra-n-butylammonium (TBADT; 3.66 mg, 1.10 µmol) andN-fluorine-N-(phenylsulfonyl)benzenesulfonamide (8.34 mg, 26.4 μmol) is treated with acetonitrile (0.75 ml), water (0.5 ml) and trifluoroacetic acid (1.69 μl, 21.9 μmol) under argon. The reaction tube is then sealed, placed in an EvoluChem™ PhotoRedOx Box equipped with a fan, and irradiated with black light (PAR20-18W LG 365 nm, 100-240 VAC) at 25°C for 16 hours. An aqueous solution of potassium phosphate (1 M, pH 7.45; 1 ml) was added to the reaction mixture, followed by alternating aliquots of water and acetonitrile to keep the purified solution in a final volume of 18 ml. Aliquots (3 ml) of this mixture were applied to Biotage Isolute C18 solid phase extraction cartridges, which were pretreated with methanol (3 ml) followed by aqueous ammonium acetate (10 mM; 3 ml). The cartridges were washed with aqueous ammonium acetate (10 mM; 3 ml) and 20% acetonitrile in 20 mM ammonium acetate (3 ml), then eluted with acetonitrile (3 ml). After evaporating the eluates in a vacuum centrifuge, the residues are reconstituted in a mixture of 1% aqueous formic acid and acetonitrile and combined to a total of 6 ml. This solution was divided in half and each half was subjected to reverse phase HPLC (Column: Phenomenex Luna C18.10×250 mm, 10 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile; Gradient: 15% B for 5 minutes, then 15% - 70% B for 70 minutes, then 70% - 95% B for 15 minutes; flow rate: 2 ml/min). Fractions are collected every 20 seconds and similar fractions of interest from the two partitions are pooled and concentrated. These fractions are then purified by reverse phase HPLC (Column: Agilent Polaris C18, 4.6×250 mm, 5 µm; Mobile phase A: water containing 10 mM ammonium acetate; Mobile phase B: acetonitrile; Gradient: 10% B over 5 minutes, then immediate rise to 20% B, then 20% - 40% B for 35 minutes, then 40% - 60% B for 15 minutes, then 60% - 95% B for 9 minutes; Flow rate: 0.8 ml/min). Fractions are collected every 20 seconds, yieldingN-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-7-fluoro-4-methoxy-1H-indole-2-carboxamide (twenty), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-5-fluoro-4-methoxy-1H-indole-2-carboxamide (21), N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-3-fluoro-4-methoxy-1H-indole-2-carboxamide (22) and N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-5,7-difluoro-4-methoxy-1H-indole-2-carboxamide (23).

Example No. Retention time, first HPLC cleanup (minutes) Retention time, second HPLC purge (minutes) twenty 61.2 49.8 21 61.2 50.2 22 62.3 50.8 23 63.5 51.9

20 - First separation, fraction numbers 183-185; Second separation, fraction numbers 150-151. Yield: 24 µg, 0.051 µmol, 0.23%. High resolution MS m/z 472.2342 [M+H] ⁺ ; calculated for C ₂₄ H ₃₁ FN ₅ O ₄ , 472.2360. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 12.05 (br s, 1H), 8.91 (d, J =8.0 Hz, 1H), 8.52 (d, J =8.0 Hz , 1H), 7.69 (s, 1H), 7.37-7.32 (m, 1H), 6.92 (dd, J =10.9, 8.4 Hz, 1H), 6.41 ( dd, J =8.5, 2.7 Hz, 1H), 5.00-4.93 (m, 1H), 4.52 (dd, J =8.5, 8.2, 3.7 Hz, 1H), 3.87 (s, 3H), 3.17-3.05 (m, 2H), 2.38-2.30 (m, 1H), 2.18-2.06 (m, 2H) , 1.85-1.64 (m, 4H), 0.94 (s, 9H). Retention time: 8.32 minutes (Analytical conditions. Column: Phenomenex Kinetex XB-C18, 2.1×100 mm, 2.6 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile ; Gradient: 5% B over 0.5 minutes, then 5% - 70% B over 10.5 minutes, then 70% - 95% B over 2 minutes; Flow rate: 0.4 ml/minute).

21 - First separation, fraction numbers 183-185; Second separation, fraction numbers 152-153. Yield: 68 µg, 0.14 µmol, 0.64%. High resolution MS m/z 472.2344 [M+H] ⁺ ; calculated for C ₂₄ H ₃₁ FN ₅ O ₄ , 472.2360. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 11.72 (br s, 1H), 8.91 (d, J =8.1 Hz, 1H), 8.59 (d, J =8.0 Hz , 1H), 7.69 (s, 1H), 7.47 (d, J =2.2 Hz, 1H), 7.09-7.04 (m, 2H), 5.00-4.93 ( m, 1H), 4.52 (ddd, J =8.5, 8.5, 3.8 Hz, 1H), 4.06 (br, 3H), 3.17-3.05 (m, 2H) , 2.39-2.31(m, 1H), 2.18-2.06(m, 2H), 1.84-1.77(m, 1H), 1.78(dd, J =13, 9, 9.0 Hz, 1H), 1.74-1.64 (m, 2H), 0.94 (s, 9H). Retention time: 8.34 minutes (Analytical conditions identical to those used for 20 ).

22 - First separation, fraction numbers 187-188; Second separation, fraction number 154. Yield: 5 μg, 0.011 μmol, 0.05%. High resolution MS m/z 472.2354 [M+H] ⁺ ; calculated for C ₂₄ H ₃₁ FN ₅ O ₄ , 472.2360. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 11.45 (s, 1H), 8.91 (d, J =7.9 Hz, 1H), 7.70 (s, 1H), 7.57 (dd, J =8.2, 3.6Hz, 1H), 7.15 (dd, J =8.8Hz, 1H), 6.95 (bd, J =8.3Hz, 1H), 6 .54 (d, J = 7.8 Hz, 1H), 5.00-4.94 (m, 1H), 4.56-4.49 (m, 1H), 3.88 (s, 3H), 3.18-3.07 (m, 2H), 2.40-2.32 (m, 1H), 2.17-2.09 (m, 2H), 1.84-1.77 (m, 1H ), 1.76-1.65 (m, 3H), 0.95 (s, 9H). Retention time: 8.51 minutes (Analytical conditions identical to those used for 20 ).

23 - First separation, fraction numbers 190-192; Second separation, fraction numbers 156-157. Yield: 21 µg, 0.043 µmol, 0.19%. High resolution MS m/z 490.2258 [M+H] ⁺ ; calculated for C ₂₄ H ₃₀ F ₂ N ₅ O ₄ , 490.2266. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 12.24 (s, 1H), 8.95 (d, J =8.0 Hz, 1H), 8.64 (d, J =8.0 Hz , 1H), 7.69 (s, 1H), 7.49-7.47 (m, 1H), 7.13 (dd, J =11.1, 11.1 Hz, 1H), 5.00- 4.93 (m, 1H), 4.54 (ddd, J =8.8, 4.1 Hz, 1H), 4.00 (s, 3H), 3.17-3.06 (m, 2H) , 2.38-2.30 (m, 1H), 2.18-2.07 (m, 2H), 1.85-1.65 (m, 4H), 0.95 (s, 9H). Retention time: 8.65 minutes (Analytical conditions identical to those used for 20 ).

Table 1. Method of synthesis, structure and physico-chemical data for examples 24-74. No. pr. Method of synthesis; Non-commercial source materials Structure ¹ H NMR (600 MHz, DMSO- d ₆ ) δ; Mass spectrum, observed ion m/z [M+H] ⁺ or HPLC retention time; Mass spectrum m/z [M+H] ⁺ (unless otherwise noted) 24 Example 10; C18

¹ H NMR (400 MHz, methanol- d ₄ ) δ 8.58 (s, 1H), 5.04 (dd, J =9.9, 6.1 Hz, 1H), 4.64 (dd, J = 8.7, 4.1 Hz, 1H), 3.34-3.23 (m, 2H; assumed; partially covered by solvent peak), 2.61-2.50 (m, 1H), 2.35-2 .24 (m, 2H), 1.96-1.76 (m, 4H), 1.01 (s, 9H); 460.4 25 Examples 8 and 9 ^1.2 ; C28

or

DIAST-1 2.29 minutes ³ ; 391.4 26 Examples 8 and 9 ^1.2 ; C28

or

DIAST-2 2.69 minutes ³ ; 391.4 27 Example 7; C28

2.18 minutes ⁴ ; 381.4 28 Example 7; C28

2.61 minutes ⁴ ; 421.5 29 Example 7; C28

2.31 minutes ⁴ ; 419.4 thirty Example 7 ¹ ; C28

1.59 minutes ⁴ ; 442.5 ⁵ 31 C29 ⁶

or

2.06 minutes ⁴ ; 416.5 32 C29 ⁶

or

2.67 minutes ⁴ ; 435.6 33 Example 4 ⁷

11.40 (s, 1H), 9.60 (brs, 1H), 8.90 (d, J = 8.1 Hz, 1H), 8.44 (d, J = 7.9 Hz, 1H), 7.70 (s, 1H), 7.31 (s, 1H), 6.95 (dd, J =8.8 Hz, 1H), 6.85 (d, J =8.2 Hz, 1H), 6.36 (d, J = 7.5 Hz, 1H), 5.02-4.93 (m, 1H), 4.50-4.41 (m, 1H), 3.17-3.07 ( m, 2H), 2.41-2.31 (m, 1H), 2.19-2.08 (m, 2H), 1.80 (ddd, J =13.6, 9.4, 6.8 Hz, 1H), 1.76-1.65 (m, 3H), 1.57-1.48 (m, 1H), 0.94 (d, J =6.3 Hz, 3H), 0.89 (d, J =6.3 Hz, 3H); High resolution MS m/z 426.2139 [M+H] ⁺ ; calculated for C ₂₂ H ₂₈ N ₅ O ₄ , 426.2141 34 Example 4 ⁷

11.34 (s, 1H), 8.91 (d, J = 8.1 Hz, 1H), 8.46 (d, J = 7.9 Hz, 1H), 7.69 (s, 1H), 7.29 (s, 1H), 6.95 (d, J = 8.6 Hz, 1H), 6.78 (d, J = 8.6 Hz, 1H), 5.02-4.93 (m , 1H), 4.50-4.41 (m, 1H), 3.91 (s, 3H), 3.18-3.06 (m, 2H, inferred; partially covered by solvent peak), 2.39- 2.31 (m, 1H), 2.20-2.07 (m, 2H), 1.84-1.65 (m, 4H), 1.58-1.49 (m, 1H), 0, 94 (d, J =6.2 Hz, 3H), 0.89 (d, J =6.2 Hz, 3H); High resolution MS m/z 456.2238 [M+H] ⁺ ; calculated for C ₂₃ H ₃₀ N ₅ O ₅ , 456.2247 35 Example 11 ⁸ ; C18

2.39 minutes ⁴ ; 437.4 (chlorine isotope pattern observed) 36 Example 11; C18

¹ H NMR (600 MHz, methanol- d ₄ ) δ 7.45-7.40 (m, 2H), 7.34-7.27 (m, 3H), 4.98 (dd, J =10.5 , 5.6 Hz, 1H), 4.27 (dd, J = 8.4, 4.3 Hz, 1H), 3.70 (s, 1H), 3.28-3.23 (m, 1H) , 3.20 (ddd, J =9.5, 9.3, 7.1 Hz, 1H), 2.57-2.49 (m, 1H), 2.27 (ddd, J =13.8, 10.5, 5.2 Hz, 1H), 2.21-2.13 (m, 1H), 2.17 (s, 6H), 1.85 (ddd, J =13.8, 9.7, 5.7 Hz, 1H), 1.79-1.70 (m, 1H), 1.67 (dd, ABX system component, J =14.4, 4.3 Hz, 1H), 1.59 (dd , component of the ABX system, J = 14.4, 8.4 Hz, 1H), 0.84 (s, 9H); 442.5 37 Examples 5 and 6 ⁹ ; Example 10

12.13 (s, 1H), 8.96 (bd, J = 7.6 Hz, 2H), 7.69 (s, 1H), 7.21 (dd, J = 8.8 Hz, 1H), 7.08 (d, J = 8.2 Hz, 1H), 6.68 (d, J = 7.8 Hz, 1H), 5.00-4.92 (m, 1H), 4.52-4 .45 (m, 1H), 3.86 (s, 3H), 3.20-3.15 (m, 1H), 3.14-3.08 (m, 1H), 2.43-2.34 (m, 1H), 2.23-2.13 (m, 2H), 1.85-1.78 (m, 1H), 1.78-1.68 (m, 2H), 1.59 (dd , J = 14.1, 6.6 Hz, 1H), 0.95 (s, 9H); High resolution MS m/z 522.2321 [M+H] ⁺ ; calculated for C ₂₅ H ₃₁ F ₃ N ₅ O ₄ , 522.2328; hold time 7.18 minutes ¹⁰ 38 Examples 5 and 6 ⁹ ; Example 10

11.41 (s, 1H), 8.96 (d, J = 7.7 Hz, 1H), 8.79 (d, J = 8.0 Hz, 1H), 7.65 (s, 1H), 7.55 (d, J = 8.2 Hz, 1H), 7.34 (s, 1H), 6.72 (d, J = 8.3 Hz, 1H), 5.01-4.94 (m , 1H), 4.62-4.55 (m, 1H), 3.97 (s, 3H), 3.17-3.11 (m, 1H), 3.11-3.04 (m, 1H ), 2.38-2.30 (m, 1H), 2.18-2.07 (m, 2H), 1.86-1.78 (m, 1H), 1.77-1.64 (m , 3H), 0.95 (s, 9H); High resolution MS m/z 522.2316 [M+H] ⁺ ; calculated for calculated for C ₂₅ H ₃₁ F ₃ N ₅ O ₄ , 522.2328; hold time 7.45 minutes ¹⁰ 39 Examples 5 and 6 ⁹ ; Example 10

9.08 (brs, 1H), 9.03 (d, J = 8.0 Hz, 1H), 7.65 (s, 1H), 7.60 (d, J = 8.3 Hz, 1H), 6.84 (d, J = 8.2 Hz, 1H), 4.98-4.91 (m, 1H), 4.53-4.47 (m, 1H), 3.95 (s, 3H) , 3.20-3.14 (m, 1H), 3.13-3.06 (m, 1H), 2.44-2.36 (m, 1H), 2.23-2.12 (m, 2H), 1.85 (dd, J =13.9, 7.8 Hz, 1H), 1.80 (dd, J =13.6, 9.6, 6.5 Hz, 1H), 1.76 -1.68 (m, 1H), 1.54 (dd, J =13.9, 5.1 Hz, 1H), 0.95 (s, 9H); High resolution MS m/z 590.2177 [M+H] ⁺ ; calculated for C ₂₆ H ₃₀ F ₆ N ₅ O ₄ , 590.2202; hold time 7.70 minutes ¹⁰ 40 Examples 5 and 6 ⁹ ; Example 10

12.76 (br, 1H), 9.19-9.10 (m, 1H), 9.06-9.00 (m, 1H), 7.69 (s, 1H), 7.49 (AB quartet , J _AB= 8.6 Hz, δν _AB= 48.9 Hz, 2H), 5.01-4.93 (m, 1H), 4.54-4.47 (m, 1H), 3.85 ( s, 3H), 3.21-3.15 (m, 1H), 3.15-3.07 (m, 1H), 2.43-2.35 (m, 1H), 2.22-2, 13 (m, 2H), 1.86-1.78 (m, 1H), 1.78-1.70 (m, 2H), 1.59 (dd, ABX system component, J =14.1, 6 .5 Hz, 1H), 0.96 (s, 9H); High resolution MS m/z 590.2181 [M+H] ⁺ ; calculated for C ₂₆ H ₃₀ F ₆ N ₅ O ₄ , 590.2202; hold time 7.79 minutes ¹⁰ 41 Examples 5 and 6 ^{9, 11} ; Example 10

12.62 (s, 1H), 9.14 (d, J =7.5 Hz, 1H), 9.03 (d, J =7.8 Hz, 1H), 7.70 (s, 1H), 7.42 (s, 1H), 6.92 (s, 1H), 5.00-4.93 (m, 1H), 4.54-4.47 (m, 1H), 3.96 (s, 3H), 3.21-3.15(m, 1H), 3.15-3.08(m, 1H), 2.43-2.35(m, 1H), 2.22-2.13( m, 2H), 1.86-1.79 (m, 1H), 1.78-1.69 (m, 2H), 1.59 (dd, component of the ABX system, J =14.0, 6.4 Hz, 1H), 0.96 (s, 9H); High resolution MS m/z 590.2175 [M+H] ⁺ ; calculated for C ₂₆ H ₃₀ F ₆ N ₅ O ₄ , 590.2202; hold time 7.84 minutes ¹⁰ 42 Examples 5 and 6 ¹² ; Example 36

8.92 (d, J =8.0 Hz, 1H), 8.45 (d, J =7.9 Hz, 1H), 7.71 (s, 2H), 7.67 (d, J =7 .6 Hz, 1H), 7.64-7.60 (m, 1H), 7.58-7.52 (m, 1H), 4.99-4.87 (m, 1H), 4.24- 4.17 (m, 1H), 3.92 (s, 1H), 3.20-3.10 (m, 1H), 3.09-3.00 (m, 1H), 2.36-2, 27 (m, 1H), 2.19-2.00 (m, 2H), 2.08 (s, 6H), 1.81-1.64 (m, 2H), 1.49 (d, J = 6.5 Hz, 2H), 0.74 (s, 9H); High resolution MS m/z 510.2679 [M+H] ⁺ ; calculated for C ₂₅ H ₃₅ F ₃ N ₅ O ₃ , 510.2692; hold time 5.83 minutes ¹⁰ 43 Examples 5 and 6 ¹² ; Example 36

8.93 (d, J = 8.1 Hz, 1H), 8.42 (d, J = 8.1 Hz, 1H), 7.71 (s, 1H), 7.63 (AB quartet, J _{AB =} 7.6 Hz, δν _AB= 48.8 Hz, 4H), 4.97-4.88 (m, 1H), 4.26-4.19 (m, 1H), 3.91 (s, 1H ), 3.19-3.11 (m, 1H), 3.07-3.00 (m, 1H), 2.35-2.26 (m, 1H), 2.18-1.99 (m , 2H), 2.08 (s, 6H), 1.80-1.64 (m, 2H), 1.56-1.44 (m, 2H), 0.77 (s, 9H); High resolution MS m/z 510.2676 [M+H] ⁺ ; calculated for C ₂₅ H ₃₅ F ₃ N ₅ O ₃ , 510.2692; hold time 5.92 minutes ¹⁰ 44 Alternative synthesis of example 6; C26

2.85 minutes ⁴ ; 478.6 45 Alternative synthesis of example 6; C18

2.95 minutes ⁴ ; 492.6 46 Examples 5 and 6 ¹³ ; 4

12.48 (s, 1H), 9.13 (d, J = 7.4 Hz, 1H), 8.97 (d, J = 7.7 Hz, 1H), 7.70 (s, 1H), 7.62 (d, J = 8.3 Hz, 1H), 6.86 (d, J = 8.3 Hz, 1H), 5.04-4.95 (m, 1H), 4.53-4 .44 (m, 1H), 3.96 (s, 3H), 3.22-3.14 (m, 1H), 3.14-3.06 (m, 1H), 2.44-2.34 (m, 1H), 2.24-2.10 (m, 2H), 1.87-1.77 (m, 1H), 1.77-1.61 (m, 2H), 1.61-1 .50 (m, 2H), 0.98-0.89 (m, 6H); High resolution MS m/z 576.2055 [M+H] ⁺ ; calculated for C ₂₅ H ₂₈ F ₆ N ₅ O ₄ , 576.2045; hold time 6.70 minutes ¹⁴ 47 Examples 5 and 6 ¹³ ; 4

12.61 (s, 1H), 9.23 (d, J = 7.6 Hz, 1H), 9.01 (d, J = 7.9 Hz, 1H), 7.71 (s, 1H), 7.20 (s, 1H), 5.04-4.95 (m, 1H), 4.54-4.45 (m, 1H), 4.07 (s, 3H), 3.22-3, 14 (m, 1H), 3.14-3.06 (m, 1H), 2.43-2.34 (m, 1H), 2.23-2.11 (m, 2H), 1.82 ( ddd, J = 13.5, 9.0, 7.0 Hz, 1H), 1.78-1.61 (m, 2H), 1.61-1.49 (m, 2H), 0.99- 0.89 (m, 6H); High resolution MS m/z 644.1914 [M+H] ⁺ ; calculated for C ₂₆ H ₂₇ F ₉ N ₅ O ₄ , 644.1919; hold time 7.43 minutes ¹⁴ 48 Examples 5 and 6 ¹³ ; 4

13.21 (s, 1H), 9.28 (d, J =7.5 Hz, 1H), 9.05 (d, J =7.8 Hz, 1H), 7.86 (s, 1H), 7.72 (s, 1H), 5.04-4.96 (m, 1H), 4.54-4.47 (m, 1H), 3.93 (s, 3H), 3.22-3, 15 (m, 1H), 3.15-3.07 (m, 1H), 2.43-2.35 (m, 1H), 2.23-2.11 (m, 2H), 1.82 ( ddd, J = 13.7, 9.0, 6.9 Hz, 1H), 1.78-1.63 (m, 2H), 1.61-1.51 (m, 2H), 0.98- 0.91 (m, 6H); High resolution MS m/z 644.1901 [M+H] ⁺ ; calculated for C ₂₆ H ₂₇ F ₉ N ₅ O ₄ , 644.1919; hold time 7.51 minutes ¹⁴ 49 Alternative synthesis of example 6; C26

2.95 minutes ⁴ ; 494.4 fifty Alternative synthesis of example 6; C18

3.04 minutes ⁴ ; 508.4 51 Alternative synthesis of example 6; C18

3.00 minutes ⁴ ; 492.4 52 Alternative synthesis of example 6; C18

2.84 minutes ⁴ ; 458.4 (chlorine isotope pattern observed) 53 Alternative synthesis of example 6; C18

2.80 minutes ⁴ ; 468.5 54 Alternative synthesis of example 6; C18

2.89 minutes ⁴ ; 458.4 (chlorine isotope pattern observed) 55 Alternative synthesis of example 6; C18

2.90 minutes ⁴ ; 458.4 (chlorine isotope pattern observed) 56 Alternative synthesis of example 6; C18

2.89 minutes ⁴ ; 458.4 (chlorine isotope pattern observed) 57 Alternative synthesis of example 6; C18

2.95 minutes ⁴ ; 492.4 58 Alternative synthesis of example 6; C18

3.16 minutes ⁴ ; 492.3 (dichlorine isotope pattern observed) 59 Alternative synthesis of example 6; C18

2.99 minutes ⁴ ; 492.4 60 Alternative synthesis of example 6; C26

2.91 minutes ⁴ ; 478.4 61 Alternative synthesis of example 6; C26

2.74 minutes ⁴ ; 444.4 (chlorine isotope pattern observed) 62 Alternative synthesis of example 6; C26

2.70 minutes ⁴ ; 454.4 63 Alternative synthesis of example 6; C26

2.80 minutes ⁴ ; 444.4 (chlorine isotope pattern observed) 64 Alternative synthesis of example 6; C26

2.81 minutes ⁴ ; 444.4 (chlorine isotope pattern observed) 65 Alternative synthesis of example 6; C26

2.80 minutes ⁴ ; 444.4 (chlorine isotope pattern observed) 66 Alternative synthesis of example 6; C26

2.86 minutes ⁴ ; 478.4 67 Alternative synthesis of example 6; C26

3.08 minutes ⁴ ; 478.3 (dichlorine isotope pattern observed) 68 Alternative synthesis of example 6; C26

2.91 minutes ⁴ ; 478.4 69 Examples 5 and 6 ¹⁵ ; 7

9.00 (d, J =7.9 Hz, 1H), 8.83 (d, J =7.9 Hz, 1H), 8.01 (s, 1H), 7.72 (s, 1H), 5.01-4.93 (m, 1H), 4.44 (ddd, J =8, 8, 4.4 Hz, 1H), 3.20-3.14 (m, 1H), 3.14- 3.08 (m, 1H), 2.66 (s, 3H), 2.42-2.34 (m, 1H), 2.20-2.08 (m, 2H), 1.81 (ddd, J = 13.5, 9.4, 6.7 Hz, 1H), 1.77-1.64 (m, 3H), 0.95 (s, 9H); High resolution MS m/z 513.1873 [M+H] ⁺ ; calculated for C ₂₂ H ₂₈ F ₃ N ₆ O ₃ S, 513.1896; hold time 6.88 minutes ¹⁰ 70 Alternative synthesis of example 6; C18

2.87 minutes ⁴ ; 474.5 71 Alternative synthesis of example 6; C18

2.70 minutes ⁴ ; 474.5 72 Alternative synthesis of example 6; C26

2.41 minutes ⁴ ; 481.5 (bromine isotope pattern observed) 73 Alternative synthesis of example 6; C26

2.30 minutes ⁴ ; 423.5 (chlorine isotope pattern observed) 74 4 ¹⁶

12.20 (s, 1H), 8.99 (d, J = 7.9 Hz, 1H), 8.97 (d, J = 7.4 Hz, 1H), 7.70 (s, 1H), 7.24 (dd, J = 8.8 Hz, 1H), 7.12 (d, half AB quartet, J = 8.2 Hz, 1H), 6.69 (d, J = 7.8 Hz, 1H ), 5.01-4.93 (m, 1H), 4.47-4.41 (m, 1H), 3.89 (s, 3H), 3.18-3.08 (m, 2H), 2.57 (s, 3H), 2.40-2.32 (m, 1H), 2.19-2.09 (m, 2H), 1.81 (ddd, J =13.5, 9.3 , 6.8 Hz, 1H), 1.75-1.63 (m, 2H), 1.59 (dd, J = 7.5, 6.8 Hz, 2H), 0.95 (d, J = 6.5 Hz, 3H), 0.91 (d, J =6.5 Hz, 3H); High resolution MS m/z 482.2391 [M+H] ⁺ ; calculated for C ₂₅ H ₃₂ N ₅ O ₅ , 482.2403; hold time 8.38 minutes ¹⁰

1. In this case, C28 is deprotected using methanesulfonic acid rather than hydrochloride.

2. The epimers of example 25 and example 26 are separated by supercritical liquid chromatography (Column: Chiral Technologies Chiralpak IB, 21×250 mm, 5 µm; Mobile phase: 9:1 carbon dioxide/methanol; Back pressure: 120 bar, Flow rate: 75 ml /minute). The first eluted diastereomer is designated Example 25 and the second eluted diastereomer is designated Example 26.

3. Conditions for analytical HPLC. Column: Chiral Technologies Chiralpak IB, 4.6×100 mm, 5 µm; Mobile phase: 85:15 carbon dioxide/methanol; Back pressure: 120 bar; Flow rate: 1.5 ml/minute.

4. Conditions for analytical HPLC. Column: Waters Atlantis dC18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v); Gradient: 5.0% - 95% B, linear over 4.0 minutes, then 95% B over 1.0 minutes; Flow rate: 2 ml/minute.

5. ¹ H NMR Example 30 to final purification: ¹ H NMR (400 MHz, methanol- d ₄ ) δ 7.48-7.42 (m, 2H), 7.36-7.26 (m, 3H), 4.90 (dd, J =10.5, 5.7 Hz, 1H), 4.37 (dd, J =7.7, 4.9 Hz, 1H), 3.69 (s, 1H), 3 .25 (ddd, J =9.9, 8.9, 2.5 Hz, 1H), 3.18 (ddd, J =9.6, 9.0, 7.1 Hz, 1H), 2.40 -2.29(m, 1H), 2.20(s, 6H), 2.2-2.10(m, 1H), 2.09-1.99(m, 1H), 1.80-1 .61 (m, 2H), 1.73 (dd, J =14.5, 5.0 Hz, 1H), 1.61 (dd, J =14.4, 7.8 Hz, 1H), 0, 95 (s, 9H).

6. Amide coupling with a suitable carboxylic acid is carried out using 2,4,6-trioxide 2,4,6-tripropyl-1,3,5,2,4,6-trioxatrifosfinan trioxide.

7. Example 4 (25 µm) incubated with human cytochrome P450 3A5 (4 nmol) in potassium phosphate buffer (100 mM, pH 7.4; 40 ml) containing magnesium chloride (3.3 mM) and NADPH (1.3 mM). Incubation is carried out for 0.75 hours in a shaking water bath maintained at 37°C. The incubation is terminated by adding an equal volume of acetonitrile, then the mixture is spun in a centrifuge at 1700 x g for 5 minutes and the supernatant is subjected to vacuum centrifugation for approximately 1.5 hours. To this mixture are added formic acid (0.5 ml), acetonitrile (0.5 ml) and water to a final volume of 50 ml, and the resulting mixture is centrifuged at 40,000 x g for 30 minutes. The supernatant was subjected to reverse phase HPLC (Column: Polaris C18, 4.6×250 mm; 5 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: methanol; Gradient: 15% B over 5 minutes, then 15% - 35% B for 75 minutes, then 35% - 95% B for 10 minutes; Flow rate: 0.8 ml/minute). Fractions are collected every 20 seconds. the first eluted product, not pure Example 33, was eluted at 54.7 minutes and Example 34 was eluted at 55.3 minutes. Illegal Example 33 was repurified using reverse phase HPLC (Column: Phenomenex Kinetex XB-C18, 2.1×100 mm, 2.6 µm; Mobile phase A: water containing 0.5% acetic acid; Mobile phase B : 9:1 acetonitrile/methanol; Gradient: 10% B over 0.5 minutes, then 10% - 35% over 26.5 minutes, then 35% - 60% B over 3 minutes; Flow rate 0.5 ml/minute); fractions are collected every 15 seconds. In this system, example 33 has a hold time of 12.7 minutes; Additional Example 34 was eluted at 13.5 minutes.

8. The desired 4-chloro-1,3-dimethyl-1 H -pyrazole-5-carboxylic acid can be obtained by hydrolysis of commercially available ethyl ester.

9. The reaction mixture is diluted with acetonitrile and 1% aqueous formic acid to a volume of approximately 2 ml; the composition of the final solvent is such that the mixture is clear, with an acetonitrile content of approximately 20% - 30%. The components of this mixture are separated by reverse phase HPLC (Column: Phenomenex Luna C18, 10×250 mm, 10 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile; Gradient: 15% B over 5 minutes, then 15% - 70% B for 70 minutes, then 70% - 95% B for 15 minutes; Flow rate: 2 ml/minute); fractions are collected every 20 seconds. Examples 37, 38, 39, 40, and 41 are eluted with the retention times given below.

Example Hold time (minutes) 37 64.9 38 68.4 39 72.1 40 73.5 41 74.2

10. Conditions for analytical HPLC. Column: Phenomenex Kinetex XB-C18, 2.1×100 mm, 2.6 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile; Gradient: 5% B over 0.5 minutes, then 5% - 70% B over 10.5 minutes, then 70% - 95% B over 2 minutes; Flow rate: 0.4 ml/min.

11. The regiochemistry of example 41 was not strictly defined; other possible structures for this example are N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4 ,4-dimethyl-1-oxopentan-2-yl]-4-methoxy-5,6-bis(trifluoromethyl)-1 H -indole-2-carboxamide and N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-4-methoxy-6,7 -bis(trifluoromethyl)-1 H -indole-2-carboxamide.

12. The reaction mixture was purified using the conditions described in Note 9. Example 42 was eluted at 58.1 minutes and Example 43 was eluted at 59.2 minutes.

13. The reaction mixture was diluted with a mixture of acetonitrile (0.3 ml) and 1% aqueous formic acid (0.7 ml). The resulting mixture was centrifuged and the supernatant was subjected to reverse phase HPLC (Column: Phenomenex Luna C18, 10×250 mm, 10 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile; Gradient: 2% - 10% B for 5.0 minutes, then 10% - 95% B for 95 minutes; Flow rate: 2 ml/minute); fractions are collected every 20 seconds. Examples 46, 47, and 48 are eluted with the retention times given below. Example 5 is also isolated from this reaction, in fractions 189-190.

Example Fraction number 46 207 47 225-226 48 231-232

14. Conditions for analytical HPLC. Column: Phenomenex Kinetex C18, 2.1×50 mm, 1.7 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile containing 0.1% formic acid; Gradient: 5% B over 0.5 minutes, then 5% - 50% B over 6.0 minutes, then 50% - 80% B over 1.5 minutes, then 80% - 95% B over 1 .0 minutes; Flow rate: 0.4 ml/min.

15. Only the indicated product is observed in this reaction.

16. Prepare a stock solution of Example 4 (5.56 mg, 12.7 µmol) and trifluoroacetic acid (4 µl, 50 µl) in dimethyl sulfoxide (420 µl). One sixth of this solution is treated with sodium 1,1-difluoroethanesulfinate 1.3 mg, 8.5 µmol) then tert -butyl hydroperoxide (70% in water; 1.4 µl, 10 µmol) and heated at 50° C. overnight. The reaction mixture is diluted with acetonitrile and 1% aqueous formic acid to a volume of approximately 2-3 ml; the composition of the final solvent is such that the mixture is clear, with an acetonitrile content of approximately 20% - 30%. The components of this mixture are separated by reverse phase HPLC (Column: Phenomenex Luna C18, 10×250 mm, 10 µm; Mobile phase A: water containing 0.1% formic acid; Mobile phase B: acetonitrile; Gradient: 15% B over 5 minutes, then 15% - 40% B for 70 minutes, then 40% - 95% B for 15 minutes; Flow rate: 2 ml/minute); fractions are collected every 20 seconds. Example 74 was eluted at 68.6 minutes.

Examples 75 and 76

( 2S , 4R )-4- tert -Butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1-{ N- [(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide and ( 2R , 4S )-4- tert -Butyl- N -{( 1S )-1-cyano-2-[( 3S ) -2-oxopyrrolidin-3-yl]ethyl}-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide [ 75 (DIAST-1) and 76 (DIAST-2)

Step 1. Synthesis of tert -butyl N -[(trifluoromethyl)sulfonyl]-L-valinate ( C44 ).

A solution of trifluoromethanesulfonic anhydride (8.88 ml, 52.8 mmol) in dichloromethane (10 ml) was added to a -78°C solution of tert -butyl L-valinate, hydrochloride (10.0 g, 47.7 mmol) and triethylamine (18 .7 ml, 134 mmol) in dichloromethane (90 ml). The reaction mixture is stirred at -78°C for 2 hours, then it is poured into water and acidified to pH approximately 4 by adding 1 M hydrochloric acid. The resulting mixture was extracted with dichloromethane and the organic layer was washed with aqueous sodium bicarbonate and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . The residue is combined with the products of two similar reactions carried out using tert -butyl L-valinate, hydrochloride (1.00 g, 4.77 mmol; 1.00 g, 4.77 mmol) and purified by silica gel chromatography (gradient: 0% - 20% ethyl acetate in petroleum ether) to give C44 as a white solid. Combined yield: 14.0 g, 45.9 mmol, 80%. ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.92 (d, J =8.8 Hz, 1H), 3.68 (dd, J =8.8, 6.2 Hz, 1H), 2 .16-2.02 (m, 1H), 1.43 (s, 9H), 0.92 (d, J = 6.9 Hz, 3H), 0.90 (d, J = 6.9 Hz, 3H).

Step 2. Synthesis of N -[(trifluoromethyl)sulfonyl]-L-valine ( C45 ).

To a solution of C44 (14.0 g, 45.9 mmol) in dichloromethane (85 ml) was added trifluoroacetic acid (85 ml). The reaction mixture is stirred at room temperature for 3 hours, then it is concentrated in vacuo ; the residue is washed with petroleum ether to give C45 as a white solid. Yield: 10.9 g, 43.7 mmol, 95%. MS m/z 248.0 [MH] ^- . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.86 (br, J = 8.3 Hz, 1H), 3.79-3.71 (m, 1H), 2.19-2.05 ( m, 1H), 0.93 (d, J =6.8 Hz, 3H), 0.90 (d, J =6.8 Hz, 3H).

Step 3. Synthesis of methyl cis -4- tert -butylpiperidine-2-carboxylate, hydrochloride ( C46 ).

To a 0°C solution of cis -4- tert -butylpiperidine-2-carboxylic acid, hydrochloride (see RT Shuman et al., J. Org. Chem . 1990, 55 , 738-741; 4.00 g, 18.0 mmol) in methanol (40 ml) thionyl chloride (6.44 g, 54.1 mmol) is added. The reaction mixture was then stirred at 25° C. for 16 hours, concentrated in vacuo to give C46 as an off-white solid (4.50 g). Part of this product is used in the next step. LCMS m/z 200.0 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.46 (br s, 1H), 9.09 (br br, 1H), 4.11-3.96 (m, 1H), 3.76 (s, 3H), 3.4-3.21 (m, 1H, estimated; significantly covered by water peak), 2.93-2.77 (m, 1H), 2.07 (br, J =10.8 Hz, 1H ), 1.75 (bd, J =10.6 Hz, 1H), 1.51-1.32 (m, 3H), 0.84 (s, 9H).

Stage 4. Synthesis of methyl (2 S ,4 R )-4- tert -butyl-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-caroxylate and methyl (2 R ,4 S )- 4- tert -Butyl-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-caroxylate ( C47 ).

4- methylmorpholine ( 365 mg , 3 .61 mmol). The resulting mixture was cooled to 0° C. and treated with O- (7-azabenzotriazol-1-yl) -N, N, N', N'- tetramethyluronium hexafluorophosphate (GATU; 549 mg, 1.44 mmol). Then the reaction mixture is purged with nitrogen for 1 minute, it is stirred at 25°C for 12 hours. LCMS analysis at this point shows the presence of C47 : LCMS m/z 431.1 [M+H] ⁺ . The reaction mixture was partitioned between ethyl acetate (20 ml) and water (20 ml) and the aqueous layer was extracted with ethyl acetate (20 ml). The combined organic layers are washed with saturated aqueous sodium chloride (4×20 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (gradient: 0% - 20% ethyl acetate in petroleum ether) to give C47 as a yellow gum. ¹ H NMR analysis confirmed that the product contained a mixture of diastereomers. Yield: 320 mg, 0.743 mmol, 62%. ¹ H NMR (400 MHz, chloroform- d ) δ 6.12-5.94 (m, 1H), [4.51 (dd, J =11.7, 6.3 Hz) and 4.32-4, 18 (m), 2H total], [3.73 (s) and 3.71 (s), 3H total], [3.63-3.49 (m) and 3.48-3.39 (m) , total 2H], 2.18-1.93 (m, 2H), 1.91-1.77 (m, 1H), 1.63-1.37 (m, 2H), 1.37-1, 22 (m, 1H), 1.13-1.04 (m, 3H), [0.94 (d, J =6.8 Hz) and 0.91 (d, J =6.8 Hz), total 3H], 0.87 (s, 9H).

Step 5. Synthesis of ( 2S , 4R )-4- tert -butyl-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxylic acid and ( 2R , 4S )-4 - tert -Butyl-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxylic acid ( C48 ).

A solution of C47 (314 mg, 0.729 mmol) in a mixture of methanol (2 ml) and tetrahydrofuran (2 ml) is treated with a solution of lithium hydroxide monohydrate (91.8 mg, 2.19 mmol) in water (1.4 ml) and the reaction mixture is stirred at 25°C for 3 hours. After removal of the solvent in vacuo, the residue was diluted with water (10 ml) and acidified to pH approximately 1 with 1 M hydrochloric acid. The resulting mixture was extracted with ethyl acetate (2 x 20 ml) and the combined organic layers were washed with saturated aqueous sodium chloride (15 ml), dried over sodium sulfate, filtered, and concentrated in vacuo to give C48 as a yellow glass. ¹ H NMR analysis confirmed that the product contained a mixture of diastereomers. Yield: 304 mg, quantitative. ¹ H NMR (400 MHz, DMSO- d ₆ ) δ [9.82 (d, J = 8.7 Hz) and 9.69 (bd, J = 8.8 Hz), total 1H], [4.28 (dd, J =11.5, 6.4 Hz), 4.24-4.14 (m), and 4.05-3.96 (m), total 2H], [3.80-3.69 (m) and 3.6-3.2 (m, estimated; essentially covered by water peak), total 2H], 2.06-1.90 (m, 2H), 1.80-1.65 (m, 1H), 1.41-1.17 (m, 3H), [0.96 (d, J =6.8 Hz) and 0.93 (d, J =6.5 Hz), total 3H], [ 0.89 (d, J = 6.9 Hz) and 0.86-0.80 (m), total 12H].

Stage 6. Synthesis of (2 S ,4 R )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2-yl} -4- tert -Butyl-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide and ( 2R , 4S ) -N -{( 2S )-1-amino-1 -oxo-3-[( 3S )-2-oxopyrrolidin-3-yl]propan-2-yl}-4- tert -butyl-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine- 2-carboxamide ( C49 ).

To a 25°C mixture of C16 (120 mg, 0.449 mmol) and C48 (144 mg, 0.346 mmol) in N,N -dimethylformamide (3 ml) was added 4-methylmorpholine (100 mg, 0.989 mmol), then the mixture was cooled to 0° C and treated with O- (7-azabenzotriazol-1-yl) -N,N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 151 mg, 0.397 mmol). The reaction mixture was purged with nitrogen for 1 minute and then stirred at 25° C. for 12 hours. LCMS analysis shows the presence of C49 : LCMS m/z 570.3 [M+H] ⁺ . The reaction mixture was then partitioned between ethyl acetate (20 ml) and water (20 ml) and the aqueous layer was saturated with solid sodium chloride and extracted with ethyl acetate (5×20 ml). The combined organic layers are dried over sodium sulfate, filtered, concentrated in vacuo , and chromatographed on silica gel (gradient: 0% - 15% methanol in dichloromethane) to give C49 as a white solid. This product contains a mixture of diastereomers as determined by ¹ H NMR. Yield: 190 mg, 0.334 mmol, 96%. ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks, integrations are approximate: δ [9.88 (d, J =8.6 Hz) and 9.82-9.68 (m), total 1H] , [8.12 (d, J = 8.8 Hz) and 8.09-7.98 (m), total 1H], [7.63 (s) and 7.57 (s), total 1H], [7.30 (shs) and 7.18 (shs), total 1H], [7.06 (shs) and 7.03 (shs), total 1H], [4.36 (dd, J = 12.0 , 6.1 Hz) and 4.32-4.08 (m), total 2H], 2.26-2.05 (m, 2H), 1.81-1.54 (m, 2H), 1, 53-1.30 (m, 2H), 0.98-0.87 (m, 6H), 0.86-0.76 (m, 9H).

Step 7. Synthesis of ( 2S , 4R )-4- tert -butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1 -{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide and ( 2R , 4S )-4- tert -butyl- N -{( 1S )-1-cyano-2-[ (3 S )-2-oxopyrrolidin-3-yl]ethyl}-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide [ 75 (DIAST-1) and 76 (DIAST-2 ) ].

A mixture of C49 (190.0 mg, 0.334 mmol) and methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 238 mg, 1.00 mmol) in dichloromethane (10 mL) was stirred at 25°C for 2 days, then the reaction mixture was diluted with water (20 ml) and extracted with dichloromethane (2×20 ml). The combined organic layers are washed with saturated aqueous sodium chloride (10 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (gradient: 0% - 8% methanol in dichloromethane) gives a white solid, which according to LCMS analysis contains approximately 3:1 mixture of products: LCMS m/z 552.2 [M+H] ⁺ and LCMS m/z 552.2 [M+H] ⁺ . These diastereomers are separated by supercritical liquid chromatography [Column: Chiral Technologies Chiralpak IG, 30×250 mm, 10 µm; Mobile phase: 3:1 carbon dioxide/(ethanol containing 0.1% ammonium hydroxide); flow rate: 70 ml/minute]. The first eluted diastereomer, isolated as a white solid, is designated 75 and the second eluted diastereomer, also a white solid, is designated 76 [( 2S , 4R )-4- tert -butyl- N -{( 1S ) -1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1-{ N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide and ( 2R , 4 S )-4- tert -butyl- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-1-{ N -[(trifluoromethyl) sulfonyl]-L-valyl}piperidine-2-carboxamide].

75- Yield: 26.2 mg, 47.5 µmol, 14%.^oneH NMR (400 MHz, DMSO-d ₆) δ 9.87 (d,J= 8.8 Hz, 1H), 8.87 (d,J= 8.4 Hz, 1H), 7.70 (s, 1H), 4.99-4.91 (m, 1H), 4.24 (dd,J=12.3, 6.0 Hz, 1H), 4.18 (dd,J= 8.3, 8.3 Hz, 1H), 3.88-3.78 (m, 1H), 3.19-3.00 (m, 2H), 2.46-2.35 (m, 1H ), 2.17-2.02 (m, 2H), 1.99-1.85 (m, 2H), 1.79-1.62 (m, 3H), 1.50-1.36 (m , 2H), 1.26-1.12 (m, 2H), 0.97-0.87 (m, 6H), 0.84 (s, 9H). Retention time: 1.30 minutes (Analytical conditions. Column: Chiral Technologies Chiralpak IG-3, 4.6×50 mm, 3 µm; Mobile phase A: carbon dioxide; Mobile phase B: ethanol containing 0.05% diethylamine; gradient : 5% - 40% B for 2 minutes, then 40% B for 1.2 minutes; Flow rate: 4 ml/minute; Back pressure: 1500 psi).

76- Yield: 8.8 mg, 16 µmol, 5%. HCMSm/z 552.3 [M+H]⁺. By^oneH NMR analysis, this sample76 contains impurities.^oneH NMR (400 MHz, DMSO-d ₆), characteristic peaks, integrations are approximate: δ 9.76 (d,J= 8.8 Hz, 1H), 8.59 (d,J= 8.1 Hz, 1H), 7.72 (s, 1H), 5.02-4.90 (m, 1H), 0.94-0.86 (m, 6H), 0.82 (s, 9H). Retention time: 1.61 minutes (Assay conditions are identical to those used for 7five).

Example 77

3-Methyl- N- (trifluoroacetyl)-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}- 4-(trifluoromethyl)-L-prolinamide ( 77 )

Step 1. Synthesis of (4 R )-1-( tert -butoxycarbonyl)-4-(trifluoromethyl)-L-prolyl-3-[(3 S )-2-oxopyrrolidin-3-yl]-L-alanineamide ( C50 ) .

K -30°C mixture of ( 4R )-1-( tert -butoxycarbonyl)-4-(trifluoromethyl)-L-proline (429 mg, 1.51 mmol) and C16, HCl salts (346 mg, 1.67 mmol ) in N, N -dimethylformamide (7.8 ml) add N, N -diisopropylethylamine (0.791 ml, 4.54 mmol), then hexafluorophosphate O - (7-azabenzotriazol-1-yl) - N, N, N ', N' -tetramethyluronium (GATU; 633 mg, 1.66 mmol). The reaction mixture is heated to 0°C for 1 hour, then it is diluted with aqueous sodium bicarbonate solution (30 ml) and extracted with a mixture of 2-butanol and dichloromethane (9:1, 3.

ml). The combined organic layers were concentrated in vacuo and purified by silica gel chromatography (Gradient: 0% - 100% methanol in dichloromethane) to give C50 as an off-white foam. According to ¹ H NMR analysis, this product exists as a mixture of rotamers, and contains impurities derived from the reagents used; part of this sample is transferred to the next stage. Yield: 613 mg, 1.40 mmol, 93%. LCMS m/z 459.3 [M+Na ⁺ ]. ¹ H NMR (400 MHz, DMSO- d ₆ ), product characteristic peaks only: δ 8.33-8.18 (m, 1H), [7.65 (br) and 7.59 (br) 1H total] , [7.39 (brs) and 7.27 br (s), total 1H], 7.05 (brs, 1H), 4.38-4.28 (m, 1H), 4.28-4.17 (m, 1H), 3.46-3.36 (m, 1H), 2.02-1.89 (m, 1H), 1.80-1.45 (m, 2H), [1.39 ( c) and 1.32 (c), total 9H].

Step 2. Synthesis of N- ( tert -butoxycarbonyl)-3-methyl-L- valyl- (4R)-4-(trifluoromethyl)-L-prolyl-3-[( 3S )-2-oxopyrrolidin-3-yl ]-L-alanine amide ( C51 ).

A mixture of C50 (242 mg, 0.554 mmol) and a solution of hydrochloride in 1,4-dioxane (4 M; 2 ml, 8 mmol) was stirred at room temperature for 5 minutes, then the reaction mixture was concentrated in vacuo to remove the solvent and residual hydrochloride. The resulting deprotected product is combined with N- ( tert -butoxycarbonyl)-3-methyl-L-valine (128 mg, 0.553 mmol) and hexafluorophosphate O- (7-azabenzotriazol-1-yl) -N, N, N', N' -tetramethyluronium (GATU: 232 mg, 0.610 mmol) in N,N -dimethylformamide (2 ml) and then cooled to -30°C. N,N -diisopropylethylamine (0.290 ml, 1.66 mmol) is added and the reaction mixture is heated to 0° C. for 1 hour. After an aqueous sodium bicarbonate solution was added, the resulting mixture was extracted three times with ethyl acetate; the combined organic layers were concentrated in vacuo and purified by silica gel chromatography (gradient: 0% - 30% methanol in dichloromethane) to give C51 as a solid. Yield: 230 mg, 0.418 mmol, 75%. LCMS m/z 550.3 [M+H] ⁺ .

Step 3. Synthesis of 3-methyl- N- (trifluoroacetyl)-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl ]ethyl}-4-(trifluoromethyl)-L-prolinamide ( 77 ).

A mixture of C51 (230 mg, 0.418 mmol) and a solution of hydrochloride in 1,4-dioxane (4 M; 2 ml, 8 mmol) was stirred at room temperature for 5 minutes, then the reaction mixture was concentrated in vacuo to remove the solvent and residual hydrochloride. The resulting deprotected product is combined with ethyl trifluoroacetate (595 mg, 4.19 mmol) and N,N -diisopropylethylamine (0.219 ml, 1.26 mmol) in methanol (1.0 ml). The reaction mixture is then stirred at room temperature for 30 minutes, ethyl trifluoroacetate (60 mg, 0.422 mmol) is added again and stirring is continued for 30 minutes. An aqueous solution of sodium bicarbonate is then added and the resulting mixture is extracted three times with ethyl acetate. The combined organic layers are dried over magnesium sulfate, filtered, concentrated in vacuo and dissolved in dichloromethane (3 ml). Ethyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 299 mg, 1.25 mmol) is added thereto and the reaction mixture is stirred at room temperature for 2 hours, then it is treated with additional methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 100 mg, 0.420 mmol) and stirred for an additional 30 minutes. Hatem dilute aqueous sodium carbonate solution is added and the mixture is extracted twice with ethyl acetate; the combined organic layers are dried over magnesium sulfate, filtered, and concentrated in vacuo . Purification by supercritical fluid chromatography (Column: Princeton Dinitrophenyl, 10 x 250 mm, 5 µm; Mobile phase: 9:1 carbon dioxide/methanol; Back pressure: 120 bar; Flow rate: 80 ml/minute) gives a product which is then suspended in heptane (2.0 ml) at 50° C. for 2 hours, cooled to room temperature and collected by filtration to give 3-methyl- N- (trifluoroacetyl)-L- valyl- (4R) -N -{(1 S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-4-(trifluoromethyl)-L-prolinamide ( 77 ) as a solid. Yield: 64 mg, 0.121 mmol, 29%. LCMS m/z 528.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.46 (d, J =8.4 Hz, 1H), 9.05 (d, J =8.6 Hz, 1H), 7.67 (s , 1H), 4.96 (ddd, J = 11.0, 8.5, 5.0 Hz, 1H), 4.56 (d, J = 8.5 Hz, 1H), 4.37 (dd, J = 7.5, 7.5 Hz, 1H), 3.98 (dd, ABX system component, J = 11.2, 7.5 Hz, 1H), 3.92 (dd, ABX system component, J = 11.3, 4.8 Hz, 1H), 3.46-3.35(m, 1H), 3.19-3.10(m, 1H), 3.09-3.00(m, 1H) , 2.5-2.38 (m, 1H, estimated; partially covered by solvent peak), 2.38-2.28 (m, 1H), 2.21-2.04 (m, 3H), 1.78 -1.65 (m, 2H), 0.99 (s, 9H).

Example 78

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 78 )

Stage 1. Synthesis of methyl (1 R ,2 S ,5 S )-6,6-dimethyl-3-(3-methyl-L-valyl)-3-azabicyclo[3.1.0]hexane-2-caroxylate, hydrochloride ( C52 ).

To a 0°C solution of C31 (1.00 g, 2.61 mmol) in dichloromethane (20 ml) was added dropwise a solution of hydrochloride in ethyl acetate (4 M; 20 ml, 80 mmol). The reaction mixture is then stirred at 25° C. overnight, concentrated in vacuo to give C52 as a white gum. Yield: 700 mg, 2.20 mmol, 84%. LCMS m/z 283.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.22 (br s, 3H), 4.25 (s, 1H), 3.87-3.77 (m, 2H), 3.72 (d, half AB quartet, J = 10.8 Hz, 1H), 3.67 (s, 3H), 1.59 (dd, component of the ABX system, J = 7.7, 5.3 Hz, 1H), 1.49 (d, half AB quartet, J = 7.7 Hz, 1H), 1.03 (s, 9H), 1.02 (s, 3H), 0.96 (s, 3H).

Step 2. Synthesis of methyl (1 R ,2 S ,5 S )-6,6-dimethyl-3-[3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0] hexane-2-carboxylate ( C53 ).

To a 0° C. solution of C52 (320 mg, 1.00 mmol) in dichloromethane (6 ml) was slowly added triethylamine (0.769 ml, 5.52 mmol) and methylcarbamyl chloride (188 mg, 2.01 mmol). The reaction mixture is heated to 20°C and stirred for 18 hours, then it is treated dropwise with saturated aqueous sodium carbonate solution (5 ml) and extracted with dichloromethane (2×5 ml). The combined organic layers are washed with saturated aqueous sodium chloride (2×20 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane) to give C53 as a light yellow gum. Yield: 190 mg, 0.560 mmol, 56%. LCMS m/z 339.9 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 6.03 (d, J =9.4 Hz, 1H), 5.89 (shw, J =5 Hz, 1H), 4.20-4.14 (m, 2H), 3.91 (d, half AB quartet, J = 10.3 Hz, 1H), 3.79 (dd, component of the ABX system, J = 10.3, 5.3 Hz, 1H), 3.65 (s, 3H), 3.17 (d, J = 5.3 Hz, 3H), 1.55-1.49 (m, 1H), 1.40 (d, half AB quartet, J = 7.4 Hz, 1H), 1.00 (s, 3H), 0.92 (s, 9H), 0.83 (s, 3H).

Step 3. Synthesis of (1 R ,2 S ,5 S )-6,6-dimethyl-3-[3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane -2-carboxylic acid ( C54 ).

To a 0°C solution of C53 (190 mg, 0.560 mmol) in a mixture of tetrahydrofuran (2 ml), water (4 ml) and methanol (1 ml) was added lithium hydroxide monohydrate (82.0 mg, 1.95 mmol). Then the reaction mixture is stirred at 20°C for 2 hours, it is diluted with ethyl acetate (10 ml); the aqueous layer is then cooled to 0°C - 5°C and acidified to pH 2-3 by adding 1 M hydrochloric acid. The aqueous mixture was extracted with ethyl acetate (3 x 15 mL) and the combined ethyl acetate layers were dried over sodium sulfate, filtered, and concentrated in vacuo to give C54 as a white solid. Yield: 120 mg, 0.369 mmol, 66%. LCMS m/z 348.3 [M+Na ⁺ ]. ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 6.04 (d, J =9.6 Hz, 1H), 5.89 (d, J =4.7 Hz, 1H), 4 .17 (d, J = 9.6 Hz, 1H), 4.09 (s, 1H), 3.87 (d, half AB quartet, J = 10.4 Hz, 1H), 3.77 (dd, ABX system component, J = 10.3, 5.4 Hz, 1H), 1.49 (dd, ABX system component, J = 7.6, 5.1 Hz, 1H), 1.38 (d, half AB quartet, J = 7.5 Hz, 1H), 1.00 (s, 3H), 0.92 (s, 9H), 0.82 (s, 3H).

Step 4. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-[3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( C55 ).

O- ( 7 _ -azabenzotriazol-1-yl) -N, N, N', N' -tetramethyluronium (GATU; 154 mg, 0.405 mmol) and 4-methylmorpholine (0.144 ml, 1.31 mmol). Then the reaction mixture is heated from 0°C to 20°C for 1.5 hours, it is stirred at 20°C for 18 hours, then it is diluted with water and treated with solid sodium sulfate until saturation. The resulting mixture was extracted with a mixture of 2-propanol and chloroform (1:4, 3×20 ml) and the combined organic layers were dried over sodium sulfate, filtered and concentrated in vacuo . Chromatography on silica gel (gradient: 0% - 20% methanol in dichloromethane) gives C55 (240 mg) as a colorless glass. Part of this product is used in the next step. LCMS m/z 479.2 [M+H] ⁺ . According to ¹ H NMR analysis, this product is contaminated with a by-product obtained from the GATU reagent. ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks of the product only: δ 8.21 (d, J = 8.7 Hz, 1H), 7.53 (shs, 1H), 7.29 (brs, 1H), 7.03 (br, 1H), 6.02 (d, J = 9.6 Hz, 1H), 5.86 (q, J = 4.6 Hz, 1H), 4.31-4, 23 (m, 1H), 4.21 (s, 1H), 4.15 (d, J =9.6 Hz, 1H), 2.18-2.08 (m, 1H), 1.98-1 .88 (m, 1H), 1.68-1.55 (m, 1H), 1.54-1.42 (m, 2H), 1.34 (d, half AB quartet, J = 7.6 Hz , 1H), 1.01 (s, 3H), 0.90 (s, 9H), 0.84 (s, 3H).

Stage 5. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 78 ).

To a solution of C55 (from the previous step; 190 mg, ≤0.292 mmol) in acetonitrile (12 mL) was added methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 303 mg, 1.26 mmol). The reaction mixture is stirred at 20°C for 22 hours, then it is combined with a similar reaction carried out using C55 (from the previous stage; 50 mg, ≤77 μmol). The resulting solution was concentrated in vacuo , diluted with water (10 ml) and extracted with ethyl acetate (3×10 ml). The combined organic layers are dried over sodium sulfate, filtered, and concentrated in vacuo ; reverse phase HPLC purification (Column: Boston Prime C18, 30×150 mm, 5 µm; Mobile phase A: 0.225% formic acid in water; Mobile phase B: acetonitrile; Gradient: 23% - 46% B; Flow rate: 25 ml/minute) gives (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6.6 -dimethyl-3-[3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 78 ) as a white solid. Combined Yield: 25 mg, 54 µmol, 15% over 2 steps. LCMS m/z 461.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 8.96 (d, J = 8.6 Hz, 1H), 7.65 (s, 1H), 6.02 (d, J = 9.5Hz, 1H), 5.85(kv, J =4.5Hz, 1H), 4.95(ddd, J =10.8, 8.4, 5.1Hz, 1H), 4, 13 (d, J =9.6 Hz, 1H), 4.11 (s, 1H), 3.88-3.79 (m, 2H), 3.18-3.09 (m, 1H), 3 .07-2.98 (m, 1H), 2.48-2.37 (m, 1H), 2.20-2.02 (m, 2H), 1.77-1.62 (m, 2H) , 1.56-1.50 (m, 1H), 1.27 (d, half AB quartet, J = 7.6 Hz, 1H), 1.02 (s, 3H), 0.89 (s, 9H ), 0.85 (s, 3H). ¹ H NMR (400 MHz, chloroform- d ), characteristic peaks: δ 8.12 (d, J =7.6 Hz, 1H), 5.78 (brisk, 1H), 5.04 (brd, J =9 .4 Hz, 1H), 4.99-4.90 (m, 1H), 4.57-4.49 (m, 1H), 4.39 (d, J =9.7 Hz, 1H), 4 .25 (s, 1H), 4.01 (d, half AB quartet, J = 10.2 Hz, 1H), 3.93 (bdd, ABX system component, J = 10.6, 4.9 Hz, 1H ), 3.43-3.25 (m, 2H), 2.71 (d, J =4.8 Hz, 3H), 2.61-2.50 (m, 1H), 2.45-2, 30 (m, 2H), 2.03-1.93 (m, 1H), 1.91-1.78 (m, 1H), 1.05 (s, 3H), 0.98 (s, 9H) , 0.91 (s, 3H).

Example 79

Methyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3,3-dimethyl-1-oxobutan-2-yl}carbamate ( 79 )

Step 1. Synthesis of methyl (1 R ,2 S ,5 S )-3-[ N- (methoxycarbonyl)-3-methyl-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0] hexane-2-carboxylate ( C56 ).

To a 0° C. solution of C52 (370 mg, 1.16 mmol) in dichloromethane (6 ml) was slowly added triethylamine (0.647 ml, 4.64 mmol) and methyl chloroformate (335 mg, 3.55 mmol). then the reaction mixture was stirred at 20°C for 16 hours, it was diluted dropwise with saturated aqueous sodium carbonate solution (5 ml) and extracted with dichloromethane (2×20 ml). The combined organic layers are washed with saturated aqueous sodium chloride (2×20 ml), dried over sodium sulfate, filtered, and concentrated in vacuo ; chromatography on silica gel (gradient: 0% - 100% ethyl acetate in petroleum ether) gives C56 as a white gum. Yield: 115 mg, 0.338 mmol, 29%. LCMS m/z 341.1 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 5.29 (brd, J =9.6 Hz, 1H), 4.46 (s, 1H), 4.23 (d, J =9.9 Hz, 1H), 3.94-3.86(m, 2H), 3.74(s, 3H), 3.63(brs, 3H), 1.49-1.41(m, 2H), 1.04 (s, 3H), 1.03 (s, 9H), 0.91 (s, 3H).

Step 2. Synthesis of (1 R ,2 S ,5 S )-3-[ N- (methoxycarbonyl)-3-methyl-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane -2-carboxylic acid ( C57 ).

To a solution of C56 (115 mg, 0.338 mmol) in a mixture of methanol (2.0 ml), tetrahydrofuran (2.0 ml), and water (2 ml) was added lithium hydroxide monohydrate (28.4 mg, 0.677 mmol). The reaction mixture is stirred at room temperature (22°C - 25°C) for 16 hours, then concentrated in vacuo . The aqueous residue was partitioned between water (5 ml) and ethyl acetate (20 ml), then the organic layer was discarded and the aqueous layer was adjusted to pH 1-2 with concentrated hydrochloric acid. The resulting mixture is extracted three times with ethyl acetate; the combined organic layers are dried over sodium sulfate, filtered, and concentrated in vacuo to give C57 as a colorless gum. Yield: 100 mg, 0.306 mmol, 91%. LCMS m/z 327.2 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 5.42 (d, J =9.9 Hz, 1H), 4.46 (s, 1H), 4.26 (d, J =10.0 Hz, 1H), 3.96 (d, half of the AB quartet, J = 10.5 Hz, 1H), 3.87 (dd, component of the ABX system, J = 10.3, 5.4 Hz, 1H), 3.64 (s, 3H), 1.68 (d, half AB quartet, J = 7.7 Hz, 1H), 1.50 (dd, component of the ABX system, J = 7.6, 5.3 Hz, 1H), 1.06 (s, 3H), 1.01 (s, 9H), 0.91 (s, 3H).

Stage 3. Synthesis of methyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3-[(3 S )- 2-oxopyrrolidin-3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3,3-dimethyl-1-oxobutan- 2-yl}carbamate ( C58 ).

To a 0°C solution of C57 (100 mg, 0.306 mmol) and C16, HCl salts (75%, 84.8 mg, 0.306 mmol) in N, N -dimethylformamide (3 ml) was added hexafluorophosphate O- (7-azabenzotriazole-1 -yl)- N, N, N', N' -tetramethyluronium (GATU; 140 mg, 0.368 mmol), then a solution of 4-methylmorpholine (93 mg, 0.919 mmol) in N, N -dimethylformamide (1 ml) is added dropwise . The reaction mixture was then warmed to room temperature (25°C) and stirred for 16 hours, then water (10 ml) was added. After adding sodium sulfate until saturation, the resulting mixture was extracted with a mixture of chloroform and 2-propanol (4:1, 3×10 ml). The combined organic layers are dried over sodium sulfate, filtered, concentrated in vacuo , and purified using silica gel chromatography (Gradient: 0% - 30% methanol in dichloromethane) to give C58 as a white solid. Yield: 93 mg, 0.19 mmol, 62%. LCMS m/z 480.0 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 8.30 (brisk, 1H), 7.18 (brisk, 1H), 5.98 (brisk, 1H), 5.64 (brisk, 1H), 5, 58-5.42 (m, 1H), 4.49-4.37 (m, 1H), 4.29 (d, J =10.0 Hz, 1H), 4.23 (s, 1H), 4 .11 (dd, component of the ABX system, J = 10.3, 5.5 Hz, 1H), 3.93 (d, half of the AB quartet, J = 10.3 Hz, 1H), 3.64 (s, 3H ), 3.43-3.29 (m, 2H), 2.55-2.33 (m, 2H), 2.15-1.81 (m, 3H), 1.54-1.47 (m , 1H), 1.45 (d, half AB quartet, J = 7.7 Hz, 1H), 1.03 (s, 3H), 1.01 (s, 9H), 0.88 (s, 3H) .

Step 4. Synthesis of methyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine- 3-yl]ethyl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3,3-dimethyl-1-oxobutan-2-yl}carbamate ( 79 ) .

To a suspension of C58 (93 mg, 0.19 mmol) in dichloromethane (5 mL) was added methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 139 mg, 0.583 mmol) and the reaction mixture was stirred at 25°C for 2 hours. It is then diluted with water (10 ml) and extracted with dichloromethane (3×10 ml); the combined organic layers are washed with saturated aqueous sodium chloride (2×10 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (gradient: 0% - 100% ethyl acetate in petroleum ether, then gradient 0% - 20% methanol in dichloromethane) gives methyl {( 2S )-1-[( 1R , 2S , 5S )-2 -({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane -3-yl]-3,3-dimethyl-1-oxobutan-2-yl}carbamate ( 79 ) as a white solid. Yield: 7.0 mg, 15 µmol, 8%. LCMS m/z 462.2 [M+H] ⁺ . 1H NMR ⁽ 400 MHz, chloroform- d ) δ 8.13 (brd, J =7.0 Hz, 1H), 5.68 (brb, 1H), 5.34 (brd, J =9.9 Hz, 1H), 4.95-4.85 (m, 1H), 4.26 (s, 1H), 4.23 (d, J =10.0 Hz, 1H), 3.94 (dd, ABX system component , J = 10.1, 4.5 Hz, 1H), 3.88 (d, half AB quartet, J = 10.3 Hz, 1H), 3.63 (s, 3H), 3.45-3, 29 (m, 2H), 2.62-2.50 (m, 1H), 2.46-2.28 (m, 2H), 2.02-1.93 (m, 1H), 1.92- 1.79 (m, 1H), 1.6-1.49 (m, 2H, estimated; partially covered by water peak), 1.06 (s, 3H), 0.98 (s, 9H), 0.90 (s, 3H).

Example 80

N -(Trifluoroacetyl)-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-(trifluoromethyl )-L-Prolinamide ( 80 )

Step 1. Synthesis of 2-benzyl 1- tert -butyl (2S, 4R )-4-(trifluoromethyl)pyrrolidine-1,2- dicaroxylate ( C59 ).

A mixture of ( 4R )-1-( tert -butoxycarbonyl)-4-(trifluoromethyl)-L-proline (400 mg, 1.41 mmol), benzyl bromide (0.335 ml, 2.82 mmol) and sodium bicarbonate (593 mg, 7.06 mmol) in N,N -dimethylformamide (8 ml) was stirred for 15 hours at 25°C. The reaction mixture was then diluted with water (30 ml) and extracted with ethyl acetate (3 x 30 ml), the combined organic layers were washed with saturated aqueous sodium chloride and 5% aqueous lithium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 30% ethyl acetate in petroleum ether) gave C59 as a colorless oil. According to ¹ H NMR analysis, this product exists as a mixture of rotamers. Yield: 355 mg, 0.951 mmol, 67%. ¹ H NMR (400 MHz, chloroform- d ) δ 7.44-7.28 (m, 5H), 5.29-5.07 (m, 2H), [4.54 (brd, J =8.6 Hz) and 4.40 (bdd, J = 8.5, 2 Hz), total 1H], 3.87-3.70 (m, 1H), [3.58 (dd, J = 11.2, 7 .4 Hz) and 3.49 (dd, J = 11.0, 7.9 Hz), total 1H], 3.13-2.95 (m, 1H), 2.47-2.27 (m, 1H), 2.25-2.11 (m, 1H), [1.46 (s) and 1.33 (s), 9H total].

Stage 2. Synthesis of benzyl ( 4R )-4-(trifluoromethyl)-L-prolinate, hydrochloride ( C60 ).

To a 0°C solution of C59 (200 mg, 0.536 mmol) in ethyl acetate (3 ml) was added a solution of hydrochloride in ethyl acetate (4 M; 6 ml, 24 mmol). The reaction mixture is then stirred at room temperature (28°C) for 3 hours, it is concentrated in vacuo to give C60 as a white solid; this product is taken directly to the next stage. LCMS m/z 274.0 [M+H] ⁺ .

Step 3. Synthesis of benzyl N -( tert -butoxycarbonyl)-L- valyl- (4R)-4-(trifluoromethyl)-L-prolinate ( C61 ).

O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 277 mg, 0.728 mmol) and 4-methylmorpholine (184 mg, 1.82 mmol) are added to 0° C mixture of C60 (from previous step; ≤0.536 mmol) and N- ( tert -butoxycarbonyl)-L-valine (158 mg, 0.727 mmol) in N,N -dimethylformamide (3 mL). The reaction mixture was stirred at 0°C for 1 hour, then it was poured into ice water (15 ml) and extracted with ethyl acetate (2×15 ml). The combined organic layers are washed successively with 1 M hydrochloric acid, saturated aqueous sodium bicarbonate and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . Purification by chromatography on silica gel (Gradient: 0% - 40% ethyl acetate in petroleum ether) gave C61 as a colorless gum. Yield: 230 mg, 0.487 mmol, 91% over 2 steps. LCMS m/z 495.0 [M+Na ⁺ ]. ¹ H NMR (400 MHz, chloroform- d ), characteristic peaks: δ 7.40-7.30 (m, 5H), 5.17 (AB quartet, J _AB= 12.3 Hz, δν _AB= 12.6 Hz, 2H), 4.21 (dd, J =9.3, 6.8 Hz, 1H), 4.00-3.86 (m, 2H), 3.18-3.04 (m, 1H) , 2.36 (ddd, ABXY component of the system, J = 13.5, 9.9 Hz, 1H), 2.20 (ddd, ABXY component of the system, J = 13.4, 7.4, 3.5 Hz, 1H), 2.05-1.94 (m, 1H), 1.42 (s, 9H), 0.98 (d, J =6.7 Hz, 3H), 0.91 (d, J =6 .8 Hz, 3H).

Step 4. Synthesis of benzyl L- valyl- (4R)-4-(trifluoromethyl)-L-prolinate, hydrochloride ( C62 ).

To a 0°C solution of C61 (230 mg, 0.487 mmol) in ethyl acetate (2 ml) was added a solution of hydrochloride in ethyl acetate (4 M; 4 ml, 16 mmol). The reaction mixture was stirred at room temperature (28° C.) for 1 hour, then LCMS analysis showed conversion to C62 : LCMS m/z 373.1 [M+H] ⁺ . Concentration of the reaction mixture in vacuo gives C62 as a white solid which is taken directly to the next step.

Step 5. Synthesis of benzyl N- (trifluoroacetyl)-L- valyl- (4R)-4-(trifluoromethyl)-L-prolinate ( C63 ).

A solution of trifluoroacetic anhydride (154 mg, 0.733 mmol) in dichloromethane (0.5 ml) was added to a 0°C suspension of C62 (from the previous step; ≤0.487 mmol) in dichloromethane (3 ml). After 3 minutes, a solution of triethylamine (148 mg, 1.46 mmol) in dichloromethane (0.5 ml) was added dropwise and stirring continued at 25° C. for 3 hours. After dilution with dichloromethane (5 ml), the reaction mixture was washed with saturated aqueous sodium carbonate (10 ml) and saturated aqueous sodium chloride (15 ml), dried, filtered, and concentrated in vacuo ; chromatography on silica gel (Gradient: 0% - 30% ethyl acetate in petroleum ether) gave C63 as a colorless oil. Yield: 129 mg, 0.275 mmol, 56% over 2 steps. LCMS m/z 491.2 [M+Na ⁺ ].

Step 6. Synthesis of N- (trifluoroacetyl)-L- valyl- (4R)-4-(trifluoromethyl)-L-proline ( C64 ).

To a 28° C. solution of C63 (129 mg, 0.275 mmol) in methanol (3 mL) was added palladium on carbon (10%, 29.3 mg, 27.5 µmol) then the mixture was hydrogenated at 15 psi. within 16 hours. Filtration gave a filter cake which was washed with methanol (10 ml); the combined filtrates are concentrated in vacuo to give C64 as a light yellow solid. Yield: 80 mg, 0.21 mmol, 76%. LCMS m/z 401.0 [M+Na ⁺ ].

Step 7. Synthesis of N- (trifluoroacetyl)-L-valyl-(4 R )-4-(trifluoromethyl)-L-prolyl-3-[(3 S )-2-oxopyrrolidin-3-yl]-L-alanineamide ( C65 ).

O- (7-Azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 88.5 mg, 0.233 mmol) and 4-methylmorpholine (64.2 mg, 0.635 mmol) are added to 0°C solution of C64 (80 mg, 0.21 mmol) and C16 (76.8 mg, 0.287 mmol) in N,N -dimethylformamide (3 ml). then the reaction mixture is stirred at 0°C for 2 hours, it is treated with water (10 ml) and an aqueous solution of citric acid (1 M; 10 ml, 10 mmol), then extracted with ethyl acetate (3×10 ml). The combined organic layers were washed successively with saturated aqueous sodium bicarbonate (15 ml) and saturated aqueous sodium chloride (15 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Purification by silica gel chromatography (Gradient: 0% - 10% methanol in dichloromethane) gave C65 as a white solid. Yield: 72 mg, 0.14 mmol, 67%. LCMS m/z 532.2 [M+H] ⁺ .

Stage 8. Synthesis of N -(trifluoroacetyl)-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- 4-(trifluoromethyl)-L-prolinamide ( 80 ).

Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 96.9 mg, 0.407 mmol) was added to a mixture of C65 (72 mg, 0.14 mmol) in dichloromethane (5 ml) and the reaction mixture was stirred at room temperature in during the night. After dilution with water (15 ml), the mixture was extracted with dichloromethane (3 x 15 ml) and the combined organic layers were washed with saturated aqueous sodium chloride (2 x 20 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane) gives N- (trifluoroacetyl)-L- valyl- (4R) -N -{( 1S )-1-cyano-2-[( 3S ) -2-oxopyrrolidin-3-yl]ethyl}-4-(trifluoromethyl)-L-prolinamide ( 80 ) as a white solid. Yield: 30.9 mg, 60.2 µmol, 43%. LCMS m/z 536.1 [M+Na ⁺ ]. ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 9.89 (d, J =7.8 Hz, 1H), 9.06 (d, J =8.4 Hz, 1H), 7 .69 (s, 1H), 4.96 (ddd, J =10.6, 8.4, 5.5 Hz, 1H), 4.38 (dd, J =7.9, 6.3 Hz, 1H ), 4.28 (dd, J =9.8, 7.8 Hz, 1H), 4.07-3.94 (m, 2H), 3.20-3.00 (m, 2H), 2, 5-2.41 (m, 1H, estimated; partially covered by solvent peak), 2.38-2.28 (m, 1H), 2.19-2.02 (m, 4H), 1.78-1, 61 (m, 2H), 0.92 (d, J =7 Hz, 3H), 0.90 (d, J =6.8 Hz, 3H).

Examples 81-84

Example 81: ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3R )-5-hydroxy-2-oxopyrrolidin-3-yl]ethyl}-6 ,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide

Example 82: ( 1R , 2S , 5S , 6R ) -N -{( 1S )-1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-6- (hydroxymethyl)-6-methyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide

Example 83: (1 R ,2 S ,5 S ,6 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6- (hydroxymethyl)-6-methyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide

Example 84: (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3- (hydroxymethyl)- N -(trifluoroacetyl)-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide

Compounds of examples 81-84 are prepared by biotransformation, both in vitro and in vivo, from (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2- oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (compound example 13) as follows. In vitro studies , ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-6.6 -dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide is incubated with mouse, rat, hamster, rabbit, monkey or human (see Table M1 below) or rat, monkey or human hepatocytes (see Table M2 below). Alternatively, in in vivo studies (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6, 6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide is administered to rats and monkeys. Get samples of plasma, urine and bile of rats and plasma of monkeys. The resulting metabolites are then analyzed using HPLC/MS and the resulting oxidative metabolites of the compounds of examples 81-84 are determined and obtained. In addition to the compounds of examples 81-84, an additional metabolite, (1 R ,2 S ,5 S )-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3 ,1,0]hexane-2-carboxylic acid produced by hydrolytic cleavage has been observed in in vivo studies .

Table M1: Compounds derived from liver microsomes Example Mouse Rat Hamster Rabbit Monkey Man 81 +++ +++ +++ +++ +++ +++ 82 + + + ++ t + 83 + + + + t + 84 + t - t t + Table M2: Compounds of Examples 81-84 Derived from Hepatocytes Example Mouse Rat Hamster 81 +++ +++ +++ 82 + t t 83 t t t 84 + t - Table M3: Compounds of examples 81-84 obtained in vivo in mice or monkeys Example Rat plasma Rat urine Rat bile monkey plasma 81 + t t ++ 82 t t t t 83 t - - t 84 + - - +

In tables M1, M2 and M3, the following designations apply: -=not defined; +=determined by mass spectrometry and slight UV peak; ++=determined by mass spectrometry and average UV peak; +++=determined by mass spectrometry and main UV peak; t=trace determined by mass spectrometry only

Examples 82, 83, 84, and 81

(1 R ,2 S ,5 S ,6 R )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6-(hydroxymethyl) -6-methyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 82 ), (1 R ,2 S ,5 S ,6 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6-(hydroxymethyl)-6-methyl-3-[ 3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 83 ), (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-(hydroxymethyl)- N- (trifluoroacetyl)-L-valyl]-6,6- dimethyl-3-azabicyclo[3,1,0]hexane-2-carboxamide ( 84 ) and (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 R )-5-hydroxy-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0 ]hexane-2-carboxamide ( 81 )

Example 13 (25 µM) was combined with human liver microsomes (2 mg/ml) in potassium phosphate buffer (100 mM, pH 7.4; 40 ml) containing magnesium chloride (3.3 mM) and NADPH (1. 3 mM). Incubation is carried out for 55 minutes in a shaking water bath maintained at 37°C. The reaction is terminated by adding an equal volume of acetonitrile, then the mixture is centrifuged at 1800 x g for 5 minutes and the supernatant is subjected to vacuum centrifugation for approximately 1.5 hours. Formic acid (0.5 ml), acetonitrile (0.5 ml) and water were added to the residue to a final volume of 50 ml, and the resulting mixture was centrifuged at 40,000 x g for 30 minutes. The supernatant is applied to an HPLC column (Polaris C18, 4.6×250 mm; 5 µm) at 1 ml/min using a Jasco HPLC pump. After loading, the column is transferred to a Waters Acquity HPLC-UV system connected to a Thermo LTQ mass spectrometer and CTC Analytics fraction collector and subjected to reverse phase HPLC separation (Mobile phase A: water containing 0.1% formic acid (v/v). vol.); Mobile phase B: acetonitrile; Gradient: 2% over 5 minutes, then increased to 15% B, then 15% - 60% B over 80 minutes, then 60% - 95% B over 5 minutes; Flow rate: 0.8 ml/min). Fractions are collected every 20 seconds to obtain (1R.2S,fiveS.6R)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6-(hydroxymethyl)-6-methyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (82), (oneR.2S,fiveS.6S)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-6-(hydroxymethyl)-6-methyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (83), (oneR.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}-3-[3-(hydroxymethyl)-N-(trifluoroacetyl)-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide (84) and (1R.2S,fiveS)-N-{(oneS)-1-cyano-2-[(3R)-5-hydroxy-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl-N-(trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (81).

Example number Retention time, HPLC cleanup (minutes) 82 37.2 83 39.3 84 46.6 81 50.5

82 - Yield: 60 µg, 0.12 µmol, 12%. High resolution MS m/z 516.2424 [M+H] ⁺ ; calculated for C ₂₃ H ₃₃ F ₃ N ₅ O ₅ , 516.2434. ¹ H NMR (600 MHz, DMSO- d ₆ ), characteristic peaks: δ 9.42 (bd, J =7.9 Hz, 1H), 9.03 (d, J =8.5 Hz, 1H), 7 .66 (s, 1H), 5.01-4.93 (m, 1H), 4.69-4.63 (m, 1H), 4.43 (d, J = 8.3 Hz, 1H), 4.15 (s, 1H), 3.94 (dd, ABX system component, J = 10.2, 5.4 Hz, 1H), 3.68 (d, half AB quartet, J = 10.4 Hz, 1H), 3.21-3.17(m, 2H), 3.17-3.11(m, 1H), 3.07-3.00(m, 1H), 1.69-1.65( m, 1H), 1.44 (d, J =7.8 Hz, 1H), 0.98 (s, 9H), 0.84 (s, 3H).

83 - Yield: 30 µg, 0.058 µmol, 6%. High resolution MS m/z 516.2425 [M+H] ⁺ ; calculated for C ₂₃ H ₃₃ F ₃ N ₅ O ₅ , 516.2434. ¹ H NMR (600 MHz, DMSO- d ₆ ), characteristic peaks: δ 9.37 (d, J =7.0 Hz, 1H), 9.04 (d, J =8.5 Hz, 1H), 7 .65 (s, 1H), 5.00-4.94 (m, 1H), 4.54-4.49 (m, 1H), 4.40 (d, J = 8.1 Hz, 1H), 4.28 (s, 1H), 3.93 (dd, ABX system component, J = 10.2, 5.8 Hz, 1H), 3.74 (d, half AB quartet, J = 10.6 Hz, 1H), 3.3-3.20 (m, 1H, estimated; partially covered by water peak), 3.17-3.11 (m, 1H), 3.07-3.00 (m, 1H), 1 .75-1.63 (m, 2H), 1.38 (d, half AB quartet, J = 7.3 Hz, 1H), 1.06 (s, 3H), 0.98 (s, 9H).

84 - Yield: 40 µg, 0.078 µmol, 8%. High resolution MS m/z 516.2423 [M+H] ⁺ ; calculated for C ₂₃ H ₃₃ F ₃ N ₅ O ₅ , 516.2434. ¹ H NMR (600 MHz, DMSO- d ₆ ), characteristic peaks: δ 9.61-9.51 (m, 1H), 9.00 (d, J =8.3 Hz, 1H), 7.68 ( s, 1H), 5.02-4.92 (m, 1H), 4.51-4.43 (m, 1H), 4.16 (s, 1H), 3.92 (bdd, J =10, 0, 5.6 Hz, 1H), 3.78 (d, J = 10.6 Hz, 1H), 3.51 (d, J = 10.1 Hz, 1H), 3.18-3.10 ( m, 1H), 3.10-3.03 (m, 1H), 1.73-1.67 (m, 2H), 1.60-1.54 (m, 1H), 1.31 (d, J =7.6Hz, 1H), 1.03(s, 3H), 1.01(s, 3H), 0.88(brs, 6H).

81 - Yield: 130 µg, 0.252 µmol, 25%. This product was determined to exist as an interconvertible mixture of stereoisomers around the carbinolaine group of the pyrrolidone (see Examples 81 and 82). High resolution MS m/z 516.2428 [M+H] ⁺ ; calculated for C ₂₃ H ₃₃ F ₃ N ₅ O ₅ , 516.2434. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ [9.40 (d, J =8.4 Hz) and 9.38 (d, J =8.2 Hz), total 1H], [8.99 (d, J = 8.5 Hz) and 8.92 (d, J = 7.6 Hz), total 1H], [8.37 (s) and 8.25 (s), total 1H], [5 .83 (brs) and 5.70 (brs), total 1H], 5.04-4.92 (m, 2H), 4.44-4.38 (m, 1H), [4.19 (s) and 4.15 (s), total 1H], 3.91 (dd, J = 10.2, 5.5 Hz, 1H), [3.69 (d, J = 10 Hz) and 3.68 (d , J= 10.2 Hz), 1H total], [2.65-2.57 (m), 2.43-2.30 (m), and 2.21-2.13 (m), 2H total ], [2.08 (ddd, J =13.7, 8.4, 6.2 Hz), 2.00-1.90 (m), and 1.87-1.79 (m), total 2H ], [1.78-1.70 (m) and 1.51-1.44 (m), total 1H], 1.60-1.53 (m, 1H), [1.32 (d, J =7.6 Hz) and 1.29 (d, J =7.6 Hz), total 1H], 1.03 (s, 3H), [0.99 (s) and 0.98 (s), total 9H], [0.85 (s) and 0.84 (s), total 3H].

Examples 81 and 85

(1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 R )-5-hydroxy-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 81 ) and (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 R )-2,5-dioxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl )-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 85 )

A mixture of Example 13 (1.0 mg, 2.0 µmol) and tetra -n- butylammonium decatungstate (TBADT; 0.33 mg, 0.10 µmol) was treated with acetonitrile (0.15 ml) and hydrochloric acid (1.0 M ; 0.05 ml, 50 µmol). The syringe needle (18 gauge) is inserted through the Teflon cap of the tube and the air-available mixture is placed in an EvoluChem™ PhotoRedOx Box equipped with a fan and irradiated with black light (PAR20-18W LG 365 nm, 100-240 VAC) at 25°C for 16 hours . An aqueous solution of potassium phosphate (1 M, pH 7.5; 0.5 mL) was added to the reaction mixture, followed by water (to a volume of approximately 6 mL), then aqueous formic acid (1%, 2 mL) and sufficient acetonitrile were added to save the solution. The resulting solution is divided by half and applied to two 5 g Biotage Isolute C18 cartridges for solid phase extraction. The cartridges were washed with aqueous ammonium acetate (10 mM; 3 ml) and 20% acetonitrile in 10 mM aqueous ammonium acetate (3 ml), then eluted with acetonitrile (3 ml). The solvents were removed using a Genevac evaporator and the two residues were reconstituted in a mixture of acetonitrile and 1% aqueous formic acid and combined to a total of 2 ml solution. This product was subjected to reverse phase HPLC (Column: Phenomenex Luna C18.10×250 mm, 10 µm; Mobile phase A: water containing 0.1% formic acid (v/v); Mobile phase B: acetonitrile; Gradient : 2% - 15% B over 5 minutes, then 15% - 60% B over 80 minutes, then 60% - 95% B over 5 minutes; Flow rate: 2 ml/min). Fractions are collected every 20 seconds; the first compound eluted is ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3R )-5-hydroxy-2-oxopyrrolidin-3-yl]ethyl}- 6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 81 ), and the second compound eluted is (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 R )-2,5-dioxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 85 ).

Yield 81 : 0.122 mg, 0.237 µmol, 12%. This product is determined to exist as an interconvertible mixture of stereoisomers around the carbinolamine group of the pyrrolidone and elutes as a double peak on HPLC. High resolution MS m/z 516.2413 [M+H] ⁺ ; calculated for C ₂₃ H ₃₃ F ₃ N ₅ O ₅ , 516.2434. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 9.44-9.34 (m, 1H), [8.99 (d, J =8.5 Hz) and 8.92 (d, J =7 .6 Hz), 1H total], [8.37 (s) and 8.25 (s), 1H total], [5.83 (sh) and 5.70 (sh), 1H total], 5.05 -4.91 (m, 2H), 4.44-4.37 (m, 1H), [4.19 (s) and 4.15 (s), total 1H], 3.91 (dd, J = 10.3, 5.5 Hz, 1H), [3.69 (d, J =10.1 Hz) and 3.68 (d, J =10.3 Hz), total 1H], [2.65- 2.57 (m), 2.43-2.30 (m), and 2.17 (ddd, J = 14.9, 10.7, 4.7 Hz), total 2H], [2.08 ( ddd, J =14.1, 8.5, 6.2 Hz), 2.01-1.90 (m), and 1.83 (ddd, J =13.7, 10.1, 5.7 Hz ), total 2H], [1.78-1.70 (m) and 1.51-1.44 (m), total 1H], 1.60-1.53 (m, 1H), [1.32 (d, J = 7.6 Hz) and 1.29 (d, J = 7.6 Hz), total 1H], 1.03 (s, 3H), [0.99 (s) and 0.98 ( s), 9H total], [0.85 (s) and 0.84 (s), 3H total]. Retention time: 7.7 minutes (Analytical conditions. Column: Phenomenex Kinetex XB-C18, 2.1×100 mm, 2.6 µm; Mobile phase A: water containing 0.1% formic acid (v/v ); Mobile phase B: acetonitrile; Gradient: 5% B over 0.5 minutes, then 5% - 70% B over 10.5 minutes, then 70% - 95% B over 2 minutes; Flow rate: 0 .4 ml/minute).

Yield 85 : 0.104 mg, 0.203 µmol, 10%. High resolution MS m/z 514.2259 [M+H] ⁺ ; calculated for C ₂₃ H ₃₁ F ₃ N ₅ O ₅ , 514.2277. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 11.17 (brs, 1H), 9.40 (brs, 1H), 9.08 (d, J =8.9 Hz, 1H), 5.06 -4.98(m, 1H), 4.42-4.36(m, 1H), 4.13(s, 1H), 3.91(dd, J =10.3, 5.6Hz, 1H ), 3.70 (d, half of the AB quartet, J = 10.4 Hz, 1H), 3.00-2.93 (m, 1H), 2.60 (dd, component of the ABX system, J = 18.0 , 5.9 Hz, 1H), 2.46 (dd, component of the ABX system, J = 18.1, 9.1 Hz, 1H), 2.25-2.18 (m, 1H), 2.04- 1.97 (m, 1H), 1.60-1.55 (m, 1H), 1.35 (d, half AB quartet, J = 7.6 Hz, 1H), 1.03 (s, 3H) , 0.95 (s, 9H), 0.86 (s, 3H). Retention time: 8.3 minutes (Analytical conditions identical to those used for 81 ).

Example 86

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [5,5,5-trifluoro-2-(2,2,2-trifluoroacetamido)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 86 )

Stage 1. Synthesis of tert -butyl (1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl ]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-3-caroxylate ( C66 ).

To 0°C suspension of (1 R ,2 S ,5 S )-3-( tert -butoxycarbonyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxylic acid (5.25 g, 20.6 mmol), C16, HCl salts (4.70 g, 22.6 mmol) and 2-hydroxypyridine 1-oxide (571 mg, 5.14 mmol) in butan-2-one (108 ml) are added N,N -diisopropylethylamine (7.97 g, 61.7 mmol) followed by 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (4.73 g, 24.7 mmol). Then the reaction mixture is stirred at 0°C for 20 minutes, it is gradually warmed to room temperature, and then stirred at room temperature overnight, then LCMS analysis shows the presence of C66 : LCMS m/z 407.1 [M−H] ⁻ . The reaction mixture was diluted with ethyl acetate (100 ml), and washed successively with the following: a mixture of water (50 ml) and a saturated aqueous sodium chloride solution (20 ml), a saturated aqueous sodium chloride solution (70 ml), twice with a mixture of hydrochloric acid (1 M; 50 ml) and a saturated aqueous sodium chloride solution (20 ml) and finally a saturated aqueous sodium chloride solution (70 ml). Each aqueous layer was extracted with ethyl acetate (100 ml) and the combined organic layers were dried over sodium sulfate and filtered. The collected sodium sulfate was washed with ethyl acetate (2×50 ml) and the combined filtrates were concentrated in vacuo , diluted with heptane (50 ml) and concentrated under reduced pressure to give C66 as a colorless glass (6.69 g). According to ¹ H NMR analysis, some solvent is present; the purity is estimated as approximately 85% by weight. ¹ H NMR analysis also showed that this product exists as a mixture of rotamers. Yield corrected for the presence of solvents: 5.7 g, 14 mmol, 68%. ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 8.23-8.13 (m, 1H), [7.63 (br) and 7.59 (br) 1H total], [ 7.36 (brs) and 7.23 (brs), total 1H], 7.08-7.00 (m, 1H), 4.31-4.19 (m, 1H), [4.03 (s ) and 3.99 (s), total 1H], [3.58 (dd, J =10.8, 4.6 Hz) and 3.49 (dd, J =10.8, 3.9 Hz), 1H total], [3.27 (d, J =10.9 Hz) and 3.26 (d, J =10.7 Hz), 1H total], 3.22-3.00 (m, 2H), 2.38-2.09 (m, 2H), 1.79-1.43 (m, 2H), [1.36 (s) and 1.29 (s), total 9H], 1.00 (s , 3H), 0.89 (s, 3H).

Stage 2. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide, methanesulfonate ( C67 ).

Methanesulfonic acid (0.920 ml, 14.2 mmol) is added to a solution of C66 (ca. ole (30 ml). The reaction mixture is stirred at room temperature for 3 hours, then it is concentrated in vacuo . The residue was sequentially taken up in the following solvent systems, then re-concentrated: acetonitrile/ethyl acetate (1:1, 2×10 ml) and ethyl acetate/heptane (1:1, 2×10 ml) to give C67 as a glass (7.18 G). Some of this product is taken to the next stage. LCMS m/z 309.3 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ) δ 4.51 (dd, J = 10.8, 4.7 Hz, 1H), 4.21 (brisk, 1H), 3.73 (dd, J = 12.4, 6.3 Hz, 1H), 3.41-3.22 (m, 3H), 2.70 (s, 3H), 2.58-2.47 (m, 1H), 2.42 -2.32(m, 1H), 2.16(ddd, J =14.0, 10.8, 4.8Hz, 1H), 1.97(brd, J =7.9Hz, 1H), 1.95-1.84 (m, 1H), 1.84-1.75 (m, 2H), 1.15 (s, 6H).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl 3-[5,5,5-trifluoro-2-(2,2,2-trifluoroacetamido)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 86 ).

To a solution of 2-[( tert -butoxycarbonyl)amino]-5,5,5-trifluoropentanoic acid (59.0 mg, 0.218 mmol) in a mixture of acetonitrile (0.60 ml) and N,N -dimethylformamide (0.40 ml ) add O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 82.7 mg, 0.218 mmol), then 4-methylmorpholine (54.4 µl, 0.495 mmol ). The reaction mixture is then stirred for 20 minutes, C67 (from the previous step; 100 mg, ≤0.19 mmol) is added as a solid. The reaction mixture is stirred for 1.5 hours, then it is partitioned between ethyl acetate and saturated aqueous sodium bicarbonate. The aqueous layer is extracted with ethyl acetate and the combined organic layers are dried over magnesium sulfate, filtered, and concentrated using a stream of nitrogen. The residue was dissolved in dichloromethane (0.70 ml), treated with trifluoroacetic acid (0.175 ml) and stirred at room temperature. After 2 hours, trifluoroacetic acid (0.10 ml) is added again; stirring is continued for another 3 hours, then the reaction mixture is concentrated under a stream of nitrogen, and then in vacuo . This product is dissolved in dichloromethane (0.75 ml), cooled in an ice bath and treated with triethylamine (54.8 μl, 0.393 mmol); trifluoroacetic anhydride (41.2 μl, 0.292 mmol) was added dropwise and the reaction mixture was stirred at 0° C. for 3 hours. The volatiles were removed using a Genevac evaporator and the residue was dissolved in dichloromethane (0.90 ml), treated with methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 132 mg, 0.554 mmol), and stirred at room temperature for 2.5 hours. The reaction mixture is then concentrated under a stream of nitrogen, diluted with ethyl acetate and washed with saturated aqueous sodium bicarbonate. The aqueous layer is extracted with ethyl acetate and the combined organic layers are dried over magnesium sulfate, filtered, and concentrated under a stream of nitrogen. Reverse phase HPLC purification (Column: Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: 0.05% trifluoroacetic acid in water; Mobile phase B: 0.05% trifluoroacetic acid in acetonitrile; Gradient: 5% - 95% B for 8.54 minutes, then 95% B for 1.46 minutes; Flow rate: 25 ml/minute) gives (1 R ,2 S ,5 S ) -N -{(1 S )- 1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[5,5,5-trifluoro-2-(2,2,2-trifluoroacetamido )pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 86 ). Yield: 14 mg, 26 µmol, 14% over 2 steps. LCMS m/z 540.6 [M+H] ⁺ . Retention time: 2.60 minutes (Analytical conditions. Column: Waters Atlantis dC18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5.0% - 95% B, linear over 4.0 minutes, then 95% B over 1.0 minutes; Speed flow: 2 ml/minute).

Example 87

(1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[(2 S )- 2-cyclohexyl-2-{[(trifluoromethyl)sulfonyl]amino}acetyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 87 )

Stage 1. Synthesis of methyl (1 R ,2 S ,5 S )-3-{(2 S )-2-[( tert -butoxycarbonyl)amino]-2-cyclohexylacetyl}-6,6-dimethyl-3-azabicyclo[ 3,1,0]hexane-2-carboxylate ( C68 ).

To a 0°C solution of methyl (1 R ,2 S ,5 S )-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate, hydrochloride (300 mg, 1.46 mmol) and ( 2S )-[( tert -butoxycarbonyl)amino](cyclohexyl)acetic acid (394 mg, 1.53 mmol) in N,N -dimethylformamide (5 mL) O- (7-azabenzotriazol-1-yl) hexafluorophosphate is added - N, N, N', N' -tetramethyluronium (GATU; 610 mg, 1.60 mmol), then 4-methylmorpholine (443 mg, 4.38 mmol) is added dropwise. The reaction mixture was stirred at 0°C for 10 minutes, then at room temperature (20°C) for 2 hours, then it was poured into ice water (30 ml) and extracted with ethyl acetate (3×40 ml). The combined organic layers were washed successively with water (40 ml), hydrochloric acid (1 M; 40 ml), saturated aqueous sodium bicarbonate (40 ml), and saturated aqueous sodium chloride, then dried over sodium sulfate, filtered, and concentrated in vacuo to give C68 as a white foam. Yield: 580 mg, 1.42 mmol, 97%. LCMS m/z 409.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 7.03 (d, J =8.5 Hz, 1H), 4.17 (s, 1H), 4.00 (d, J =10.4 Hz , 1H), 3.88 (dd, J =9.9 Hz, 1H), 3.75 (dd, J =10.3, 5.3 Hz, 1H), 3.64 (s, 3H), 1 .83-1.47 (m, 8H), 1.46-1.28 (m, 2H), 1.33 (s, 9H), 1.18-1.06 (m, 3H), 1.00 (s, 3H), 0.86 (s, 3H).

Stage 2. Synthesis of methyl (1 R ,2 S ,5 S )-3-[(2 S )-2-amino-2-cyclohexylacetyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane -2-carboxylate, hydrochloride ( C69 ).

A solution of the hydrochloride in 1,4-dioxane (4 M; 15 ml) was added dropwise to a 5°C solution of C68 (580 mg, 1.42 mmol) in 1,4-dioxane (3 ml). The reaction mixture is then stirred at room temperature (20°C) for 1.5 hours, it is concentrated in vacuo . The residue was co-evaporated with dichloromethane to give C69 as a light yellow foam (490 mg), all of which was used in the next experiment. ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.22 (br s, 3H), 4.26 (s, 1H), 3.99-3.90 (m, 1H), 3.76 (dd, component of the ABX system, J = 10.7, 5.1 Hz, 1H), 3.71 (d, half of the AB quartet, J = 10.6 Hz, 1H), 3.66 (s, 3H), 1.83 -1.60 (m, 6H), 1.59 (dd, J =7.6, 5.1 Hz, 1H), 1.49 (d, half AB quartet, J =7.6 Hz, 1H), 1.27-1.02 (m, 5H), 1.03 (s, 3H), 0.94 (s, 3H).

Stage 3. Synthesis of methyl (1 R ,2 S ,5 S )-3-[(2 S )-2-cyclohexyl-2-{[(trifluoromethyl)sulfonyl]amino}acetyl]-6,6-dimethyl-3- azabicyclo[3,1,0]hexane-2-carboxylate ( C70 ).

To a -10°C solution of C69 (from the previous step; 480 mg, ≤1.39 mmol) in dichloromethane (10 ml) is added N,N -diisopropylethylamine (630 mg, 4.87 mmol), then trifluoromethanesulfonic anhydride ( 0.328 ml, 1.95 mmol). The reaction mixture was stirred at -10°C for 1 hour, then at room temperature (20°C) for 1 hour, then it was diluted with saturated aqueous sodium chloride solution (50 ml) and extracted with dichloromethane (3×30 ml). The combined organic layers are dried over sodium sulfate, filtered, concentrated in vacuo , and purified by silica gel chromatography (eluent: 1:4 ethyl acetate/petroleum ether) to give C70 as a light yellow oil. Yield: 124 mg, 0.282 mmol, 20% over 2 steps. LCMS m/z 441.1 [M+H] ⁺ .

Stage 4. Synthesis of (1 R ,2 S ,5 S )-3-[(2 S )-2-cyclohexyl-2-{[(trifluoromethyl)sulfonyl]amino}acetyl]-6,6-dimethyl-3-azabicyclo [3,1,0]hexane-2-carboxylic acid ( C71 ).

To a solution of C70 (120 mg, 0.272 mmol) in a mixture of water (2 ml), methanol (2 ml) and tetrahydrofuran (2 ml) was added lithium hydroxide monohydrate (28.6 mg, 0.682 mmol). Then the reaction mixture is stirred at room temperature (20°C) for 18 hours at room temperature, LCMS shows that the reaction is complete: LCMS m/z 427.2 [M+H] ⁺ . The reaction mixture is concentrated in vacuo to remove organic solvents. The residue was diluted with water (5 ml) and then acidified to pH 2-3 with 1 M hydrochloric acid; the resulting mixture was extracted with ethyl acetate (2×30 ml). The combined organic layers are dried over sodium sulfate, filtered, and concentrated in vacuo to give C71 as a light yellow solid. Yield: 92.0 mg, 0.216 mmol, 79%.

Step 5. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-3-[(2 S )-2-cyclohexyl-2-{[(trifluoromethyl)sulfonyl]amino}acetyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2- carboxamide ( C72 ).

To a 0°C solution of C71 (92.0 mg, 0.216 mmol) and C16, HCl salts (72%, 68.8 mg, 0.238 mmol) in N, N -dimethylformamide (3 ml) was added hexafluorophosphate O- (7-azabenzotriazole -1-yl)- N, N, N', N' -tetramethyluronium (GATU; 98.4 mg, 0.259 mmol), then 4-methylmorpholine (76.4 mg, 0.755 mmol). The reaction mixture was then stirred at 20°C for 2 hours, then it was poured into ice water (10 ml) and extracted with a mixture of chloroform and 2-propanol (4:1, 4×20 ml). The combined organic layers are dried over sodium sulfate, filtered, concentrated in vacuo , and purified by silica gel chromatography (Gradient: 0% - 10% methanol in dichloromethane) to give C72 as a colorless glass. Yield: 100 mg, 0.173 mmol, 80%. LCMS m/z 580.2 [M+H] ⁺ .

Stage 6. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[( 2 S )-2-cyclohexyl-2-{[(trifluoromethyl)sulfonyl]amino}acetyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 87 ).

A solution of trifluoroacetic anhydride (47.1 mg, 0.224 mmol) in dichloromethane (1 ml) was added dropwise to a 0°C solution of C72 (100 mg, 0.173 mmol) and pyridine (41.7 μl, 0.516 mmol) in dichloromethane (3 ml ). The mixture is then stirred for 20 hours at room temperature (10°C - 20°C), it is concentrated in vacuo and redissolved in dichloromethane (3 ml). Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 103 mg, 0.432 mmol) was added and the reaction mixture was stirred at room temperature (20° C.) for 20 hours. It is then diluted with saturated aqueous sodium chloride (10 ml) and extracted with dichloromethane (3×20 ml); the combined organic layers are dried over sodium sulfate, filtered, concentrated in vacuo , and subjected to supercritical liquid chromatography [Column: Chiral Technologies Chiralcel OD, 30×250 mm, 10 µm; Mobile phase: 4:1 carbon dioxide/(ethanol containing 0.1% ammonium hydroxide); Flow rate: 60 ml/minute]. Fractions containing 87 are concentrated under vacuum below 40° C. to remove the alcohol co-solvent. The residue was diluted with ethyl acetate (50 ml) and dichloromethane (5 ml) and washed successively with hydrochloric acid (1 M; 20 ml), saturated aqueous sodium bicarbonate (20 ml), and saturated aqueous sodium chloride. The resulting organic layer was dried over sodium sulfate, filtered, and concentrated in vacuo, then mixed with water (20 ml) and acetonitrile (5 ml); this mixture is lyophilized to give ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-3-[ (2 S )-2-cyclohexyl-2-{[(trifluoromethyl)sulfonyl]amino}acetyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 87 ) as white solid. Yield: 27.6 mg, 49.1 µmol, 28%. LCMS m/z 562.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 9.93 (d, J =8.3 Hz, 1H), 9.10 (d, J =8.7 Hz, 1H), 7 .70 (s, 1H), 5.03-4.94 (m, 1H), 4.15 (s, 1H), 3.88-3.78 (m, 2H), 3.55 (d, J =10.2 Hz, 1H), 3.20-3.11 (m, 1H), 3.10-3.00 (m, 1H), 2.19-2.08 (m, 1H), 2, 06-1.95 (m, 1H), 1.85-1.53 (m, 9H), 1.33 (d, half AB quartet, J = 7.7 Hz, 1H), 1.03 (s, 3H), 0.89 (s, 3H).

Examples 88 and 89

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclobutyl- N -(trifluoroacetyl)-L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide and ( 1R , 2S , 5S ) -N -{( 1S ) -1-Cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclobutyl- N- (trifluoroacetyl)-D-alanyl]-6,6-dimethyl-3- azabicyclo[3,1,0]hexane-2-carboxamide ( 88 and 89 )

Stage 1. Synthesis of tert -butyl {1-[(1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidine -3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3-cyclobutyl-1-oxopropan-2-yl}carbamate ( C73 ).

A 0°C solution of C67 (150 mg, 0.371 mmol) and N- ( tert -butoxycarbonyl)-3-cyclobutylalanine (99.2 mg, 0.408 mmol) in N,N -dimethylformamide (3 ml) was treated with O- (7- azabenzotriazol-1-yl) -N, N, N', N'- tetramethyluronium (GATU; 169 mg, 0.444 mmol). Then add dropwise 4-methylmorpholine (131 mg, 1.30 mmol), then the reaction mixture is heated to 25°C and stirred overnight. Ice water (10 ml) was added and the resulting mixture was extracted with a mixture of chloroform and 2-propanol (4:1, 4×20 ml); the combined organic layers are dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane) is carried out twice to give C73 as a white solid which contains a mixture of two diastereomers. Yield: 106 mg, 0.199 mmol, 54%. LCMS m/z 534.2 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ [8.59 (d, J =5.5 Hz) and 7.85 (d, J =7.7 Hz), total 1H], 7.20-7 .06 (m, 1H), [5.78 (brs) and 5.67 (brs), 1H total], [5.51 (br) and 5.40 br (s), 1H total], 5.22 -5.12 (m, 1H), [4.49-4.39 (m) and 4.38-4.23 (m), total 3H], 4.17-4.06 (m, 1H), [3.83 (d, J =10.4 Hz) and 3.50 (d, J =10.5 Hz), total 1H], 3.42-3.28 (m, 2H), 2.50- 2.30 (m, 3H), 2.23-2.00 (m, 4H), 2.00-1.77 (m, 4H), 1.73-1.44 (m, 5H), [1 .40 (s) and 1.39 (s), 9H total], [1.07 (s) and 1.03 (s), 3H total], [0.98 (s) and 0.92 (s) , total 3H].

Stage 2. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3 -cyclobutyl- N- (trifluoroacetyl)-L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide and (1 R ,2 S ,5 S ) -N- { (1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclobutyl- N- (trifluoroacetyl)-D-alanyl]-6,6- dimethyl-3-azabicyclo[3,1,0]hexane-2-carboxamide [ 88 (DIAST-1) and 89 (DIAST-2) ].

A solution of hydrochloride in 1,4-dioxane (4 M; 5 ml, 20 mmol) was added dropwise to a 0° C. solution of C73 (106 mg, 0.199 mmol) in dichloromethane (5 ml). The reaction mixture was stirred at 0°C for 15 minutes and then at 25°C for 2 hours, then it was concentrated in vacuo to give the deprotected product as a white solid: LCMS m/z 434.2 [M+H ] ⁺ . This is dissolved in dichloromethane (3 ml), cooled in an ice bath, and treated with pyridine (79.9 mg, 1.01 mmol) and a solution of trifluoroacetic anhydride (170 mg, 0.809 mmol) in dichloromethane (1.5 ml). The reaction mixture is stirred at 20°C for 20 hours; then pyridine (40.0 mg, 0.506 mmol) is added and stirring is continued for another 12 hours at 25°C. After dilution with dichloromethane (15 ml), the reaction mixture was washed successively with hydrochloric acid (1 M; 10 ml), saturated aqueous sodium bicarbonate solution (3×10 ml), and saturated aqueous sodium chloride solution (10 ml), dried, filtered, and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 20% methanol in dichloromethane) then supercritical liquid chromatography [Column: Chiral Technologies Chiralpak IC, 30×250 mm, 10 μm; Mobile phase: 3:1 carbon dioxide/(methanol containing 0.1% ammonium hydroxide); Flow rate: 70 ml/minute] to give separated diastereomers (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-3-[3-cyclobutyl- N- (trifluoroacetyl)-L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide and (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclobutyl- N -(trifluoroacetyl)- D-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide. Which product contains the D-alanyl configuration and which contains the L-alanyl configuration was not determined; the first eluted diastereomer is designated 88 , and the second eluted diastereomer is designated 89 . Both are obtained as a white solid.

88 - Yield: 9.3 mg, 18.2 µmol, 9% over 2 steps. LCMS m/z 512.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.75 (bd, J =7.0 Hz, 1H), 8.96 (d, J =8.1 Hz, 1H), 7.71 (s , 1H), 5.00-4.90 (m, 1H), 4.32-4.23 (m, 1H), 4.13 (s, 1H), 3.82 (dd, ABX system component, J \u003d 10.2, 5.4 Hz, 1H), 3.69 (d, half AB quartet, J \u003d 10.2 Hz, 1H), 3.20-3.02 (m, 2H), 2.44- 2.28 (m, 2H), 2.16-2.04 (m, 2H), 2.04-1.91 (m, 2H), 1.87-1.54 (m, 9H), 1, 32 (d, half AB quartet, J = 7.6 Hz, 1H), 1.03 (s, 3H), 0.88 (s, 3H). Retention time: 2.78 minutes (Analytical conditions. Column: Chiral Technologies Chiralpak IC-3, 4.6×150 mm, 3 µm; Mobile phase A: carbon dioxide; Mobile phase B: methanol containing 0.05% diethylamine (vol ./v); Gradient: 5% - 40% B over 5 minutes, then 40% B over 2.5 minutes; Back pressure: 1500 psi; Flow rate: 2.5 ml/ minute).

89 - Yield: 23 mg, 45.0 mmol, 23% over 2 steps. ¹ H NMR analysis shows that this product exists as a mixture of rotamers. LCMS m/z 512.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ [9.92 br (s) and 9.65 (br s), total 1H], [9.22 (d, J = 7.7 Hz) and 8, 85 (d, J = 8.4 Hz), 1H total], [7.76 (s) and 7.67 (s), 1H total], [5.11-5.00 (m) and 4.98 -4.87 (m), 1H total], [4.51 (s) and 4.07 (s), 1H total], [4.47-4.36 (m) and 4.09-4.00 (m), total 1H], [3.90 (dd, J = 10.2, 5.3 Hz) and 3.60-3.45 (m), total 2H], 3.21-3.00 ( m, 2H), 2.44-2.33 (m, 1H), 2.28-1.98 (m, 3H), 1.98-1.52 (m, 10H), [1.49-1 .38 (m) and 1.32 (d, half an AB quartet, J = 7.6 Hz), 2H total], [1.04 (s) and 1.02 (s), 3H total], [0, 93 (s) and 0.82 (s), total 3H]. Retention time: 4.14 minutes (Analytical conditions identical to those used for 88 ).

Example 90

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-(pyridin-2-yl) -N- (trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 90 )

Stage 1. Synthesis of tert -butyl [(2 S )-1-[(1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-1-oxo-3-(pyridine -2-yl)propan-2-yl]carbamate ( C74 ).

To a 0°C solution of C67 (250 mg, 0.618 mmol) and N- ( tert -butoxycarbonyl)-3-pyridin-2-yl-L-alanine (198 mg, 0.744 mmol) in N,N -dimethylformamide (2 mL) O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 282 mg, 0.742 mmol) was added, then a solution of 4-methylmorpholine (188 mg, 1.86 mmol) in N,N -dimethylformamide (1 ml). The reaction mixture is then heated to 20°C and stirred for 2 hours, then it is diluted with water (10 ml) and extracted with ethyl acetate (10 ml). Solid sodium sulfate is added to the aqueous layer until saturation is reached, then the aqueous layer is extracted with a mixture of dichloromethane and methanol (10:1, 3×20 ml). The combined organic layers were concentrated in vacuo and purified by silica gel chromatography (Gradient: 0% - 10% methanol in dichloromethane) to give C74 as a yellow gum. Yield: 250 mg, 0.449 mmol, 73%. LCMS m/z 557.0 [M+H] ⁺ . ¹ H NMR (400 MHz, methanol- d ₄ ), characteristic peaks: δ 8.49 (d, J =5.2 Hz, 1H), 7.78-7.71 (m, 1H), 7.35 ( d, J = 7.8 Hz, 1H), 7.30-7.25 (m, 1H), 4.73 (dd, J = 8.5, 5.2 Hz, 1H), 4.40 (dd , J = 11.8, 4.2 Hz, 1H), 4.30 (s, 1H), 4.01-3.90 (m, 1H), 3.26 (dd, J = 14.2.5 .6 Hz, 1H), 2.94 (dd, J = 14.1, 8.9 Hz, 1H), 2.65-2.53 (m, 1H), 2.37-2.25 (m, 1H), 2.14-2.04 (m, 1H), 1.91-1.78 (m, 2H), 1.61-1.55 (m, 1H), 1.53 (d, half AB quartet, J = 7.7 Hz, 1H), 1.34 (s, 9H), 1.08 (s, 3H), 1.00 (s, 3H).

Stage 2. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-[3-(pyridin-2-yl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide hydrochloride ( C75 ).

A solution of hydrochloride in 1,4-dioxane (4 M; 6 ml) was added to a 0°C solution of C74 (250 mg, 0.449 mmol) in dichloromethane (2 ml) and the reaction mixture was stirred for 5 hours at 20°C. LCMS analysis shows conversion to C75 : LCMS m/z 457.1 [M+H] ⁺ . Removal of solvents in vacuo gave C75 as a yellow solid (250 mg); part of this product is used directly in the next step. ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 8.81 (bd, J =5.4 Hz, 1H), 8.43 (d, J =8.3 Hz, 1H), 8 .39-8.31 (m, 1H), 7.90 (d, J = 8.0 Hz, 1H), 7.87-7.81 (m, 1H), 7.65 (s, 1H), 7.43(brs, 1H), 7.06(brs, 1H), 4.77-4.67(m, 1H), 4.35(s, 1H), 4.32-4.23(m, 1H), 3.94-3.86 (m, 1H), 3.80 (d, half AB quartet, J = 10.6 Hz, 1H), 3.36-3.25 (m, 1H), 3 .20-3.08 (m, 2H), 2.35-2.23 (m, 1H), 2.17-2.06 (m, 1H), 2.01-1.90 (m, 1H) , 1.55-1.48 (m, 1H), 1.43 (d, half AB quartet, J = 7.6 Hz, 1H), 1.04 (s, 3H), 0.96 (s, 3H ).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl 3-[3-(pyridin-2-yl) -N- (trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 90 ).

To a 0°C solution of C75 (from the previous step; 175 mg, ≤0.314 mmol) in dichloromethane (6 mL) was added pyridine (197 mg, 2.49 mmol) followed by a solution of trifluoroacetic anhydride (186 mg, 0.886 mmol) in dichloromethane ( 2 ml). The reaction mixture is then stirred at 20°C for 2 hours, diluted with water and extracted with a mixture of dichloromethane and methanol (10:1, 3×15 ml). The combined organic layers are concentrated in vacuo and subjected to silica gel chromatography (Gradient: 0% - 10% methanol in dichloromethane), then supercritical liquid chromatography [Column: Chiral Technologies Chiralpak AS, 30×250 mm, 10 μm; Mobile phase: 3:1 carbon dioxide/(ethanol containing 0.1% ammonium hydroxide); Flow rate: 70 ml/minute] to give (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}-6,6-dimethyl-3-[3-(pyridin-2-yl) -N- (trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 90 ) as a white solid. Yield: 25 mg, 46.8 µmol, 15% over 2 steps. LCMS m/z 535.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.92 (d, J =7.5 Hz, 1H), 8.91 (d, J =8.0 Hz, 1H), 8.51 (bd , J = 5 Hz, 1H), 7.75-7.68 (m, 2H), 7.28 (d, J = 7.8 Hz, 1H), 7.25 (dd, J = 7.3, 5.1 Hz, 1H), 5.00-4.88 (m, 2H), 4.15 (s, 1H), 3.86 (dd, ABX system component, J =10.3, 5.4 Hz , 1H), 3.70 (d, half AB quartet, J = 10.3 Hz, 1H), 3.21-3.04 (m, 4H), 2.42-2.31 (m, 1H), 2.18-2.04 (m, 2H), 1.76 (ddd, J =13.5, 9.6, 6.6 Hz, 1H), 1.74-1.62 (m, 1H), 1.62-1.53 (m, 1H), 1.33 (d, half AB quartet, J = 7.6 Hz, 1H), 1.03 (s, 3H), 0.89 (s, 3H) .

Example 91

(1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-{ N -[(4 -fluorophenoxy)acetyl]-3-methyl-L-valyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 91 )

Stage 1. Synthesis of tert -butyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3,3-dimethyl-1- oxobutan-2-yl}carbamate ( C76 ).

A solution of C32 (15.4 g, 41.8 mmol) and C16, HCl salts (75%, 11.6 g, 41.9 mmol) in N,N -dimethylformamide (380 ml) is cooled to -5°C - 0 °C O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 18.3 g, 48.1 mmol) and 4-methylmorpholine (12.7 g, 126 mmol) at -5°C - 0°C. Then the reaction mixture was stirred at 0°C for 1.5 hours, it was poured into ice water (400 ml) and the resulting mixture was extracted with ethyl acetate (3×200 ml). The combined organic layers are washed successively with aqueous citric acid (1 M; 120 ml), saturated aqueous sodium bicarbonate (120 ml) and saturated aqueous sodium chloride (3×60 ml), dried over sodium sulfate, filtered, concentrated in vacuo . and combined with the crude product from a similar reaction using C32 (1.08 g, 2.93 mmol). Chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane) gave C76 as a white solid (9.80 g). The combined aqueous layers are extracted with a mixture of chloroform and 2-propanol (4:1, 3×100 ml); concentration of these combined extracts followed by silica gel chromatography (Gradient: 0% - 10% methanol in dichloromethane) gave additional C76 as a white solid (2.3 g). Combined yield: 12.1 gm, 23.2 mmol, 52%. LCMS m/z 522.5 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 8.28 (brb, J = 5.9 Hz, 1H), 7.20 (bbr, 1H), 5.71 (brb, 1H), 5.38 ( brs, 1H), 5.10 (brd, J =10.3 Hz, 1H), 4.47-4.38 (m, 1H), 4.28-4.20 (m, 2H), 4.12 (dd, component of the ABX system, J = 10.2, 4.0 Hz, 1H), 3.99 (d, half of the AB quartet, J = 10.2 Hz, 1H), 3.40-3.33 (m , 2H), 2.53-2.35 (m, 2H), 2.17-2.07 (m, 1H), 2.00-1.81 (m, 2H), 1.52-1.4 (m, 2H, estimated; significantly obscured by water and t- butyl peak signal), 1.39 (s, 9H), 1.02 (s, 3H), 1.01 (s, 9H), 0.88 (s , 3H).

Stage 2. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-(3-methyl-L-valyl)-3-azabicyclo[3.1.0]hexane-2-carboxamide hydrochloride ( C77 ).

To a 0°C solution of C76 (12.1 g, 23.2 mmol) in dichloromethane (50 ml) was added a solution of hydrochloride in 1,4-dioxane (4 M; 250 ml). Then the reaction mixture is stirred at 20°C for 2 hours, it is filtered. The filter cake was stirred with methyl t- butyl ether (250 mL) for 18 hours; filtration gave C77 as a light yellow/white solid (10.89 g). LCMS m/z 422.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.38 (d, J = 9.0 Hz, 1H), 8.19 (brine, 3H), 7.57 (brine, 1H), 7.41 (brs, 1H), 7.04 (brs, 1H), 4.35-4.27 (m, 1H), 4.34 (s, 1H), 3.85-3.72 (m, 2H), 3.65 (d, half AB quartet, J = 10.8 Hz, 1H), 3.16-3.09 (m, 1H), 3.05-2.95 (m, 1H), 2.43- 2.31 (m, 1H), 2.17-2.04 (m, 1H), 2.00-1.89 (m, 1H), 1.71-1.42 (m, 3H), 1, 38 (d, half AB quartet, J = 7.7 Hz, 1H), 1.02 (s, 3H), 1.02 (s, 9H), 0.97 (s, 3H).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-3-[ N- (chloroacetyl)-3-methyl-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( C78 ).

Triethylamine (2.21 g, 21.8 mmol) is added to a 0°C solution of C77 (from the previous step; 2.50 g, ≤5.33 mmol) in dichloromethane (100 ml). A solution of chloroacetyl chloride (1.23 g, 10.9 mmol) in dichloromethane (9 ml) was added dropwise to the reaction mixture, and stirring was continued at 0° C. for 1 hour. Water (50 ml) was then added and the resulting mixture was extracted into a mixture. chloroform and 2-propanol (4:1, 3×50 ml); the combined organic layers are washed with saturated aqueous sodium chloride (50 ml), dried over sodium sulfate, filtered, and concentrated in vacuo . Purification by chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane) gave C78 as a white solid. Yield: 1.21 g, 2.43 mmol, 46% over 2 steps. LCMS m/z 498.1 (chlorine isotope pattern observed) [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.30-8.22 (m, 2H), 7.54 (br br, 1H), 7.30 (br br, 1H), 7.03 (br br, 1H), 4.35 (d, J = 8.9 Hz, 1H), 4.29 (ddd, J = 12.1, 8.7, 3.4 Hz, 1H), 4.24 (s, 1H ), 4.11 (AB quartet, J _AB= 12.4 Hz, δν _AB= 14.3 Hz, 2H), 3.86 (dd, ABX system component, J = 10.2, 5.4 Hz, 1H ), 3.72 (d, half AB quartet, J = 10.3 Hz, 1H), 3.18-3.08 (m, 1H), 3.07-2.97 (m, 1H), 2, 48-2.36 (m, 1H), 2.19-2.08 (m, 1H), 1.99-1.88 (m, 1H), 1.69-1.43 (m, 3H), 1.37 (d, half AB quartet, J = 7.7 Hz, 1H), 1.01 (s, 3H), 0.94 (s, 9H), 0.84 (s, 3H).

Step 4. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-3-{ N -[(4-fluorophenoxy)acetyl]-3-methyl-L-valyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( C79 ).

4-Fluorophenol (49.5 mg, 0.442 mmol) was added to a mixture of cesium fluoride (67.1 mg, 0.442 mmol) in N,N -dimethylformamide (3 ml) and the mixture was stirred at 65°C for 1 hour, then added C78 (110.0 mg, 0.221 mmol) and the reaction mixture was stirred at 65°C for 8 hours. It is then combined with a similar reaction carried out using C78 (30 mg, 60 μmol), poured into water (10 ml) and extracted with ethyl acetate (3×10 ml). The combined organic layers were washed successively with water (20 ml), hydrochloric acid (1 M; 10 ml), saturated aqueous sodium carbonate (10 ml) and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane) gives C79 as a white glass. Combined yield: 100 mg, 0.174 mmol, 62%. LCMS m/z 574.2 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 8.23 (brd, J =6 Hz, 1H), 7.19 (brb, 1H), 7.04 (brd, J =10.2 Hz, 1H) , 6.99 (dd, J =9.2, 8.0 Hz, 2H), 6.85 (dd, ABX system component, J =9.1, 4.2 Hz, 2H), 5.78 (ws , 1H), 5.46 (brs, 1H), 4.65 (d, J = 9.8 Hz, 1H), 4.52-4.38 (m, 3H), 4.22 (s, 1H) , 4.14 (dd, component of the ABX system, J = 10.3, 5.3 Hz, 1H), 3.93 (d, half of the AB quartet, J = 10.3 Hz, 1H), 3.41-3 .32 (m, 2H), 2.55-2.34 (m, 2H), 2.14-2.05 (m, 1H), 2.03-1.80 (m, 3H), 1.03 (s, 3H), 1.00 (s, 9H), 0.86 (s, 3H).

Stage 5. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-{ N -[(4-fluorophenoxy)acetyl]-3-methyl-L-valyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 91 ).

A solution of methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 125 mg, 0.524 mmol) in dichloromethane (2 mL) was added dropwise to a 10°C (room temperature) solution of C79 (100 mg, 0.174 mmol) in dichloromethane ( 4 ml). Then the reaction mixture was stirred at 10°C for 16 hours, it was diluted with water (10 ml) and extracted with dichloromethane (3×10 ml). The combined organic layers are washed with saturated aqueous sodium chloride (2×10 ml) and concentrated in vacuo ; chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane), then supercritical liquid chromatography [Column: Chiral Technologies Chiralcel OD, 30×250 mm, 10 μm; Mobile phase: 4:1 carbon dioxide/(ethanol containing 0.1% ammonium hydroxide); Flow rate: 60 ml/minute] gives (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl }-3-{ N -[(4-fluorophenoxy)acetyl]-3-methyl-L-valyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 91 ) in form of a white solid. Yield: 55 mg, 99.0 µmol, 57%. LCMS m/z 556.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.01 (d, J =8.5 Hz, 1H), 7.96 (d, J =9.1 Hz, 1H), 7.68 (s , 1H), 7.13-7.04 (m, 2H), 6.90-6.84 (m, 2H), 4.97 (ddd, J =10.9, 8.5, 5.1 Hz , 1H), 4.61-4.50 (m, 2H), 4.39 (d, J = 8.9 Hz, 1H), 4.12 (s, 1H), 3.87 (dd, system component ABX, J = 10.4, 5.5 Hz, 1H), 3.73 (d, half AB quartet, J = 10.4 Hz, 1H), 3.19-3.09 (m, 1H), 3 .09-2.99 (m, 1H), 2.47-2.36 (m, 1H, estimated; partially covered by solvent peak), 2.21-2.02 (m, 2H), 1.79-1 .65 (m, 2H), 1.53 (dd, J = 7.6, 5.4 Hz, 1H), 1.29 (d, half AB quartet, J = 7.6 Hz, 1H), 1, 00 (s, 3H), 0.93 (s, 9H), 0.75 (s, 3H).

Example 92

3-Methyl- N -[(4-methylphenyl)acetyl]-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-4-(trifluoromethyl)-L-prolinamide ( 92 )

A mixture of C51 (68.0 mg, 0.12 mmol) and a solution of hydrochloride in 1,4-dioxane (4 M; 1 ml, 4 mmol) was stirred at room temperature for 5 minutes, then the reaction mixture was concentrated in vacuo to remove the solvent , then further aspirated using high vacuum to remove residual hydrochloride. (4-Methylphenyl)acetic acid (18.6 mg, 0.124 mmol) was added to the residue; the resulting mixture was dissolved in N,N -dimethylformamide (1.0 ml) and cooled to -30°C. After adding N, N -diisopropylethylamine (64.7 μl, 0.371 mmol), then O - (7-azabenzotriazol-1-yl) - N, N, N', N' -tetramethyluronium hexafluorophosphate (GATU; 61.2 mg, 0.161 mmol), the reaction mixture is warmed to room temperature for 1 hour and then treated with aqueous sodium bicarbonate. The mixture is extracted 5 times with ethyl acetate and the combined organic layers are concentrated under reduced pressure. The residue was then dissolved in dichloromethane (1 ml), treated with methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 88.5 mg, 0.371 mmol), and stirred at room temperature for 1 hour, then the reaction mixture was treated with dilute aqueous sodium carbonate solution and extracted twice with ethyl acetate. The combined organic layers are dried over magnesium sulfate, filtered, concentrated in vacuo , and purified by reverse phase HPLC (Column: Waters XBridge C18, 19×100 mm, 5 µm; Mobile phase A: water; Mobile phase B: acetonitrile; Gradient: 20 % - 60% B for 8.5 minutes, then 60% - 95% B for 0.5 minutes, then 95% B for 1.0 minutes; Flow rate: 25 ml / minute) to obtain 3-methyl - N -[(4-methylphenyl)acetyl]-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl }-4-(trifluoromethyl)-L-prolinamide ( 92 ). Yield: 20.7 mg, 36.7 µmol, 31%. LCMS m/z 564.8 [M+H] ⁺ . Retention time: 2.71 minutes (Analytical conditions. Column: Waters Atlantis dC18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5.0% - 95% B, linear over 4.0 minutes, then 95% B over 1.0 minutes; Speed flow: 2 ml/minute).

Example 93

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-( 1H -pyrazol-1-yl) -N- (trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 93 )

Stage 1. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide, hydrochloride ( C67, HCl salt ).

To a solution of C66 (9.97 g, 24.4 mmol) in dichloromethane (50 ml) was added a solution of hydrochloride in 1,4-dioxane (4 M; 90 ml). The reaction mixture was stirred at room temperature (25° C.) for 2 hours, then LCMS analysis showed conversion to C67 : LCMS m/z 309.0 [M+H] ⁺ . Concentration in vacuo gives C67, the HCl salt as a white solid. Yield: 8.10 g, 23.5 mmol, 96%. ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 10.20-10.08 (m, 1H), 8.93 (d, J =8.0 Hz, 1H), 8.86-8.71 ( m, 1H), 7.68 (brs, 1H), 7.63 (brs, 1H), 7.12 (brs, 1H), 4.30 (ddd, J =10.9, 8.1, 4, 1 Hz, 1H), 4.09-4.02 (m, 1H), 3.63-3.53 (m, 1H, estimated; partially covered by water peak), 3.22-2.99 (m, 3H ), 2.34-2.22 (m, 1H), 2.21-2.11 (m, 1H), 2.01 (ddd, J = 13.6, 11.1, 3.6 Hz, 1H ), 1.80-1.66 (m, 3H), 1.55 (ddd, J = 13.6, 11.4, 4.1 Hz, 1H), 1.08 (s, 3H), 1, 05(s, 3H).

Stage 2. Synthesis of tert -butyl [(2 S )-1-[(1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-1-oxo-3-(1 H -pyrazol-1-yl)propan-2-yl]carbamate ( C80 ).

0°C C67 solution, HCl salts (300 mg, 0.870 mmol) and N- ( tert -butoxycarbonyl)-3-(1 H -pyrazol-1-yl)-L-alanine (222 mg, 0.870 mmol) in N, N -dimethylformamide (10 ml) was treated with O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 430 mg, 1.13 mmol) and 4-methylmorpholine (264 mg , 2.61 mmol). Then the reaction mixture was stirred at 0°C for 2 hours, it was diluted with water (50 ml) and extracted with ethyl acetate (20 ml). The aqueous layer was then saturated with the addition of solid sodium sulfate and extracted with a mixture of dichloromethane and methanol (10:1, 4×30 ml). The combined organic layers were concentrated in vacuo and subjected to silica gel chromatography twice (Gradient #1: 0%-10% methanol in dichloromethane; Gradient #2: 0%-25% methanol in dichloromethane) to give C80 as a bright yellow solid . Yield: 340 mg, 0.623 mmol, 72%. LCMS m/z 546.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 8.29 (brd, J = 8.6 Hz, 1H), 7.70-7.66 (m, 1H), 7.64 (s, 1H) , 7.51(brs, 1H), 7.38(brd, J =8.2Hz, 1H), 7.15-7.06(m, 2H), 6.23-6.19(m, 1H ), 4.53-4.43 (m, 1H), 4.34-4.18 (m, 3H), 4.17 (s, 1H), 3.76 (d, half of the AB quartet, J = 10 .5 Hz, 1H), 3.65 (dd, ABX system component, J = 10.4, 5.4 Hz, 1H), 3.20-3.06 (m, 2H), 2.36-2, 24 (m, 1H), 2.19-2.08 (m, 1H), 2.02-1.90 (m, 1H), 1.71-1.55 (m, 2H), 1.53- 1.46 (m, 1H), 1.39 (d, half AB quartet, J = 7.6 Hz, 1H), 1.30 (s, 9H), 1.01 (s, 3H), 0.89 (s, 3H).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-[3-( 1H -pyrazol-1-yl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide hydrochloride ( C81 ).

A solution of the hydrochloride in 1,4-dioxane (4 M; 15 ml) was added to a 0° C. solution of C80 (340 mg, 0.623 mmol) in dichloromethane (4 ml). The reaction mixture is then stirred at 20°C for 1 hour, it is filtered and the filter cake is washed with dichloromethane (3×10 ml). The combined filtrates are concentrated in vacuo to give C81 as a white solid. Yield: 244 mg, 0.506 mmol, 81%. LCMS m/z 446.0 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 8.57-8.48 (m, 3H), 8.42 (bd, J =8.6 Hz, 1H), 7.81 ( d, J = 2.3 Hz, 1H), 7.65 (brs, 1H), 7.58 (d, J = 1.9 Hz, 1H), 7.35 (brs, 1H), 7.09 ( brs, 1H), 6.27 (dd, J =2.2 Hz, 1H), 4.59 (dd, ABX system component, J =14.4, 4.7 Hz, 1H), 4.54-4 .41 (m, 2H), 4.36-4.25 (m, 1H), 4.30 (s, 1H), 3.66-3.60 (m, 1H), 3.42-3.33 (m, 1H), 3.21-3.03 (m, 2H), 2.31-2.20 (m, 1H), 2.18-2.06 (m, 1H), 1.97 (ddd , J = 13.5, 11.5, 3.7 Hz, 1H), 1.46 (dd, ABX system component, J = 7.7, 5.3 Hz, 1H), 1.41 (d, half AB quartet, J = 7.7 Hz, 1H), 1.01 (s, 3H), 0.92 (s, 3H).

Stage 4. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl 3-[3-( 1H -pyrazol-1-yl) -N- (trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 93 ).

A 0°C solution of C81 (120 mg, 0.249 mmol) in dichloromethane (6.0 mL) was treated with pyridine (170 mg, 2.15 mmol) followed by a solution of trifluoroacetic anhydride (158 mg, 0.752 mmol) in dichloromethane (2.0 ml). The reaction mixture is then heated to 20°C and stirred for 3 hours, then it is diluted with water (20 ml) and extracted with dichloromethane (3×20 ml). The combined organic layers are washed with hydrochloric acid (1 M; 20 ml) and saturated aqueous sodium chloride solution (2×20 ml), dried, filtered, and concentrated in vacuo . Chromatography on silica gel (Gradient: 0% - 10% methanol in dichloromethane), then supercritical liquid chromatography [Column: Chiral Technologies Chiralpak AD, 30×250 mm, 10 μm; Mobile phase: 4:1 carbon dioxide/(ethanol containing 0.1% ammonium hydroxide); Flow rate: 60 ml/minute] gives (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl }-6,6-dimethyl-3-[3-(1 H -pyrazol-1-yl)- N- (trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 93 ) as a white solid. Yield: 15.0 mg, 28.6 µmol, 11%. LCMS m/z 524.0 [M+H] ⁺ . ¹ H NMR (500 MHz, DMSO- d ₆ ) δ 10.03 (br s, 1H), 8.92 (d, J =8.0 Hz, 1H), 7.74 (br s, 1H), 7.69 (d, J =2.3 Hz, 1H), 7.51 (d, J =1.9 Hz, 1H), 6.23 (dd, J =2.2 Hz, 1H), 5.01-4 .93 (m, 1H), 4.91-4.83 (m, 1H), 4.49-4.39 (m, 2H), 4.13 (s, 1H), 3.73 (dd, component ABX system, J = 10.4, 5.4 Hz, 1H), 3.60 (d, half AB quartet, J = 10.4 Hz, 1H), 3.21-3.09 (m, 2H), 2.40-2.31 (m, 1H), 2.20-2.10 (m, 2H), 1.79 (ddd, J =13.7, 9.5, 6.8 Hz, 1H), 1.76-1.66 (m, 1H), 1.56 (dd, J = 7.5, 5.4 Hz, 1H), 1.36 (d, half AB quartet, J = 7.6 Hz, 1H), 1.02 (s, 3H), 0.85 (s, 3H).

Example 94

(1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[(2 S )- 4,4-difluoro-2-(2,2,2-trifluoroacetamido)butanoyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 94 )

Stage 1. Synthesis of 9 H -fluoren-9-ylmethyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3 -[( 3S )-2-oxopyrrolidin-3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-4,4 -difluoro-1-oxobutan-2-yl}carbamate ( C82 ).

To a 0°C solution of C67 (230 mg, 0.569 mmol) and (2S)-2-{[(9H- fluoren -9- ylmethoxy )carbonyl]amino}-4,4-difluorobutanoic acid (247 mg, 0.684 mmol ) in N, N -dimethylformamide (5 ml) add O - (7-azabenzotriazol-1-yl) - N, N, N', N' -tetramethyluronium hexafluorophosphate (GATU; 281 mg, 0.739 mmol) in one portion; then 4-methylmorpholine (173 mg, 1.71 mmol) is added dropwise. Then the reaction mixture was stirred at 0°C for 10 minutes, it was warmed to room temperature (20°C) and stirring continued for 2 hours, then the reaction mixture was poured into ice water (15 ml) and extracted with ethyl acetate (2×15 ml ). The combined organic layers are washed successively with 1 M hydrochloric acid, saturated aqueous sodium bicarbonate and saturated aqueous sodium chloride, dried over sodium sulfate, filtered, concentrated in vacuo , and purified by silica gel chromatography (Gradient: 0% - 10% methanol in dichloromethane) to give C82 as a white solid. Yield: 245 mg, 0.376 mmol, 66%. LCMS m/z 652.5 [M+H] ⁺ . ¹ H NMR (400 MHz, chloroform- d ) δ 8.86 (br s, 1H), 7.74 (d, J =7.6 Hz, 2H), 7.58-7.51 (m, 2H), 7.38 (dd, J = 7.4, 7.4 Hz, 2H), 7.33-7.25 (m, 2H, estimated; partially covered by solvent peak), 7.08 (br, 1H), 6 .49-6.32(m, 1H), 6.17-5.79(m, 3H), 4.78-4.67(m, 1H), 4.41-4.24(m, 4H) , 4.22-4.07 (m, 2H), 3.83 (d, J =10.5 Hz, 1H), 3.45-3.32 (m, 2H), 2.62-2.36 (m, 3H), 2.26-2.10 (m, 2H), 1.99-1.83 (m, 2H), 1.53 (d, half AB quartet, J = 7.6 Hz, 1H ), 1.51-1.44 (m, 1H), 1.03 (s, 3H), 0.89 (s, 3H).

Stage 2. Synthesis of (1 R ,2 S ,5 S )-3-[(2 S )-2-amino-4,4-difluorobutanoyl]- N -{(2 S )-1-amino-1-oxo- 3-[( 3S )-2-oxopyrrolidin-3-yl]propan-2-yl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( C83 ).

To a 0°C suspension of C82 (195 mg, 0.299 mmol) in dichloromethane (3 ml) was added a solution of diethylamine (32.8 mg, 0.448 mmol) in dichloromethane (0.5 ml). The reaction mixture is stirred at 30°C for 16 hours, then it is combined with a similar reaction carried out using C82 (50 mg, 77 μmol) and concentrated in vacuo . Purification by chromatography on silica gel [Gradient: 0% - 10% (10:1 mixture of methanol and ammonium hydroxide) in dichloromethane] gives C83 as a colorless gum. Combined yield: 149 mg, 0.347 mmol, 92%. LCMS m/z 452.3 [M+Na ⁺ ]. ¹ H NMR (400 MHz, chloroform- d ) δ 8.73 (bd, J = 5.3 Hz, 1H), 7.11 (brb, 1H), 6.03 (tdd, J = 56.8, 6 .2, 3.0Hz, 1H), 5.64(brs, 1H), 5.28(brs, 1H), 4.35-4.27(m, 1H), 4.27(s, 1H) , 4.11 (dd, J =10.3, 5.5 Hz, 1H), 3.72 (dd, J =9.3, 4.1 Hz, 1H), 3.61 (d, J =10 .3 Hz, 1H), 3.41-3.35 (m, 2H), 2.53-2.36 (m, 3H), 2.20-2.11 (m, 1H), 2.00- 1.84 (m, 3H), 1.6-1.45 (m, 2H, estimated; partially covered by water peak), 1.06 (s, 3H), 0.94 (s, 3H).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[( 2 S )-4,4-difluoro-2-(2,2,2-trifluoroacetamido)butanoyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 94 ).

To a 0°C solution of C83 (99 mg, 0.23 mmol) in dichloromethane (4 ml) was added pyridine (146 mg, 1.85 mmol) and a solution of trifluoroacetic anhydride (194 mg, 0.924 mmol) in dichloromethane (2 ml). The reaction mixture was stirred at room temperature (15°C) for 20 hours, treated with additional pyridine (30 mg, 0.38 mmol) and stirred at room temperature (15°C) for another 16 hours. It was then partitioned between dichloromethane (15 ml) and hydrochloric acid (1 M; 15 ml) and the organic layer was washed with saturated aqueous sodium bicarbonate (15 ml) and saturated aqueous sodium chloride (10 ml), dried, filtered, and concentrated in a vacuum . The residue was combined with the product from a similar reaction using C83 (50 mg, 0.12 mmol) and purified using reverse phase HPLC (Column: Waters XBridge BEH C18, 25x150 mm, 5 µm; Mobile phase A: water containing 0.05% ammonium hydroxide (v/v) Mobile phase B: acetonitrile Gradient: 23% - 63% B). (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[(2 S )- 4,4-difluoro-2-(2,2,2-trifluoroacetamido)butanoyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 94 ) is isolated as a white solid . Combined yield: 28.8 mg, 56.8 µmol, 16%. LCMS m/z 508.0 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 10.01 (br s, 1H), 8.96 (d, J =8.0 Hz, 1H), 7.72 (s, 1H), 6.16 (tt, J =55.9, 4.6 Hz, 1H), 4.99-4.90 (m, 1H), 4.65-4.57 (m, 1H), 4.14 (s, 1H ), 3.85 (dd, component of the ABX system, J = 10.2, 5.4 Hz, 1H), 3.67 (d, half of the AB quartet, J = 10.3 Hz, 1H), 3.21- 3.04 (m, 2H), 2.42-2.19 (m, 3H), 2.17-2.04 (m, 2H), 1.83-1.64 (m, 2H), 1, 60 (dd, J =7.6, 5.3 Hz, 1H), 1.35 (d, half AB quartet, J =7.6 Hz, 1H), 1.03 (s, 3H), 0.89 (s, 3H).

Example 95

N- (Methoxycarbonyl)-3-methyl-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- 4-(trifluoromethyl)-L-prolinamide ( 95 )

Step 1. Synthesis of (4 R )-1-( tert -butoxycarbonyl)-4-(trifluoromethyl)-L-prolyl-3-[(3 S )-2-oxopyrrolidin-3-yl]-L-alanineamide ( C50 ) .

N,N -diisopropylethylamine (14.8 mL, 85.0 mmol) was added to a -30°C mixture of ( 4R )-1-( tert -butoxycarbonyl)-4-(trifluoromethyl)-L-proline (8.00 g , 28.2 mmol), C16, HCl salts (6.45 g, 31.1 mmol) and O- (7-azabenzotriazol-1-yl) -N, N,N',N'- tetramethyluronium hexafluorophosphate (GATU; 11.8 g, 31.0 mmol) in N,N -dimethylformamide (100 ml). The reaction mixture is heated to 0°C for 1 hour, then LCMS analysis shows the presence of C50 : LCMS m/z 437.3 [M+H] ⁺ . An aqueous solution of sodium bicarbonate (300 ml) was added and the resulting mixture was extracted with a mixture of 2-propanol and dichloromethane (1:4, 5×100 ml); the combined organic layers were concentrated in vacuo and purified by silica gel chromatography (Gradient: 0% - 100% methanol in dichloromethane) to give C50 as an oil. ¹ H NMR analysis shows that this product exists as a mixture of rotamers. Yield: 10.9 g, 25.0 mmol, 89%. ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ [8.28 (d, J =8.5 Hz) and 8.22 (d, J =8.2 Hz), 1H total], [7.64 (s) and 7.59 (s), 1H total], [7.38 (br) and 7.27 (br) 1H total], 7.05 (br, 1H), 4.38 -4.28(m, 1H), 4.28-4.17(m, 1H), 3.45-3.36(m, 1H), 3.12-3.00(m, 1H), 2 .42-2.03 (m, 4H), 2.02-1.89 (m, 1H), 1.80-1.45 (m, 2H), [1.39 (s) and 1.32 ( c), total 9H].

Step 2. Synthesis of N- ( tert -butoxycarbonyl)-3-methyl-L-valyl-(4 R )-4-(trifluoromethyl)-L-prolyl-3-[(3 S )-2-oxopyrrolidin-3-yl ]-L-alanine amide ( C51 ).

A solution of hydrochloride in 1,4-dioxane (4 M; 80 ml) was added to a solution of C50 (7.00 g, 16.0 mmol) in dichloromethane (15 ml). The reaction mixture is then stirred at room temperature for 5 minutes, concentrated in vacuo to remove the solvent, and then suctioned off under high vacuum to remove residual hydrochloride. The residue is mixed with N- ( tert -butoxycarbonyl)-3-methyl-L-valine (4.08 g, 17.6 mmol) and hexafluorophosphate O- (7-azabenzotriazol-1-yl) -N, N, N', N' -tetramethyluronium (HATU; 6.71 g, 17.6 mmol) in N,N -dimethylformamide (25 ml), cooled to -30°C and treated with N,N -diisopropylethylamine (8.38 ml, 48.1 mmol). The reaction mixture is heated to 0°C for 1 hour, then LCMS analysis shows the presence of C51 : LCMS m/z 550.4 [M+H] ⁺ . The reaction mixture was then diluted with aqueous sodium bicarbonate and extracted three times with a 4:1 mixture of dichloromethane and 2-propanol. The combined organic layers are then concentrated in vacuo , the residue is purified by silica gel chromatography (Gradient: 0% - 30% methanol in dichloromethane) to give C51 as a solid. Yield: 3.95 g, 7.19 mmol, 45%, ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks: δ 8.28 (d, J =8.8 Hz, 1H), 7, 55(s, 1H), 7.29(brs, 1H), 7.03(brs, 1H), 6.77(brd, J =9.1Hz, 1H), 4.52-4.43(m , 1H), 4.24 (ddd, J =12.2, 8.7, 3.5 Hz, 1H), 4.12 (d, J =9.2 Hz, 1H), 4.02-3, 84 (m, 2H), 3.16-3.08 (m, 1H), 3.08-2.98 (m, 1H), 2.50-2.37 (m, 1H), 2.31- 2.20 (m, 1H), 2.19-2.05 (m, 2H), 2.00-1.87 (m, 1H), 1.69-1.55 (m, 1H), 1, 55-1.44 (m, 1H), 1.36 (s, 9H), 0.93 (s, 9H).

Stage 3. Synthesis of N -(methoxycarbonyl)-3-methyl-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl ]ethyl}-4-(trifluoromethyl)-L-prolinamide ( 95 ).

A mixture of C51 (230 mg, 0.418 mmol) and a solution of hydrochloride in 1,4-dioxane (4 M; 2 ml, 8 mmol) was stirred at room temperature for 5 minutes, then the reaction mixture was concentrated in vacuo to remove the solvent, then it was filtered with suction through high vacuum to remove residual hydrochloride. The residue was dissolved in dichloromethane (2 ml), cooled to 0°C and treated with N,N -diisopropylethylamine (0.219 ml, 1.26 mmol) then methyl chloroformate (59.3 mg, 0.628 mmol). The reaction mixture was then stirred at 0° C. for 10 minutes, diluted with an aqueous sodium bicarbonate solution and extracted three times with a 4:1 mixture of dichloromethane and 2-propanol; the combined organic layers are dried over magnesium sulfate, filtered, and concentrated in vacuo . The residue is dissolved in dichloromethane (3 ml); after adding methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 299 mg, 1.25 mmol), the reaction mixture was stirred at room temperature for 1 hour. It is then treated with dilute sodium carbonate aqueous solution and extracted 3 times with ethyl acetate. The combined organic layers are dried over magnesium sulfate, filtered, and purified by silica gel chromatography (Gradient: 50% - 100% ethyl acetate in heptane). The resulting product is suspended in heptane (4 ml) at 50°C for 2 hours, cooled to room temperature and stirred at room temperature overnight; collection of the solid yields N- (methoxycarbonyl)-3-methyl-L- valyl- (4R) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl ]ethyl}-4-(trifluoromethyl)-L-prolinamide ( 95 ) as a solid. Yield: 123 mg, 0.251 mmol, 60%. LCMS m/z 490.4 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.02 (d, J =8.6 Hz, 1H), 7.65 (s, 1H), 7.27 (br, J =8.7 Hz , 1H), 4.94 (ddd, J =11.1, 8.5, 5.0 Hz, 1H), 4.36 (dd, J =7.3, 7.2 Hz, 1H), 4, 14 (d, J = 8.7 Hz, 1H), 4.00-3.91 (m, 2H), 3.52 (s, 3H), 3.46-3.34 (m, 1H, estimated; partially covered by water peak), 3.18-2.98 (m, 2H), 2.5-2.39 (m, 1H, estimated; partially covered by solvent peak), 2.35-2.23 (m, 1H ), 2.22-2.01 (m, 3H), 1.77-1.63 (m, 2H), 0.94 (s, 9H).

Example 96

(1 R ,2 S ,5 S )- N -{(1 R )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 96 ) and ( 1R , 2S , 5S ) -N -{( 1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl] -3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ), Solid Form 5

To a solution of 13, methyl t-butyl ether solvate ( from the alternative synthesis from Example 13, methyl t -butyl ether solvate; 15.0 g, 25.5 mmol) in acetonitrile (80 mL) was added methanesulfonic acid (6.4 mL, 99 mmol). The reaction mixture was stirred at room temperature for 4 hours, then made basic by adding a mixture of saturated aqueous sodium bicarbonate (80 ml) and saturated aqueous sodium chloride (10 ml). The resulting mixture was extracted with dichloromethane (2×100 ml) and the combined organic layers were dried over sodium sulfate, filtered, and concentrated in vacuo . The separation of the two epimers is carried out by supercritical liquid chromatography (Column: Chiral Technologies Chiralcel OX-H, 30×250 mm, 5 µm; Mobile phase: 9:1 carbon dioxide/2-propanol; Back pressure: 100 bar; Flow rate: 80 ml/ minute). The first product eluted is reduced ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-6,6 -dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide ( 13 ). According to powder x-ray diffraction, this product is amorphous; this is referred to as Solid Form 5. The second eluted product is obtained as a glass, which is dissolved in dichloromethane, treated with heptane and concentrated in vacuo to give (1 R ,2 S ,5 S ) -N -{(1 R )-1-cyano -2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3, 1.0]hexane-2-carboxamide ( 96 ) as a solid.

Reduced 13 - Yield: 6.00 g, 12.0 mmol, 47%. LCMS m/z 500.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.46-9.33 (m, 1H), 9.01 (d, J =8.5 Hz, 1H), 7.66 (s, 1H) , 5.03-4.91 (m, 1H), 4.46-4.37 (m, 1H), 4.16 (s, 1H), 3.97-3.86 (m, 1H), 3 .69 (d, half AB quartet, J = 10.4 Hz, 1H), 3.19-3.09 (m, 1H), 3.09-2.98 (m, 1H), 2.46-2 .33 (m, 1H), 2.21-2.03 (m, 2H), 1.79-1.65 (m, 2H), 1.61-1.53 (m, 1H), 1.32 (d, half AB quartet, J = 7.7 Hz, 1H), 1.03 (s, 3H), 0.98 (s, 9H), 0.85 (s, 3H). Retention time: 3.93 minutes (Analytical conditions. Column: Chiral Technologies Chiralcel OX-H, 4.6×250 mm, 5 µm; Mobile phase A: carbon dioxide; Mobile phase B: 2-propanol; Gradient: 5% B for 1.00 minutes, then 5% - 60% B for 8.00 minutes; Back pressure: 120 bar; Flow rate: 3.0 ml/minute). The powder X-ray diffraction pattern for this amorphous product is given in Figure 9. The method for collecting powder X-ray diffraction data is described in Alternative Synthesis of Example 13, Methyl tert -butyl ether solvate, Step 8.

96 - Yield: 2.58 g, 5.16 mmol, 20%. LCMS m/z 500.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 9.42 (d, J =8.4 Hz, 1H), 9.06 (d, J =7.6 Hz, 1H), 7.79 (s , 1H), 4.96-4.86 (m, 1H), 4.41 (d, J = 8.7 Hz, 1H), 4.20 (s, 1H), 3.92 (bdd, J = 10.7, 5.4 Hz, 1H), 3.66 (d, half AB quartet, J = 10.5 Hz, 1H), 3.22-3.12 (m, 2H), 2.43-2 .31 (m, 1H), 2.31-2.20 (m, 1H), 2.16-2.04 (m, 1H), 1.84-1.63 (m, 2H), 1.57 -1.49 (m, 1H), 1.32 (d, half AB quartet, J =7.7 Hz, 1H), 1.00 (brs, 12H), 0.84 (s, 3H). Retention time: 4.20 minutes (Analytical conditions identical to those used for reconstituted 13 above).

Example 97

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C86 (DIAST-2) ( 97 )

Step 1. Synthesis of 2-[( tert -butoxycarbonyl)amino]-3-(trifluoromethyl)pentanoic acid ( C84 ).

An aqueous solution of sodium hydroxide (1 M; 1.48 ml, 1.48 mmol) is added to a suspension of 2-amino-3-(trifluoromethyl)pentanoic acid (Wang et al., J. Amer. Chem. Soc . 2003 , 125 , 6900-6906; 137 mg, 0.740 mmol) in 1,4-dioxane (3 ml) and the resulting mixture was cooled to 0°C. Di- tert -butyl dicarbonate (0.204 ml, 0.888 mmol) was added slowly, then the reaction mixture was stirred at room temperature overnight. After dilution with ethyl acetate, the reaction mixture was cooled in an ice bath and then acidified to pH 2 by adding 1 M aqueous potassium hydrogen sulfate solution. The aqueous layer is extracted twice with ethyl acetate and the combined organic layers are dried over magnesium sulfate, filtered, and concentrated in vacuo to give C84 as a solid. This product is believed to be a mixture of 4 diastereomers, potentially showing rotamers as well. Yield: 197 mg, 0.690 mmol, 93%. LCMS m/z 284.3 [M−H] ⁻ . ¹ H NMR (400 MHz, DMSO- d ₆ ) δ 13.16 (br s, 1H), [7.29 (d, major, J =9.8 Hz) and 6.95-6.85 (m, minor ), total 1H], [4.55 (dd, major, J = 9.8, 3.3 Hz), 4.46 (bd, minor, J = 9.1 Hz), and 4.40 (dd, negligible, J = 9.4, 4.5 Hz), total 1H], 2.86-2.67 (m, 1H), 1.71-1.47 (m, 2H), 1.39 (brs, 9H), [0.98 (t, minor, J =7.4 Hz) and 0.91 (t, major, J =7.5 Hz), total 3H].

Stage 2. Synthesis of tert -butyl {1-[(1 R ,2 S ,5 S )-2-({(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidine -3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-1-oxo-3-(trifluoromethyl)pentan-2- yl} carbamate , DIAST -1 ( C85 ) and tert -butyl {1-[(1R,2S, 5S )-2-({( 2S )-1-amino-1-oxo-3-[( 3 S )-2-oxopyrrolidin-3-yl]propan-2-yl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-1-oxo-3- (trifluoromethyl)pentan-2-yl}carbamate, DIAST-2 ( C86 ).

A 0°C solution of C84 (128 mg, 0.449 mmol) in a mixture of acetonitrile (2.7 mL) and N,N -dimethylformamide (1.5 mL) is treated with O- (7-azabenzotriazol-1-yl) -N,N hexafluorophosphate ,N',N'- tetramethyluronium (GATU; 176 mg, 0.463 mmol) and 4-methylmorpholine (0.116 ml, 1.06 mmol). The reaction mixture is then stirred at 0° C. for 30 minutes, C67 (170 mg, 0.420 mmol) is added as a solid and stirring is continued for 2 hours. The reaction mixture was then diluted with ethyl acetate and water, and washed with saturated aqueous sodium bicarbonate. The aqueous layer was extracted twice with ethyl acetate, and the combined organic layers were washed with saturated aqueous sodium chloride, dried over magnesium sulfate, filtered, and concentrated in vacuo . The resulting oil was azeotropically distilled twice with heptane and twice with methyl t- butyl ether, then subjected to silica gel chromatography (Gradient: 0% - 20% methanol in dichloromethane). The first eluted diastereomer is designated C85 and the second eluted diastereomer is designated C86 .

C85 (DIAST-1)- Yield: 77.3 mg, 0.134 mmol, 32%. This product contains a mixture of isomers or rotamers according to^oneH NMR analysis. HCMSm/z 576.2 [M+H]⁺.^oneH NMR (400 MHz, DMSO-d ₆), characteristic peaks, integrations are approximate: δ [8.35 (d,J= 7.9 Hz) and 8.16 (d,J= 8.5 Hz), 1H total], 7.62-7.54 (m, 1H), 7.41-7.18 (m, 2H), [7.02 (br) and 6.98 (br ), total 1H], 4.59-4.50 (m, 1H), 4.29-4.13 (m, 2H), 3.89 (dd,J=10.4, 5.4 Hz, 1H), 3.44 (d,J=10.4 Hz, 1H), 3.20-3.04 (m, 2H), 2.70-2.59 (m, 1H), 2.43-2.31 (m, 1H), 2, 21-2.08(m, 1H), 1.99-1.88(m, 1H), [1.38(s) and 1.36(s), total 9H], 1.01(brs, 3H ), 0.94-0.82 (m, 6H).

C86 (DIAST-2)- Yield: 87.8 mg, 0.153 mmol, 36%. This product is for the most part the only isomer in terms of^oneH NMR analysis. HCMSm/z 576.2 [M+H]⁺.^oneH NMR (400 MHz, DMSO-d ₆), characteristic peaks: δ 8.27 (d,J= 8.6 Hz, 1H), 7.59 (s, 1H), 7.31 (d,J=9.5 Hz, 1H), 7.22 (ws, 1H), 7.04 (ws, 1H), 4.55 (dd,J=9.5, 5.9 Hz, 1H), 4.30-4.17 (m, 1H), 4.23 (s, 1H), 3.79 (dd, ABX system component,J=10.1, 5.2 Hz, 1H), 3.71 (d, half AB quartet,J=10.1 Hz, 1H), 3.09-2.99 (m, 1H), 2.68-2.55 (m, 1H), 2.42-2.30 (m, 1H), 2, 17-2.04 (m, 1H), 1.97-1.86 (m, 1H), 1.36 (s, 9H), 1.02 (s, 3H), 0.94-0.83 ( m, 6H).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-[2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C86 (DIAST-2) ( C87 ).

A solution of C86 (DIAST-2) (87.8 mg, 0.153 mmol) in dichloromethane (1 ml) was treated with a solution of hydrochloride in 1,4-dioxane (4 M; 0.381 ml, 1.52 mmol). After the reaction mixture was stirred at room temperature for 40 minutes, methanol (0.5 ml) was added to improve solubility. After another 40 minutes, add a solution of hydrochloride in 1,4-dioxane (4 M; 0.10 ml, 0.4 mmol); after 30 minutes, LCMS analysis shows complete deprotection: LCMS m/z 476.2 [M+H] ⁺ . The reaction mixture is concentrated in vacuo and azeotropically distilled twice with heptane; the residue is triturated twice with diethyl ether, suspended in dichloromethane (1.2 ml) and cooled to 0°C. After adding triethylamine (42.4 µl, 0.304 mmol) followed by trifluoroacetic anhydride (47.9 µl, 0.339 mmol), the reaction mixture was stirred at 0°C for 30 minutes, then it was removed from the ice bath and partitioned between water and ethyl acetate . The aqueous layer was extracted with ethyl acetate and the combined organic layers were washed with saturated aqueous sodium bicarbonate and saturated aqueous sodium chloride, dried over magnesium sulfate, filtered, concentrated in vacuo , and azeotropically distilled twice with methyl t- butyl ether. According to ¹ H NMR and LCMS analysis, this product contains a mixture of C87 and the corresponding methyl ester (LCMS m/z 587.4 [M+H] ⁺ ). ¹ H NMR (400 MHz, DMSO- d ₆ ), characteristic peaks for C87 : δ 8.31 (d, J = 8.7 Hz, 1H), 7.59 (s, 1H), 7.26 (s, 1H), 7.05 (s, 1H), 3.59 (d, half AB quartet, J = 10.0 Hz, 1H), 1.97-1.87 (m, 1H), 1.40 (d , half AB quartet, J = 7.6 Hz, 1H), 1.03 (s, 3H), 0.84 (s, 3H). Purification by chromatography on silica gel (Gradient: 0% - 20% methanol in dichloromethane), then azeotroping the resulting oil with heptane, then azeotroping with a mixture of diethyl ether and heptane, gives C87 as a white solid. Yield: 17.9 mg, 31.3 µmol, 20%. LCMS m/z 572.0 [M+H] ⁺ .

Stage 4. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl 3-[2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C86 (DIAST-2) ( 97 ).

Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 18.8 mg, 78.9 µmol) was added to a solution of C87 (18 mg, 31 µmol) in ethyl acetate (0.8 ml). After the reaction mixture was stirred at room temperature for 1 hour, methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent) was added again at the end of a spatula. Stirring is continued for 2 hours, then the reaction mixture is filtered and the filter cake is washed with ethyl acetate. The combined filtrates are washed with saturated aqueous sodium bicarbonate and the aqueous layer is extracted once with ethyl acetate; the combined organic layers are washed with saturated aqueous sodium chloride, dried over magnesium sulfate, filtered, and concentrated in vacuo to give the crude product. ¹ H NMR (600 MHz, DMSO- d ₆ ), main component: δ 9.90 (d, J =8.8 Hz, 1H), 8.85 (d, J =8.1 Hz, 1H), 7 .56 (brs, 1H), 4.97-4.89 (m, 2H), 4.17 (s, 1H), 3.89 (dd, J =10.1, 5.4 Hz, 1H), 3.62 (d, J = 10.1 Hz, 1H), 3.21-3.14 (m, 1H), 3.12-3.06 (m, 1H), 2.92-2.82 ( m, 1H), 2.43-2.35 (m, 1H), 2.18-2.10 (m, 2H), 1.78 (ddd, J =13.6, 9.6, 6.0 Hz, 1H), 1.75-1.66 (m, 2H), 1.62-1.55 (m, 2H), 1.35 (d, half AB quartet, J = 7.7 Hz, 1H) , 1.04 (s, 3H), 0.97-0.92 (t, J= 7.6 Hz, 3H), 0.85 (s, 3H). This product was purified by reverse phase HPLC (Column: Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: 0.05% trifluoroacetic acid in water (v/v); Mobile phase B: 0.05% trifluoroacetic acid in acetonitrile (v/v); Gradient: 5% - 95% B over 8.54 minutes, then 95% B over 1.46 minutes; Flow rate: 25 ml/minute) to give (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[2 -(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C86 (DIAST-2) ( 97 ). Yield: 8.3 mg, 15 µmol, 48%. LCMS m/z 554.6 [M+H] ⁺ . Retention time: 2.72 minutes (Analytical conditions. Column: Waters Atlantis dC18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5.0% - 95% B, linear over 4.0 minutes, then 95% B over 1.0 minute; Flow rate : 2 ml/minute).

Example 98

(1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C85 (DIAST-1) ( 98 )

Stage 1. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-3-[2-amino-3-(trifluoromethyl)pentanoyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide, hydrochloride, from C85 (DIAST-1) ( C88 ).

A solution of hydrochloride in 1,4-dioxane (4 M; 0.336 ml, 1.34 mmol) was added to a solution of C85 (DIAST-1) (77.3 mg, 0.134 mmol) in dichloromethane (1 ml). After the reaction mixture was stirred at room temperature for 40 minutes, methanol (0.5 ml) was added to improve solubility. Stirring is continued for 2 hours, then LCMS analysis shows completion of deprotection: LCMS m/z 476.2 [M+H] ⁺ . The reaction mixture is concentrated in vacuo ; the residue is azeotropically distilled twice with heptane, then triturated twice with diethyl ether to give C88 as a white solid. Yield: 54.5 mg, 0.106 mmol, 79%. ¹ H NMR (400 MHz, DMSO- d ₆ ), main characteristic peaks: δ 8.53 (br s, 3H), 8.36 (d, J =8.5 Hz, 1H), 7.61 (s, 1H ), 7.28 (shs, 1H), 7.06 (shs, 1H), 4.25 (ddd, J =10.9, 8.4, 4.5 Hz, 1H), 4.17 (s, 1H), 4.08 (dd, J =10.5, 5.6 Hz, 1H), 2.80-2.68 (m, 1H), 2.37-2.26 (m, 1H), 2 .23-2.13 (m, 1H), 2.05-1.96 (m, 1H), 1.73-1.51 (m, 5H), 1.44 (d, half of the AB quartet, J = 7.7 Hz, 1H), 1.04 (s, 3H), 0.90 (s, 3H).

Stage 2. Synthesis of (1 R ,2 S ,5 S )- N -{(2 S )-1-amino-1-oxo-3-[(3 S )-2-oxopyrrolidin-3-yl]propan-2 -yl}-6,6-dimethyl-3-[2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C85 (DIAST-1) ( C89 ).

A 0°C suspension of C88 (54.5 mg, 0.106 mmol) in dichloromethane (1 ml) was treated with triethylamine (26 μl, 0.19 mmol) followed by trifluoroacetic anhydride (19.5 μl, 29.1 mg, 0.138 mmol). Then the reaction mixture is stirred at 0°C for 1 hour and 10 minutes, trifluoroacetic anhydride (1 equivalent) is added; after 30 minutes, trifluoroacetic anhydride (9.4 μl, 67 μmol) is added again. Stirring is continued for 45 minutes, then LCMS analysis shows complete conversion to C89 : LCMS m/z 572.4 [M+H] ⁺ . The reaction mixture is partitioned between water and ethyl acetate, and the aqueous layer is extracted with ethyl acetate. The combined organic layers are washed successively with saturated aqueous sodium bicarbonate and saturated aqueous sodium chloride, dried over magnesium sulfate, filtered and concentrated in vacuo to give C89 . Yield: 41.2 mg, 72.1 µmol, 68%. ¹ H NMR (400 MHz, DMSO- d ₆ ), main component, characteristic peaks: δ 10.04 (d, J =9.3 Hz, 1H), 8.23 (d, J =8.5 Hz, 1H ), 7.58(s, 1H), 7.31(brs, 1H), 7.01(brs, 1H), 4.92-4.83(m, 1H), 4.23(s, 1H) , 3.95 (dd, J =10.2, 5.5 Hz, 1H), 2.98-2.86 (m, 1H), 2.38-2.27 (m, 1H), 1.90 (ddd, J =13.5, 11.2, 4.0 Hz, 1H), 1.39 (d, half AB quartet, J =7.7 Hz, 1H), 1.02 (s, 3H), 0.90 (s, 3H).

Stage 3. Synthesis of (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl 3-[2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C85 (DIAST-1) ( 98 ).

Methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent; 42.7 mg, 0.179 mmol) was added to a solution of C89 (41.0 mg, 71.7 µmol) in ethyl acetate (0.8 ml). After the reaction mixture was stirred at room temperature for 1 hour, methyl N- (triethylammoniosulfonyl)carbamate, internal salt (Burgess reagent) was added at the tip of a spatula. Stirring is continued for 2 hours, then the reaction mixture is filtered and the filter cake is washed with ethyl acetate. The combined filtrates are washed with saturated aqueous sodium bicarbonate and the aqueous layer is extracted with ethyl acetate; the combined organic layers are washed with saturated aqueous sodium chloride, dried over magnesium sulfate, filtered, and concentrated in vacuo to give the crude product. ¹ H NMR (600 MHz, DMSO- d ₆ ), main component, characteristic peaks: δ 10.12 (d, J =9.1 Hz, 1H), 8.99 (d, J =8.1 Hz, 1H ), 7.70 (s, 1H), 4.94 (ddd, J =9.4, 8.1, 6.5 Hz, 1H), 4.87 (dd, J =9.0, 9.0 Hz, 1H), 4.11 (s, 1H), 3.96 (dd, J =10.2, 5.6 Hz, 1H), 3.52 (d, J =10.0 Hz, 1H), 3.18-3.07 (m, 2H), 2.98-2.88 (m, 1H), 2.40-2.33 (m, 1H), 1.79-1.52 (m, 4H ), 1.61 (dd, J =7.6, 5.5 Hz, 1H), 1.33 (d, half AB quartet, J =7.7 Hz, 1H), 1.04 (s, 3H) , 0.91-0.86 (m, 6H).

Purification of this reverse phase HPLC product (Column: Waters Sunfire C18, 19×100 mm, 5 µm; Mobile phase A: 0.05% trifluoroacetic acid in water; Mobile phase B: 0.05% trifluoroacetic acid in acetonitrile; Gradient: 5% - 95% B for 8.54 minutes, then 95% B for 1.46 minutes; Flow rate: 25 ml/minute) gives (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl )pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C85 (DIAST-1) ( 98 ). Yield: 4.3 mg, 7.8 µmol, 11%. LCMS m/z 554.6 [M+H] ⁺ . Retention time: 2.80 minutes (Analytical conditions. Column: Waters Atlantis dC18, 4.6×50 mm, 5 µm; Mobile phase A: water containing 0.05% trifluoroacetic acid (v/v); Mobile phase B: acetonitrile containing 0.05% trifluoroacetic acid (v/v) Gradient: 5.0% - 95% B, linear over 4.0 minutes, then 95% B over 1.0 minute; Flow rate : 2 ml/minute).

Preparation of 3- tert -butyl 2-methyl(1R,2S, 5S )-6,6-dimethyl-3-azabicyclo[ 3.1,0 ]hexane-2,3- dicaroxylate ( C90 )

This preparation is carried out using the general procedure described by C. Uyeda and J. Werth, Angew. Chem. Int. Ed . 2018 , 57 , 13902-13906. A 3-necked flask equipped with a magnetic stirrer, reflux condenser, thermometer, and nitrogen inlet was charged with cobalt(II) bromide (0.15 equiv; 0.146 g, 0.667 mmol), ( 1E.1'E )-1.1' -pyridine-2,6-diylbis[ N- (2- tert -butylphenyl)ethanimine] ( ² - ^{t -Bu} PDI; 0.15 equivalents; 0.284 g, 0.667 mmol) and tetrahydrofuran (11 mL). The thick green suspension is stirred overnight at room temperature and zinc (2.4 equivalents; 0.70 g, 11 mmol) and zinc bromide (1.1 equivalents; 1.1 g, 4.9 mmol) are added. After stirring for 15 minutes, the reaction mixture turns purple and a solution of 1- tert -butyl 2-methyl (2S)-2,5-dihydro- 1H -pyrrole-1,2- dicaroxylate (1.0 equivalent; 1.0 g, 4.4 mmol) in tetrahydrofuran (7.5 ml) and 2,2-dichloropropane (2.0 equivalents; 1.0 g, 8.8 mmol). The reaction mixture was stirred at room temperature for 5 days, then it was filtered through a pad of diatomaceous earth and washed with tetrahydrofuran (10.8 ml). The filtrate was combined with saturated aqueous ammonium chloride (3.5 ml) and ethyl acetate (9.5 ml); then the layers are separated and the aqueous phase is extracted with ethyl acetate (8.4 ml). The combined organic extracts were washed with saturated aqueous sodium bicarbonate (10.5 ml), dried over magnesium sulfate, filtered, and concentrated to dryness to give 3- tert -butyl 2-methyl( 1R , 2S , 5S )-6.6 -dimethyl-3-azabicyclo[3.1.0]hexane-2,3-dicaroxylate ( C90 ) as a yellow oil. Yield: 0.90 g, 3.3 mmol, 75%. According to ¹ H NMR analysis, this product exists as two carbamate rotamers (~3:2 ratio). ¹ H NMR (400 MHz, chloroform- d ) δ 4.20 & 4.09 (2 s, 1H), 3.74 & 3.75 (2 s, 3H), 3.68-3.60 (m, 1H), 3.44 & 3.38 (2d, J=10.9Hz, 1H), 1.43 & 1.38 (2s, 9H), 1.38-1.34 (m, 2H) , 1.03 & 0.98 & 0.96 (3 s, 6H). ESI-MS (pol.) m/z (%)=255.1 (12.5) [M - Me+H] ⁺ , 214.1 (100) [M - t -Bu+H] ⁺ , 170, 2 (50) [M - Boc+H] ⁺ .

Alternative production of C42

(1 R ,2 S ,5 S )-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxylic acids ( C42 )

Step 1. Synthesis of 3-methyl- N- (trifluoroacetyl)-L-valine ( C91 ).

A solution of sodium methoxide in methanol (25% by weight; 28.5 ml, 124 mmol) is added to a solution of 3-methyl-L-valine (99%, 15 g, 113 mmol) in methanol (30 ml). Ethyl trifluoroacetate (130 mmol) is then added and the reaction mixture is stirred at 40°C until the reaction is complete (approximately 2.5 hours), then it is cooled to 20°C. After adding hydrochloric acid (1 M; 136 ml, 136 mmol), the mixture is diluted with ethyl acetate (150 ml) and the layers are separated. The organic layer was washed twice with saturated aqueous sodium chloride, dried over magnesium sulfate, and filtered. Heptane is added to the filtrate, then the solution is concentrated at 50° C. to a volume of 5 ml/g. This procedure is repeated twice; after a second distillation, seed crystals C91 (50 mg; see below) are added. The resulting solid is collected by filtration, washed with heptane and dried at 40° C. to give C91 as an off-white solid. Yield: 22.2 g, 97.7 mmol, 86%.

The seed crystals used above are obtained from a similar reaction carried out using 3-methyl-L-valine; then the organic layer containing C91 is dried over magnesium sulfate and filtered, concentration in vacuo gives a solid. A portion of this solid is used as a seed product.

Physico-chemical data obtained on samples of C91 obtained from reactions carried out according to the same method. HRMS-ESI ⁺ ( m/z ): [M+H] ⁺ Calculated for C ₈ H ₁₃ F ₃ NO ₃ , 228.0842; found, 228.0842. The primary ion is observed as C ₈ H ₁₁ F ₃ NNa ₂ O ₃ [M+Na ⁺ ]: Calc., 272.0481; Found, 272.0482. ¹ H NMR (600 MHz, DMSO- d ₆ ) δ 13.05 (s, 1H), 9.48 (d, J =8.9 Hz, 1H), 4.21 (d, J =8.9 Hz , 1H), 1.00 (s, 9H). ¹³ C NMR (150.8 MHz, DMSO- d ₆ ) δ 170.9, 156.6 (q, ² J _CF = 36.9 Hz), 115.8 (q, ¹ J _CF = 287.7 Hz) , 61.0, 33.6, 26.5. Powder x-ray diffraction pattern for C91 is given in Figure 11; characteristic peaks are listed in Table R.

Table R. Selected X-ray Powder Diffraction Peaks for C91 Corner
(°2-theta) Rel. intensity Corner
(°2-theta) Rel. intensity Corner
(°2-theta) Rel. intensity 9.7 27 28.1 4 37.7 one 11.8 6 28.7 2 38.2 2 12.6 3 28.9 five 38.7 2 13.2 twenty 29.7 4 39.3 3 14.7 28 29.9 4 39.6 one 15.6 69 30.3 10 40.0 one 17.2 4 30.8 3 40.5 five 17.9 2 31.0 one 40.7 3 18.3 2 31.3 one 40.9 3 19.5 100 32.6 3 41.5 one 20.0 fourteen 33.5 2 41.8 one 20.4 2 34.0 one 42.4 3 21.5 22 34.2 2 43.1 one 23.2 eight 34.7 3 44.2 one 23.3 eleven 36.2 2 44.8 one 23.8 6 36.4 3 45.7 one 25.2 one 36.7 2 45.9 3 25.4 17 37.1 one 46.4 one 25.7 2 37.2 2 47.0 one 26.6 7 37.4 one 47.3 2 26.7 6 37.6 3 49.5 2

The X-ray crystallography crystal was obtained by recrystallization from ethyl acetate and hexane using seed crystals from the same batch as above. The ORTEP chart for C91 single crystal data is shown in Figure 12.

Single crystal X-ray structural analysis of C91

Single crystal x-ray analysis

Data collection is carried out on a Bruker D8 Quest diffractometer at -100°C. Data collection consists of omega and phi scans.

The structure was resolved by internal phasing using the SHELX software package in the space group of the orthorhombic class P 2 ₁ 2 ₁ 2 ₁ . In the future, the structure is refined by the full-matrix least squares method. All non-hydrogen atoms are found and refined using anisotropic displacement parameters.

Hydrogen atoms located on nitrogen were found from the difference Fourier map and refined with limited distances. Hydrogen into O ₂ (H ₂ Z) and O ₃ (H ₃ Z) is separated as a charge and refined as an employment of 10.5 each. The remaining hydrogen atoms are placed in calculated positions and allowed to move on their carrier atoms. The final refinement includes isotropic displacement parameters for all hydrogen atoms.

Population disruption is defined and modeled as a ratio of ~67/33 in the -CF ₃ segment.

Absolute structure analysis using probability methods (Hooft, 2008) is performed using PLATON (Spek). The results show that the absolute structure is assigned correctly. The method calculates that the probability that the structure is assigned correctly is 100%. The Hooft parameter is presented as -0.02 s os (estimated standard deviation) (4), and the Parson parameter is presented as -0.02 s s o (4).

The resulting R-index is 4.1%. The final difference Fourier map showed no missing or shifted electron density.

The corresponding crystal, data acquisition, and refinement information are summarized in Table S. Atomic coordinates, bond lengths, bond angles, and bias parameters are listed in Tables T-V.

A list of software and references used can be found in Single Crystal X-ray Structure Determination Example 13, Solid Form 1.

Table S. Crystal data and structure refinement for C91 . Empirical Formula
formula mass
Temperature
Wavelength
Crystal system
space group
Lattice constant
Volume
Z
Density (calculated)
Absorption coefficient
F (000)
Crystal size
Theta interval for data collection
Index ranges
Collected Reflections
Independent Reflections
Completeness to theta=67.679°
Absorption correction
Refinement method
Data/Restrictions/Parameters
Compliance with F ²
Finite R indices [ I>2σ(I) ]
R indices (all data)
Absolute Structure Parameter
Excitation factor
The largest diff. peaks and valleys C ₈ H ₁₂ F ₃ NO ₃
227.19
173(2) K
1.54178 Å
tetragonal
P 4 ₁ 2 ₁ 2
a =9.9168(6) Å
b =9.9168(6) Å
c = 22.721(2) Å
2234.5(4) Å ³
eight
1.351 Mg/ ^m3
1.184 mm ^-1
944
0.200×0.170×0.080 mm ³
4.866-70.114°
-11<= h <=10, -12<= k <=12, -27<= l <=27
48160
2122 [ _Rint = 0.0392 ]
99.8%
empirical
Least squares in the full-matrix approximation in F ²
2122/9/158
1.010
R 1=0.0408, wR 2=0.1012
R 1=0.0429, wR 2=0.1039
0.03(4)
n/a
0.280 and -0.215 e.Å ^-3 α= 90°
β= 90°
γ =90°

Table T. Atomic coordinates (x 10 ⁴ ) and equivalent isotropic substitution parameters (Å ² ×10 ³ ) for C91 . U(eq) are defined as one third of the trace of the orthogonalized tensor U ^ij .

Table U. Bond lengths [Å] and angles [°] for C91 .

Symmetric transformations are used to create equivalent atoms.

Table V. Anisotropic substitution parameters (Å ² ×10 ³ ) for C91 . The exponent of the anisotropic substitution factor takes the form: -2π ² [h ² a* ² U ¹¹ + ... + 2 hka* b* U ¹² ].

Step 2. Synthesis of lithium ( 1R ,2S, 5S )-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2- caroxylate ( C92 ).

Lithium hydroxide monohydrate (29.0 g, 678 mmol) is added to a mixture of methyl (1R, 2S , 5S )-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2- caroxylate , hydrochloride (68.5 g, 333 mmol) in tetrahydrofuran (950 ml) and water (48 ml). The reaction mixture was stirred at 25° C. until hydrolysis was complete, then the solid was collected by filtration, washed with 5% tetrahydrofuran water (400 ml) and dried under vacuum at 70° C. to give C92 as a white/off-white solid. Yield: 47.6 g, 295 mmol, 89%. The physicochemical data obtained for the C92 samples are obtained from reactions carried out in the same manner.

ISWR-IER⁺ (m/z): [M+H]⁺ Calculated for C_eightH_fourteenNO₂.156.1019; Found, 156.1019.^oneH NMR (600 MHz, D₂O) δ 3.23 (d,J=1.1 Hz, 1H), 3.09 (dd,J=11.1, 5.2 Hz, 1H), 2.63 (d,J=11.1 Hz, 1H), 1.33-1.24 (m, 2H), 0.86 (s, 2H), 0.83 (s, 3H).¹³C NMR (150.8 MHz, D₂O) δ 182.7, 62.3, 45.6, 35.5, 30.0, 25.8, 19.3, 12.7. Powder X-ray diffraction pattern forC92 given in Figure 13; characteristic peaks are listed in Table W.

Table W. Selected X-ray Powder Diffraction Peaks for C92 Angle (°2-theta) Rel. intensity Corner
(°2-theta) Rel. intensity Corner
(°2-theta) Rel. intensity 5.1 4 20.2 6 30.2 4 6.0 100 20.8 nineteen 30.5 2 7.3 17 21.4 2 31.4 eighteen 8.7 3 21.9 4 32.2 five 10.2 6 22.3 16 32.6 22 12.1 twenty 22.8 7 33.8 3 12.7 2 23.8 3 34.3 3 13.7 2 24.5 nine 35.2 3 15.3 fifteen 24.8 3 35.9 10 15.7 6 25.5 21 36.7 4 16.7 78 26.2 five 37.4 4 17.2 eight 26.5 4 37.8 4 18.0 3 28.4 10 39.0 2 18.8 95 29.0 10 39.2 one 19.5 fourteen 29.5 4 39.8 one

Step 3. Synthesis of (1 R ,2 S ,5 S )-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane -2-carboxylic acid ( C42 ).

Mixture of C91 (1.29 g, 5.68 mmol), 4-(dimethylamino)pyridine (0.60 g, 4.8 mmol) and N,N -diisopropylethylamine (1.70 ml, 9.75 mmol) in tetrahydrofuran (10 ml), treated with p -toluenesulfonyl chloride (0.99 g, 5.2 mmol). The reaction mixture is then stirred for 2 hours at 20°C, C92 (75.7% wt., 1.00 g, 4.70 mmol) is charged and stirring is continued overnight at 20°C. The resulting suspension is mixed with propan-2-ylacetate (10 ml) and washed successively with an aqueous citric acid solution (10%, 10 ml) and water (10 ml). The organic layer is then concentrated, then propan-2-ylacetate (5 ml) is added, then heptane (15 ml) is added dropwise from an addition funnel. The solids are isolated by filtration and dried in vacuo to give ( 1R , 2S , 5S )-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3 ,1,0]hexane-2-carboxylic acid ( C42 ) as a white solid. Yield: 1.2 g. This compound shows two sets of NMR signals. The major and minor sets correspond to the Z and E isomers of the tertiary amide, respectively, in a molar ratio of 20:1. The sample also contains isopropyl acetate at a 37% mole ratio relative to C42 showing ¹ H resonances at 4.86, 1.96, and 1.17 ppm, and ¹³ C resonances at 169.7, 66.9, 21, 5, and 21.0 ppm ¹ H and ¹³ C signals are denoted using the TMS signal set to 0 ppm. in both.

ISWR-IER⁺ (m/z): [M+H]⁺ Calculated for C₁₆H₂₄F₃N₂O₄.365.1683; Found: 365,1684.^oneH NMR (600 MHz, DMSO-d ₆) δbasic: 9.44 (d,J= 8.5 Hz, 1H), 4.44 (d,J= 8.5 Hz, 1H), 4.15 (s, 1H), 3.85 (dd,J=10.5, 5.4 Hz, 1H), 3.73 (d,J=10.5 Hz, 1H), 1.53 (dd,J=7.6, 5.3 Hz, 1H), 1.43 (d,J=7.6 Hz, 1H), 1.01 (s, 3H), 1.01 (s, 9H), 0.83 (s, 3H);minor: 9.11 (d,J=9.4 Hz, 1H), 4.53 (s, 1H), 4.33 (d,J=9.4 Hz, 1H), 3.53 (dd,J=12.5, 5.3 Hz, 1H), 3.41 (d,J=12.5 Hz, 1H), 1.55 (d,J\u003d 7.5 Hz, 1H), 1.41 (dd,J=7.5, 5.3 Hz, 1H), 1.02 (s, 3H), 0.97 (s, 3H), 0.91 (s, 9H).¹³C NMR (150.8 MHz, DMSO-d ₆) δbasic: 172.3, 167.5, 156.8 (² J _CF=37.0 Hz), 115.7 (^one J _CF=287.7 Hz), 59.1, 58.0, 47.1, 34.6, 29.6, 26.7, 26.1, 25.6, 18.7, 12.0;minor: 172.3, 168.1, 155.9 (² J _CF=36.8 Hz), 115.8 (^one J _CF=288.1 Hz), 59.9, 57.3, 46.4, 36.2, 32.1, 26.2, 26.0, 24.4, 19.0, 12.7. Powder X-ray diffraction pattern forC42 given in figure 14; characteristic peaks are listed in Table X.

Crystallization for powder x-ray diffraction and single-crystal x-ray structural analysis is carried out as follows. A solution of C42 (2.96 g) in ethanol (9 ml) was heated to 40° C. with stirring (3500 rpm), then water (10.5 ml) was added over 10 minutes. Then add additional water (16.5 ml) over 4 hours, and the mixture is cooled to 10°C and stirred overnight. After filtration, the filter cake was washed with water (6 ml) and dried at 50° C. to give crystalline C42 (2.6 g).

Table X. Selected X-ray Powder Diffraction Peaks for C42 Corner
(°2-theta) Rel. intensity Corner
(°2-theta) Rel. intensity Corner
(°2-theta) Rel. intensity 7.4 100 24.2 3 37.4 one 9.4 73 24.4 2 38.3 one 12.0 fifteen 25.3 4 39.0 one 12.9 16 25.5 4 39.7 one 14.1 nine 27.6 6 40.1 one 14.9 26 28.6 one 41.2 one 17.4 7 29.5 4 41.9 one 17.7 35 31.4 2 42.6 one 19.0 eighteen 31.5 3 46.3 one 19.2 12 32.4 one 19.7 17 33.2 one 20.4 nine 34.1 one 20.6 4 34.5 2 22.4 3 35.7 one 23.0 3 36.1 0 23.2 2 36.6 one

The ORTEP chart for C42 single crystal data is shown in Figure 15.

Single-crystal X-ray structural analysis of C42

Single crystal x-ray analysis

Data collection is carried out on a Bruker D8 Venture diffractometer at room temperature. A special data strategy of 0.3 degrees width per frame is used to separate the domains, thereby eliminating any TWIN and pseudo-symmetry issues.

The structure is resolved by internal phasing using the SHELX software package in the R 3 rhombohedral class group. The structure is further refined by the full matrix least squares method. All non-hydrogen atoms are found and refined using anisotropic displacement parameters.

Hydrogen atoms located on nitrogen and oxygen were found from the difference Fourier map and refined with limited distances. The remaining hydrogen atoms are placed in calculated positions and allowed to move on their carrier atoms. The final refinement includes isotropic displacement parameters for all hydrogen atoms.

Absolute structure analysis using probability methods (Hooft, 2008) is performed using PLATON (Spek). The results show that the absolute structure is assigned correctly. The method calculates that the probability that the structure is assigned correctly is 100%. Hooft's parameter is presented as -0.08 s os (estimated standard deviation) (7), and Parson's parameter is presented as -0.09 s os (6).

Population occupancy disruption is determined and modeled as a ratio of 78:22 in the C1_F1_F2 segment.

The final R-index is 5.8%. The final difference Fourier map showed no missing or shifted electron density.

The corresponding crystal, data acquisition, and refinement information are summarized in Table Y. Atomic coordinates, bond lengths, bond angles, and bias parameters are listed in Tables Z-BB.

A list of software and references used can be found in Single Crystal X-ray Structure Determination Example 13, Solid Form 1.

Table Y. Crystal data and structure refinement for C42 . Empirical Formula
formula mass
Temperature
Wavelength
Crystal system
space group
Lattice constant
Volume
Z
Density (calculated)
Absorption coefficient
F (000)
Crystal size
Theta interval for data collection
Index ranges
Collected Reflections
Independent Reflections
Completeness to theta=67.679°
Absorption correction
Refinement method
Data/Restrictions/Parameters
Compliance with F ²
Finite R indices [ I>2σ(I) ]
R indices (all data)
Absolute Structure Parameter
Excitation factor
The largest diff. peaks and valleys C ₁₆ H ₂₃ F ₃ N ₂ O ₄
364.36
296(2) K
1.54178 Å
Trigonal
R3
a =14.1740(6) Å
b =14.1740(6) Å
c = 14.1740(2) Å
1715.9(4) Å ³
3
1.058 Mg/ ^m3
0.788 mm ^-1
576
0.200×0.100×0.100 mm ³
6.445-80.034°
-17<= h <=16, -14<= k <=16,
-14<= l <=17
13310
4011 [ _Rint = 0.0369 ]
98.9%
empirical
Least squares in the full-matrix approximation in F ²
4011/6/244
1.056
R 1=0.0582, wR 2=0.1675
R 1=0.0611, wR 2=0. 0.1710
-0.09(6)
n/a
0.292 and -0.174 e.Å ^-3 α= 114.11°
β= 114.11°
γ =114.11°

Table Z. Atomic coordinates (x 10 ⁴ ) and equivalent isotropic substitution parameters (Å ² × 10 ³ ) for C42 . U(eq) are defined as one third of the trace of the orthogonalized tensor U ^ij .

Table A.A. Bond lengths [Å] and angles [°] for C42 .

Symmetric transformations are used to create equivalent atoms.

BB table. Anisotropic substitution parameters (Å ² ×10 ³ ) for C42 . The exponent of the anisotropic substitution factor takes the form: -2π ² [h ² a* ² U ¹¹ + ... + 2 hka* b* U ¹² ].

In addition to obtaining (1 R ,2 S ,5 S )-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane -2-carboxylic acid ( C42 ) according to the methods described above, the compound can also be obtained as depicted in the reaction schemes shown directly below. In step 1, methyl (1R,2S,5S)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate hydrochloride is first treated with triethylamine in a mixture of tetrahydrofuran and water to neutralize the hydrochloride, then methyl ether using sodium hydroxide in a mixture of tetrahydrofuran and water to obtain sodium (1R,2S,5S)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate sodium.

Preparation of sodium (1R,2S,5S)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate

To a suitable vessel add tetrahydrofuran (30 ml), water (7.5 ml), triethylamine (7.62 ml, 54.7 mmol) and methyl (1R,2S,5S)-6,6-dimethyl-3-azabicyclo hydrochloride [3.1.0]hexane-2-caroxylate (7.59 g, 36.9 mmol). The mixture is stirred at 25°C for at least 30 minutes. Stirring is stopped and the layers are separated. In a separate vessel 28% wt./mass. aqueous sodium hydroxide (4.19 ml, 38.3 mmol) and tetrahydrofuran (71 ml) are added with stirring at 40°C. 25% organic layer containing a solution of methyl (1R,2S,5S)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate in tetrahydrofuran was added after separation and the solution was seeded (1R, 2S,5S)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-sodium carboxylate (0.1182 g, 0.7336 mmol - previously obtained by a similar method). The mixture is kept at 40° C. for at least 15 minutes and the remaining 75% of the organic layer is slowly added. The mixture is kept under stirring at 40°C for 16 hours, then slowly cooled to 20°C and held for at least 4 hours. sodium azabicyclo[3.1.0]hexane-2-caroxylate is isolated by filtration, washed with a solution consisting of tetrahydrofuran (43 ml) and water (2.25 ml). The solid is dried at 70° C. under vacuum to give 6.13 g (93.8%) of (1R,2S,5S)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate sodium as a crystalline solid. PDP is determined by the methods described above.

Selected XRD peaks of crystalline (1R,2S,5S)-6,6-dimethyl-3-azabicyclo[3,1,0]hexane-2-caroxylate sodium.

Corner
degrees
2-θ
± 0.2 °2-θ Relative intensity Corner
degrees
2-θ
± 0.2 °2-θ Relative intensity Corner
degrees
2-θ
± 0.2 °2-θ Relative intensity 5.5 nineteen 19.0 fifteen 29.8 4 5.9 nine 19.3 100 30.3 eight 6.6 59 21.3 45 30.8 eight 10.7 2 23.2 12 32.0 3 16.0 nine 23.5 17 32.5 five 16.3 6 24.2 twenty 33.8 7 16.7 10 25.8 eleven 34.6 4 17.0 52 26.0 eight 35.8 6 17.3 42 26.7 4 36.6 eight 17.7 fifteen 28.1 3 36.9 7 18.5 7 28.9 6 37.6 3

Selected XRD peaks of (S)-3,3-dimethyl-2-(2,2,2-trifluoroacetamido)butanoic acid ( C91 )

Angle degrees
2-θ ±0.2 °2-θ Relative intensity Angle degrees
2-θ
±0.2 °2-θ Relative intensity Angle degrees
2-θ
±0.2 °2-θ Relative intensity 9.7 27 28.1 4 37.7 one 11.8 6 28.7 2 38.2 2 12.6 3 28.9 five 38.7 2 13.2 twenty 29.7 4 39.3 3 14.7 28 29.9 4 39.6 one 15.6 69 30.3 10 40.0 one 17.2 4 30.8 3 40.5 five 17.9 2 31.0 one 40.7 3 18.3 2 31.3 one 40.9 3 19.5 100 32.6 3 41.5 one 20.0 fourteen 33.5 2 41.8 one 20.4 2 34.0 one 42.4 3 21.5 22 34.2 2 43.1 one 23.2 eight 34.7 3 44.2 one 23.3 eleven 36.2 2 44.8 one 23.8 6 36.4 3 45.7 one 25.2 one 36.7 2 45.9 3 25.4 17 37.1 one 46.4 one 25.7 2 37.2 2 47.0 one 26.6 7 37.4 one 47.3 2 26.7 6 37.6 3 49.5 2

Refinement of crystal data and refinement of the structure for (S)-3,3-dimethyl-2-(2,2,2-trifluoroacetamido)butanoic acid ( C91 ).

Identification code E178 Empirical formula
formula mass
Temperature
Wavelength
Crystal system
space group
Lattice constant
Volume
Z
Density (calculated)
Absorption coefficient
F (000)
Crystal size
Theta interval for data collection
Index ranges
Collected Reflections
Independent Reflections
Completeness to theta=67.679°
Absorption correction
Refinement method
Data/Restrictions/Parameters
Compliance with F ²
Finite R indices [ I>2σ(I) ]
R indices (all data)
Absolute Structure Parameter
Excitation factor
The largest diff. peaks and valleys C ₈ H ₁₂ F ₃ NO ₃
227.19
173(2) K
1.54178 Å
tetragonal
P 4 ₁ 2 ₁ 2
a =9.9168(6) Å
b =9.9168(6) Å
c = 22.721(2) Å
2234.5(4) Å ³
eight
1.351 Mg/ ^m3
1.184 mm ^-1
944
0.200×0.170×0.080 mm ³
4.866-70.114°
-11<= h <=10, -12<= k <=12, -27<= l <=27
48160
2122 [ _Rint = 0.0392 ]
99.8%
empirical
Least squares in the full-matrix approximation in F ²
2122/9/158
1.010
R 1=0.0408, wR 2=0.1012
R 1=0.0429, wR 2=0.1039
0.03(4)
n/a
0.280 and -0.215 e.Å ^-3 α= 90°
β= 90°
γ =90°

In step 2, the resulting sodium (1R,2S,5S)-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-caroxylate is then combined with (S)-3,3-dimethyl-2- (2,2,2-trifluoroacetamido)butanoic acid with the presence of tosyl chloride and dimethylaminopyridine in tetrahydrofuran. The tetrahydrofuran is removed and replaced with isopropyl acetate, then treated with HCl in brine, then treated with water and heptane to give C42.

Crystalline C42 characterizes the PDP and identifies an additional form obtained after prolonged drying or higher temperature.

Selected XRD peaks for C42 - crystalline form obtained after prolonged drying or higher temperature

Corner
degrees
2-θ
± 0.2 °2-θ Relative intensity Corner
degrees
2-θ
± 0.2 °2-θ Relative intensity Corner
degrees
2-θ
± 0.2 °2-θ Relative intensity 10.2 3 25.7 eleven 33.7 one 10.8 100 26.1 3 34.1 one 12.6 22 26.8 2 34.6 4 13.3 21 27.1 2 35.1 3 13.8 88 27.4 7 35.5 one 16.3 59 27.8 7 36.3 4 17.1 39 28.0 3 36.9 6 18.7 five 28.2 3 38.2 6 18.9 2 29.0 2 38.8 2 19.5 twenty 29.5 4 39.3 one 19.9 eighteen 29.8 2 39.6 2 20.3 2 30.1 2 39.8 2 20.6 26 30.6 7 40.2 2 20.8 63 30.9 3 42.2 3 21.7 6 31.2 3 44.2 2 22.2 twenty 32.0 eight 44.7 3 23.4 eight 32.7 4 47.0 2 23.7 eighteen 32.9 4 47.3 2 24.5 7 33.1 five 25.4 nine 33.5 one

Antiviral activity against SARS-CoV-2 infection

The ability of compounds to prevent SARS-CoV-2 coronavirus-induced cell death or cytopathic effect can be assessed by cell viability using an assay format that uses luciferase to measure intracellular ATP as an endpoint. Briefly, VeroE6 cells that are enriched in hACE2 expression are inoculated with SARS-CoV-2 (USA_WA1/2020) at an MOI of 0.002 in a BSL-3 lab. Cells inoculated with virus are then added to assay-ready compound plates at a density of 4000 cells/well. After 3 days of incubation, when the virus-induced cytopathic effect is 95% under untreated infected control conditions, cell viability is assessed using Cell Titer-Glo (Promega) according to the manufacturer's protocol, which quantifies ATP levels. The cytotoxicity of the compound is evaluated in parallel to non-infected cells. Test compounds are tested either alone or in the presence of P-glycoprotein (P-gp) inhibitor CP-100356 at a concentration of 2 µM. The inclusion of CP 100356 is intended to assess whether test compounds are cleared from VeroE6 cells that have high levels of P-glycoprotein expression. The proportion of effect for each test compound is calculated from the values for wells without virus and control wells containing virus on each assay plate. The concentration required for a 50% response (EC ₅₀ ) is determined from these data using a 4-parameter logistic model. Curves EC ₅₀ correspond to slope 3 Hill at >3 and the maximum dose, giving ≥50% effect. If cytotoxicity is detected at an effect greater than 30%, the corresponding data is excluded from the definition of the EU ₅₀ .

For plates with cytotoxicity, calculate the proportion of effect for each type of test compound based on the values for the cell-only control wells and the hyamine-containing control wells on each assay plate. The CC ₅₀ value is calculated using a 4-parameter logistic model. The TI is then calculated by dividing the CC ₅₀ value by the EC ₅₀ value.

FRET study and analysis of SARS-CoV-2 coronavirus 3C protease

The proteolytic activity of the major protease, 3CL ^pro , of SARS-CoV-2 is monitored by a continuous fluorescence resonance energy transfer (FRET) assay. The SARS-CoV-2 3CL ^pro assay measures the activity of the full-length SARS-CoV-2 3CL protease by cleaving a synthetic fluorogenic substrate peptide with the following sequence: Dabcyl-KTSAVLQ-SGFRKME-Edans, modeled on a consensus peptide (V. Grum-Tokars et al. Evaluating the 3C-like protease activity of SARS-coronavirus: recommendations for standardized assays for drug discovery Virus Research 133 (2008) 63-73). The fluorescence of the digested Edans peptide (excitation 340 nm/emission 490 nm) is measured using the fluorescence intensity protocol on a Flexstation reader (Molecular Devices). The fluorescent signal is reduced in the presence of PF-835231, a potent SARS-CoV-2 3CL ^pro inhibitor. The assay reaction buffer contains 20 mM Tris-HCl (pH 7.3), 100 nM NaCl, 1 mM EDTA, and 25 μM peptide substrate. Enzymatic reactions initiated by adding 15 nm 3CL protease SARS-CoV-2 and carried out for 60 minutes at 23°C. The proportion of inhibition or activity is calculated based on control wells without compound (0% inhibition/100% activity) and control compound (100% inhibition/0% activity). IC ₅₀ values are obtained using a four parameter fitting model using ABASE software (IDBS). The Ki values correspond to the Morrison equation with the enzyme concentration parameter set at 15 nM, the Km parameter set at 14 μm, and the substrate concentration parameter set at 25 μm using ABASE software (IDBS).

The proteolytic activity of the SARS-CoV-2 coronavirus 3CL protease was measured by continuous resonance fluorescence energy transfer assay. The SARS-CoV-2 3CL ^pro FRET assay measures protease-catalyzed cleavage of TAMRA-SITSAVLQSGFRKMK-(DABCYL)-OH to TAMRA-SITSAVLQ and SGFRKMK(DABCYL)-OH. The fluorescence of the digested TAMRA peptide (ex. 558 nm/exp. 581 nm) was measured using a TECAN SAFIRE Fluorescence Plate Reader for 10 minutes. Typical reaction solutions contain 20 mM HEPES (pH 7.0), 1 mM EDTK, 4.0 μm FRET substrate, 4% DMSO, and 0.005% Tween-20. The assays are initiated by adding 25 nM SARS 3CL ^pro (nucleotide sequence 9985-10902 of the full genome sequence of the SARS coronavirus strain Urbani (NCBI accession number AY278741)). The proportion of inhibition is determined twice at an inhibitor level of 0.001 mm. The data is analyzed with the Kalidagraph non-linear regression analysis program using the equation:

FU=divergence + (limit) (1- e ^-(kobs) t)

where the divergence is equal to the fluorescence signal of the uncleaved peptide substrate, and the limit is equal to the fluorescence of the fully cleaved peptide substrate. Kobs is the first order rate constant for this reaction, and in the absence of any inhibitor represents substrate utilization. The enzyme starts a reaction that contains an irreversible inhibitor, and if the calculated limit is less than 20% of the theoretical maximum limit, the calculated kobs is the rate of inactivation of the 3C protease of the coronavirus. Slope (kobs/I) plot kobs vs. [I] is a measure of the avidity of an inhibitor for an enzyme. For very fast irreversible inhibitors, kobs/I is calculated from observations for only one or two [I], and not as a slope.

Table 2. Biological activity and IUPAC name for examples 1-84. Example number Geometric mean K _i (µm) Number of K _i used (µm) Geometric mean EC ₅₀ (µm) Number of EC ₅₀ used (µm) Name IUPAC one 0.013 4 0.246 7 (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [ N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 2 1.08 3 7.52 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -(pyrrolidin-1-ylacetyl)-L-leucinamide, trifluoroacetate 3 0.439 2 6.74 6 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -(2,6-dichlorobenzoyl)-4-methyl-L-leucinamide 4 0.026 3 1.36 fifteen N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-1 H -indole-2-carboxamide five >0.351 2 >3.33 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-3-(trifluoromethyl)-1 H -indole-2-carboxamide 6 0.023 2 0.279 4 N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-7-(trifluoromethyl)-1 H -indole-2-carboxamide 7 0.798 one 43.1 2 N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-3-methylimidazo[2,1- b ][1,3]thiazole-2-carboxamide eight 0.917 3 5.75 2 N -{1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide, DIAST-1 nine 0.254 4 0.970 4 N -{1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[cyclohexyl(methoxy)acetyl]-4-methyl-L-leucinamide, DIAST-2 10 0.056 4 2.09 4 N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-1 H -indole-2-carboxamide eleven 0.297 3 4.78 4 N ² -[(4-bromo-1-ethyl-3-methyl-1 H -pyrazol-5-yl)carbonyl]- N -{(1 S )-1-cyano-2-[(3 S )-2 -oxopyrrolidin-3-yl]ethyl}-4-methyl-L-leucinamide 12 0.539 3 6.33 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[(3,3-difluorocyclobutyl)acetyl]-4-methyl-L -leucinamide 13 0.003 6 0.075 eighteen (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide fourteen 0.302 2 N.D.1 N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-5,5,5-trifluoro- 1-oxopentan-2-yl]-4-methoxy-1 H -indole-2-carboxamide fifteen 0.002 one 0.360 2 N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-7-fluoro-4-methoxy-1 H -indol-2-carboxamide 16 0.018 2 >2.53 2 N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-5-fluoro-4-methoxy-1 H -indol-2-carboxamide 17 0.053 one >0.333 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-3-fluoro-4-methoxy-1 H -indol-2-carboxamide eighteen 0.019 2 >0.333 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-5,7-difluoro-4-methoxy-1 H -indole-2-carboxamide nineteen 0.208 2 >0.333 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-3,5-difluoro-4-methoxy-1 H -indole-2-carboxamide twenty 0.005 one >0.333 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-7-fluoro-4-methoxy-1 H -indole-2-carboxamide 21 BUT. BUT. >3.17 2 N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-5-fluoro-4-methoxy-1 H -indole-2-carboxamide 22 0.066 one >0.333 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-3-fluoro-4-methoxy-1 H -indole-2-carboxamide 23 0.021 one >0.333 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-5,7-difluoro-4-methoxy-1 H -indole-2-carboxamide 24 1.93 2 6.30 6 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -{[2-(trifluoromethyl)-1,3- thiazol-4-yl]carbonyl}-L-leucinamide 25 BUT. 37 2 N -{1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -(cyclohexylcarbonyl)-4-methyl-L-leucinamide, DIAST-1 26 BUT. >100 one N -{1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -(cyclohexylcarbonyl)-4-methyl-L-leucinamide, DIAST-2 27 4.50 2 44.9 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -[(propan-2-yloxy)acetyl]-L -leucinamide 28 1.79 2 7.26 4 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[(cyclohexyloxy)acetyl]-4-methyl-L-leucinamide 29 2.23 2 28.1 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -(4,4,4-trifluoro-3-methylbutanoyl )-L-leucinamide thirty >10.8 one 9.37 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[(2 S )-2-(dimethylamino)-2-phenylacetyl] -4-methyl-L-leuinamide 31 0.606 3 40.7 2 N ² -[( trans -4-cyanocyclohexyl)carbonyl]- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L -leucinamide
or
N ² -[( trans -4-cyanocyclohexyl)carbonyl] -N -{(1 R )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl-L -leucinamide 32 0.690 3 4.90 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[2-(cyclohexyloxy)propanoyl]-4-methyl-L-leucinamide
or
N -{(1 R )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[2-(cyclohexyloxy)propanoyl]-4-methyl-L-leucinamide 33 0.068 2 >0.333 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-hydroxy-1 H -indole-2-carboxamide 34 0.074 2 >0.333 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-5-hydroxy-4-methoxy-1 H -indol-2-carboxamide 35 0.176 3 3.56 4 N ² -[(4-chloro-1,3-dimethyl-1 H -pyrazol-5-yl)carbonyl]- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine -3-yl]ethyl}-4-methyl-L-leucinamide 36 0.241 4 1.03 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -[(2 R )-2-(dimethylamino)-2-phenylacetyl] -4-methyl-L-leucinamide 37 0.168 one >3.33 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-3-(trifluoromethyl)-1 H -indole-2-carboxamide 38 0.023 one >0.333 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-7-(trifluoromethyl)-1 H -indole-2-carboxamide 39 0.111 one >3.33 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-3,7-bis(trifluoromethyl)-1 H -indole-2-carboxamide 40 0.131 one >0.333 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-3,5-bis(trifluoromethyl)-1 H -indole-2-carboxamide 41 0.104 one >0.333 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-3,6-bis(trifluoromethyl)-1 H -indole-2-carboxamide ² 42 >0.356 one >0.333 one N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -{(2 R )-2-(dimethylamino)-2-[3 -(trifluoromethyl)phenyl]acetyl}-4-methyl-L-leucinamide 43 >0.356 one >0.333 one N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- N ² -{(2 R )-2-(dimethylamino)-2-[4 -(trifluoromethyl)phenyl]acetyl}-4-methyl-L-leucinamide 44 0.302 2 5.88 2 N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-6-(trifluoromethyl)-1 H -indol-2-carboxamide 45 0.227 2 6.42 2 N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-6-(trifluoromethyl)-1 H -indole-2-carboxamide 46 0.283 one >3.33 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-3,7-bis(trifluoromethyl)-1 H -indole-2-carboxamide 47 >0.359 one >3.33 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-3,6,7-tris(trifluoromethyl)-1 H -indole-2-carboxamide 48 >0.359 one >3.33 one N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-3,5,7-tris(trifluoromethyl)-1 H -indole-2-carboxamide 49 0.083 3 2.68 2 N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-(trifluoromethoxy)-1 H -indol-2-carboxamide fifty 0.147 3 3.95 one N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-(trifluoromethoxy)-1 H -indole-2-carboxamide 51 0.192 3 BUT. N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-5-(trifluoromethyl)-1 H -indole-2-carboxamide 52 0.032 3 BUT. 7-chloro- N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4- dimethyl-1-oxopentan-2-yl]-1 H -indole-2-carboxamide 53 0.025 3 BUT. N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-methoxy-7-methyl-1 H -indole-2-carboxamide 54 0.049 3 BUT. 6-chloro- N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4- dimethyl-1-oxopentan-2-yl]-1 H -indole-2-carboxamide 55 0.104 3 BUT. 4-chloro- N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4- dimethyl-1-oxopentan-2-yl]-1 H -indole-2-carboxamide 56 0.151 3 BUT. 5-chloro- N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4- dimethyl-1-oxopentan-2-yl]-1 H -indole-2-carboxamide 57 0.052 3 BUT. N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-7-(trifluoromethyl)-1 H -indole-2-carboxamide 58 0.091 3 BUT. 4,6-dichloro- N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4, 4-dimethyl-1-oxopentan-2-yl]-1 H -indole-2-carboxamide 59 0.152 3 BUT. N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-4-(trifluoromethyl)-1 H -indole-2-carboxamide 60 0.261 3 BUT. N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-5-(trifluoromethyl)-1 H -indol-2-carboxamide 61 0.034 3 BUT. 7-chloro- N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl- 1-oxopentan-2-yl]-1 H -indole-2-carboxamide 62 0.029 3 BUT. N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-methoxy-7-methyl-1 H -indol-2-carboxamide 63 0.122 3 BUT. 6-chloro-N-[(2S)-1-({(1S)-1-cyano-2-[(3S)-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentane -2-yl]-1H-indole-2-carboxamide 64 0.038 3 BUT. 4-chloro- N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl- 1-oxopentan-2-yl]-1 H -indole-2-carboxamide 65 0.117 3 BUT. 5-chloro- N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl- 1-oxopentan-2-yl]-1 H -indole-2-carboxamide 66 0.073 3 BUT. N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-7-(trifluoromethyl)-1 H -indol-2-carboxamide 67 0.041 3 BUT. 4,6-dichloro- N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4- methyl-1-oxopentan-2-yl]-1 H -indole-2-carboxamide 68 0.092 3 BUT. N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl-1-oxopentan- 2-yl]-4-(trifluoromethyl)-1 H -indol-2-carboxamide 69 0.083 one >2.67 2 N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1- oxopentan-2-yl]-3-methyl-5-(trifluoromethyl)imidazo[2,1- b ][1,3]thiazole-2-carboxamide 70 6.95 2 4.09 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -{[5-methyl-2-(trifluoromethyl)- 1,3-thiazol-4-yl]carbonyl}-L-leucinamide 71 0.254 3 4.99 2 N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-methyl- N ² -{[4-methyl-2-(trifluoromethyl)- 1,3-thiazol-5-yl]carbonyl}-L-leucinamide 72 0.173 3 1.35 2 N ² -[(4-bromo-1-ethyl-3-methyl-1 H -pyrazol-5-yl)carbonyl]- N -{(1 S )-1-cyano-2-[(3 S )-2 -oxopyrrolidin-3-yl]ethyl}-L-leucinamide 73 0.226 3 2.03 2 N ² -[(4-chloro-1,3-dimethyl-1 H -pyrazol-5-yl)carbonyl]- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine -3-yl]ethyl}-L-leucinamide 74 >0.356 one >3.33 one 3-acetyl- N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4-methyl- 1-oxopentan-2-yl]-4-methoxy-1 H -indole-2-carboxamide 75 0.017 2 0.551 4 Diastereomer 1: ( 2S , 4R )-4- tert -butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1- { N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide
or
( 2R , 4S )-4- tert -butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1-{ N- [(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide 76 >8.16 2 >100 one Diastereomer 2: ( 2S , 4R )-4- tert -butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1- { N -[(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide
or
( 2R , 4S )-4- tert -butyl- N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-1-{ N- [(trifluoromethyl)sulfonyl]-L-valyl}piperidine-2-carboxamide 77 0.004 2 0.085 4 3-methyl- N- (trifluoroacetyl)-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- 4-(trifluoromethyl)-L-prolinamide 78 0.005 3 2.74 2 (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (methylcarbamoyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 79 0.001 4 0.080 4 methyl {(2 S )-1-[(1 R ,2 S ,5 S )-2-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}carbamoyl)-6,6-dimethyl-3-azabicyclo[3.1.0]hexan-3-yl]-3,3-dimethyl-1-oxobutan-2-yl}carbamate 80 0.037 2 0.158 3 N- (trifluoroacetyl)-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-4-(trifluoromethyl )-L-prolinamide 81 0.003 2 0.690 3 (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 R )-5-hydroxy-2-oxopyrrolidin-3-yl]ethyl}-6,6- dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 82 0.139 one BUT. BUT. (1 R ,2 S ,5 S ,6 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6-(hydroxymethyl) -6-methyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 83 0.092 one BUT. BUT. (1 R ,2 S ,5 S ,6 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6-(hydroxymethyl) -6-methyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 84 0.003 one BUT. BUT. (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-(hydroxymethyl) - N- (trifluoroacetyl)-L-valyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide 85 0.004 one 0.334 2 (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 R )-2,5-dioxopyrrolidin-3-yl]ethyl}-6,6-dimethyl- 3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 86 0.018 2 0.237 2 (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [5,5,5-trifluoro-2-(2,2,2-trifluoroacetamido)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 87 0.021 2 0.230 2 (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[(2 S )- 2-cyclohexyl-2-{[(trifluoromethyl)sulfonyl]amino}acetyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide 88 0.006 2 0.050 4 First-eluting diastereomer: ( 1R , 2S , 5S ) -N -{( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}-3-[ 3-cyclobutyl- N- (trifluoroacetyl)-L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide
or
(1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclobutyl- N -(trifluoroacetyl)-D-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide 89 0.007 3 0.040 4 Second-eluting diastereomer: (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[ 3-cyclobutyl- N- (trifluoroacetyl)-L-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide
or
(1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[3-cyclobutyl- N -(trifluoroacetyl)-D-alanyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide 90 0.015 2 0.571 2 (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-(pyridin-2-yl) -N- (trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 91 0.007 2 0.011 2 (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-{ N -[(4 -fluorophenoxy)acetyl]-3-methyl-L-valyl}-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide 92 0.004 2 0.019 4 3-methyl- N -[(4-methylphenyl)acetyl]-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-4-(trifluoromethyl)-L-prolinamide 93 0.014 2 0.882 2 (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-(1 H -pyrazol-1-yl)- N -(trifluoroacetyl)-L-alanyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 94 0.011 2 0.379 2 (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-3-[(2 S )- 4,4-difluoro-2-(2,2,2-trifluoroacetamido)butanoyl]-6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide 95 0.003 2 0.098 4 N- (methoxycarbonyl)-3-methyl-L-valyl-(4 R )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}- 4-(trifluoromethyl)-L-prolinamide 96 0.844 one 9.190 2 (1 R ,2 S ,5 S ) -N -{(1 R )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide 97 0.011 five 0.082 16 (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C86 (DIAST-2) 98 0.188 one 1.738 2 (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3- [2-(2,2,2-trifluoroacetamido)-3-(trifluoromethyl)pentanoyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, from C85 (DIAST-1)

1. Not defined.

2. The regiochemistry of Example 41 is not well defined; other possible structures for this example are N -[( 2S )-1-({( 1S )-1-cyano-2-[( 3S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4 ,4-dimethyl-1-oxopentan-2-yl]-4-methoxy-5,6-bis(trifluoromethyl)-1 H -indole-2-carboxamide and N -[(2 S )-1-({(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}amino)-4,4-dimethyl-1-oxopentan-2-yl]-4-methoxy-6,7 -bis(trifluoromethyl)-1 H -indole-2-carboxamide.

Predicted pharmacokinetic parameters of (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl -3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide (compound of Example 13) in humans

Based on modeling of physiologically based pharmacokinetics (PBPK) in-vitro data including CL _int from human liver microsomes and CL _bile from human hepatocytes under sandwich culture conditions, predicted CL and V _ss in human plasma for (1 R ,2 S ,5 S )- N -{(1 S )-1-Cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N -( trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide are 5.9 ml / min / kg and 0.97 l / kg, respectively, and give an effective half-life, t _{1 /2} , 1.9 hours. Target C _eff 0.16 mM (unbound plasma concentration) is determined based on antiviral inhibition data obtained from in vitro studies of (1 R ,2 S ,5 S ) -N -{(1 S )-1-Cyano-2-[ (3 S )-2-oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0] hexane-2-carboxamide with either VeroE6 cells in the presence of a P-gp inhibitor (EC ₉₀ value 0.156 mM) or in a differentiated normal human bronchial epithelial cell (dNHBE) assay (EC ₉₀ value 0.149 mM). 380 mg dose _ _ _ -3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide administered orally three times a day (TID) is expected to cover the effective unbound concentration 0.16 mm at C _min .

All patents and publications described above are incorporated herein by reference in their entirety. Although the invention has been described in terms of various preferred embodiments and specific examples, the invention is to be understood as not being limited by the above detailed description, but as defined by the appended claims and their equivalents.

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SEQUENCE LIST

<110> Pfizer Inc.

Owen, Dafydd

Pettersson, Martin

Reese, Matthew

Sammons, Matthew

Tuttle, Jamison

Yang, Qingyi

Yang, Xiaojing

Verhoest, Patrick

<120> NITRILE-CONTAINING ANTIVIRAL COMPOUNDS

<130> PC072660A

<150> US 63/073,982

<151> 2020-09-03

<150> US 63/143,435

<151> 2021-01-29

<150> US 63/170,158

<151> 2021-04-02

<150> US 63/194,241

<151> 2021-05-28

<160> 2

<170>PatentIn version 3.5

<210> 1

<211> 306

<212> PROTEIN

<213> Homo sapiens

<400> 1

Ser Gly Phe Arg Lys Met Ala Phe Pro Ser Gly Lys Val Glu Gly Cys

1 5 10 15

Met Val Gln Val Thr Cys Gly Thr Thr Thr Leu Asn Gly Leu Trp Leu

20 25 30

Asp Asp Val Val Tyr Cys Pro Arg His Val Ile Cys Thr Ser Glu Asp

35 40 45

Met Leu Asn Pro Asn Tyr Glu Asp Leu Leu Ile Arg Lys Ser Asn His

50 55 60

Asn Phe Leu Val Gln Ala Gly Asn Val Gln Leu Arg Val Ile Gly His

65 70 75 80

Ser Met Gln Asn Cys Val Leu Lys Leu Lys Val Asp Thr Ala Asn Pro

85 90 95

Lys Thr Pro Lys Tyr Lys Phe Val Arg Ile Gln Pro Gly Gln Thr Phe

100 105 110

Ser Val Leu Ala Cys Tyr Asn Gly Ser Pro Ser Gly Val Tyr Gln Cys

115 120 125

Ala Met Arg Pro Asn Phe Thr Ile Lys Gly Ser Phe Leu Asn Gly Ser

130 135 140

Cys Gly Ser Val Gly Phe Asn Ile Asp Tyr Asp Cys Val Ser Phe Cys

145 150 155 160

Tyr Met His His Met Glu Leu Pro Thr Gly Val His Ala Gly Thr Asp

165 170 175

Leu Glu Gly Asn Phe Tyr Gly Pro Phe Val Asp Arg Gln Thr Ala Gln

180 185 190

Ala Ala Gly Thr Asp Thr Thr Ile Thr Val Asn Val Leu Ala Trp Leu

195 200 205

Tyr Ala Ala Val Ile Asn Gly Asp Arg Trp Phe Leu Asn Arg Phe Thr

210 215 220

Thr Thr Leu Asn Asp Phe Asn Leu Val Ala Met Lys Tyr Asn Tyr Glu

225 230 235 240

Pro Leu Thr Gln Asp His Val Asp Ile Leu Gly Pro Leu Ser Ala Gln

245 250 255

Thr Gly Ile Ala Val Leu Asp Met Cys Ala Ser Leu Lys Glu Leu Leu

260 265 270

Gln Asn Gly Met Asn Gly Arg Thr Ile Leu Gly Ser Ala Leu Leu Glu

275 280 285

Asp Glu Phe Thr Pro Phe Asp Val Val Arg Gln Cys Ser Gly Val Thr

290 295 300

Phe Gln

305

<210> 2

<211> 306

<212> PROTEIN

<213> Homo sapiens

<400> 2

Ser Gly Phe Arg Lys Met Ala Phe Pro Ser Gly Lys Val Glu Gly Cys

1 5 10 15

Met Val Gln Val Thr Cys Gly Thr Thr Thr Leu Asn Gly Leu Trp Leu

20 25 30

Asp Asp Val Val Tyr Cys Pro Arg His Val Ile Cys Thr Ser Glu Asp

35 40 45

Met Leu Asn Pro Asn Tyr Glu Asp Leu Leu Ile Arg Lys Ser Asn His

50 55 60

Asn Phe Leu Val Gln Ala Gly Asn Val Gln Leu Arg Val Ile Gly His

65 70 75 80

Ser Met Gln Asn Cys Val Leu Lys Leu Lys Val Asp Thr Ala Asn Pro

85 90 95

Lys Thr Pro Lys Tyr Lys Phe Val Arg Ile Gln Pro Gly Gln Thr Phe

100 105 110

Ser Val Leu Ala Cys Tyr Asn Gly Ser Pro Ser Gly Val Tyr Gln Cys

115 120 125

Ala Met Arg Pro Asn Phe Thr Ile Lys Gly Ser Phe Leu Asn Gly Ser

130 135 140

Cys Gly Ser Val Gly Phe Asn Ile Asp Tyr Asp Cys Val Ser Phe Cys

145 150 155 160

Tyr Met His His Met Glu Leu Pro Thr Gly Val His Ala Gly Thr Asp

165 170 175

Leu Glu Gly Asn Phe Tyr Gly Pro Phe Val Asp Arg Gln Thr Ala Gln

180 185 190

Ala Ala Gly Thr Asp Thr Thr Ile Thr Val Asn Val Leu Ala Trp Leu

195 200 205

Tyr Ala Ala Val Ile Asn Gly Asp Arg Trp Phe Leu Asn Arg Phe Thr

210 215 220

Thr Thr Leu Asn Asp Phe Asn Leu Val Ala Met Lys Tyr Asn Tyr Glu

225 230 235 240

Pro Leu Thr Gln Asp His Val Asp Ile Leu Gly Pro Leu Ser Ala Gln

245 250 255

Thr Gly Ile Ala Val Leu Asp Met Cys Ala Ser Leu Lys Glu Leu Leu

260 265 270

Gln Asn Gly Met Asn Gly Arg Thr Ile Leu Gly Ser Ala Leu Leu Glu

275 280 285

Asp Glu Phe Thr Pro Phe Asp Val Val Arg Gln Cys Ser Gly Val Thr

290 295 300

Phe Gln

305

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Claims (35)

1. Compound (1 R ,2 S ,5 S ) - N - {(1 S ) -1-cyano-2-[(3 S ) -2-oxopyrrolidin-3-yl] ethyl} -6,6-dimethyl -3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide having the structure

; or its solvate or hydrate. 2. The compound according to claim 1 which is crystalline ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide. 3. The compound according to claim 2, which is (1 R ,2 S ,5 S )- N - {(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl }-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 1, which is characterized by an X-ray powder diffraction pattern along essentially as shown in FIG. 3 and containing diffraction peaks at 7.6, 9.8, 11.4, 11.9, 12.7, 15.7, 15.8, 17.3, 17.8, 18.3, 18.9 , 19.7, 19.9, 20.5, 21.0, 21.7, 22.2, 22.5, 23.1, 23.6, 24.7, 25.3, 27.0, 27 .2, 27.9, 28.1, 29.5, 32.6, 35.7 and 37.0 degrees 2 theta, where each peak is ± 0.2 degrees 2 theta. 4. The compound according to claim 2, which is characterized by a peak in solid state ¹⁹ F NMR with a chemical shift at -73.3 ± 0.1 ppm and peaks in solid state ¹³ C NMR with chemical shifts at 31.0 ± 0.1 ppm, 27.9±0.1 ppm and 178.9±0.2 ppm 5. The compound according to claim 2, which is (1 R ,2 S ,5 S )- N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl]ethyl }-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 4, which is characterized by an X-ray powder diffraction pattern along essentially as shown in FIG. 6 and containing diffraction peaks at 7.6, 9.8, 10.8, 11.2, 11.4, 11.7, 12.0, 12.3, 12.7, 13.7, 14.9 , 15.1, 15.9, 17.5, 18.0, 18.2, 18.5, 18.8, 20.0, 20.4, 20.7, 21.1, 21.6, 21 .8, 22.3, 23.1, 23.4, 24.2, 24.9, 25.2, 26.1, 27.0, 27.2, 28.1, 28.9, 29.4 , 29.5, 29.8, 30.0, 30.6, 30.8, 31.3, 31.8, 32.5, 32.8, 33.2, 33.4, 35.5, 35 .6, 36.0, 36.4, 37.1, 38.7, 39.4, 39.5 and 39.8 degrees 2 theta, where each peak is ± 0.2 degrees 2 theta. 6. The compound according to claim 2, which is characterized by one or more peaks selected from the group consisting of a peak in solid state ¹⁹ F NMR with a chemical shift at -73.6 ± 0.1 ppm and peaks in solid ¹³ C NMR with chemical shifts at 26.9±0.1 ppm, 21.6±0.1 ppm and 41.5±0.1 ppm. 7. The compound according to claim 1 which is amorphous ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide. 8. The compound according to claim 1, which is a solvate of (1 R ,2 S ,5 S ) -N - {(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide with methyl tert -butyl ether. 9. A compound according to claim 8, characterized in that the compound is crystalline. 10. The compound according to claim 9, which is a solvate of (1 R ,2 S ,5 S ) -N - {(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidin-3-yl] ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide with methyl tert -butyl ether, solid form 2 , which is characterized by an X-ray powder diffraction pattern essentially as shown in FIG. 1 and containing diffraction peaks at 7.1, 10.5, 11.3, 11.8, 12.5, 12.9, 14.2, 15.7, 16.0, 16.8, 17.0 , 18.5, 18.8, 19.1, 19.9, 20.2, 20.8, 21.1, 21.4, 21.7, 22.2, 23.1, 23.4, 23 .7, 25.3, 27.3, 27.9, 28.3, 28.5, 29.1, 29.4, 30.2, 30.8, 32.0, 33.3, 33.8 , 35.4, 36.4 and 38.1 degrees 2 theta, where each peak is ± 0.2 degrees 2 theta. 11. Pharmaceutical composition for the treatment of coronavirus infection , containing a therapeutically effective amount of the compound -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide having the structure

; or a solvate or hydrate thereof, together with a pharmaceutically acceptable carrier. 12. A method of treating a coronavirus infection in a patient, wherein the method comprises administering a therapeutically effective amount of the compound (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2- oxopyrrolidin-3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide having the structure

; or a solvate or hydrate thereof to a patient in need thereof. 13. The method of claim 12, wherein the coronavirus infection is COVID-19. 14. Pharmaceutical composition according to claim 11, in which the compound is a crystalline 3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide. 15. Pharmaceutical composition according to claim 14, wherein the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3 -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 1, which is characterized by peak in solid state ¹⁹ F NMR chemical shift at -73.3 ± 0.1 ppm and peaks in solid state ¹³ C NMR chemical shift at 31.0 ± 0.1 ppm, 27.9 ± 0 .1 ppm and 178.9 ± 0.2 ppm 16. Pharmaceutical composition according to claim 14, wherein the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3 -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 4, which is characterized by one or more peaks selected from the group consisting of solid-state ¹⁹ F NMR chemical shift peaks at -73.6 ± 0.1 ppm and solid-state ¹³ C NMR chemical shift peaks at 26.9 ± 0, 1 ppm, 21.6 ± 0.1 ppm and 41.5 ± 0.1 ppm 17. Pharmaceutical composition according to claim 11, wherein the compound is amorphous (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxo 3-yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide. 18. The method according to claim 12, wherein the compound is crystalline (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3 -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide. 19. The method according to claim 18, wherein the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 1, which is characterized by the peak in solid state ¹⁹ F NMR with chemical shift at -73.3 ± 0.1 ppm and peaks in solid state ¹³ C NMR with chemical shifts at 31.0 ± 0.1 ppm, 27.9 ± 0, 1 ppm and 178.9 ± 0.2 ppm 20. The method according to claim 18, wherein the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 4, which is characterized by one or several peaks selected from the group consisting of a solid-state ¹⁹ F NMR chemical shift peak at -73.6 ± 0.1 ppm and a solid-state ¹³ C NMR chemical shift peak at 26.9 ± 0.1 ppm, 21.6 ± 0.1 ppm and 41.5 ± 0.1 ppm 21. The method of claim 12 wherein the compound is amorphous (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3 -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide. 22. Compound (1 R ,2 S ,5 S ) - N - {(1 S ) -1-cyano-2-[(3 S ) -2-oxopyrrolidin-3-yl] ethyl} -6,6-dimethyl -3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide having the structure

23. Pharmaceutical composition according to claim 11, in which the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3 -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide having the structure

24. The method according to claim 12, wherein the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3- yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide having the structure

25. Pharmaceutical composition according to claim 14, wherein the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3 -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 1, which is characterized by an X-ray powder diffraction pattern essentially as shown in FIG. 3 and containing diffraction peaks at 7.6, 9.8, 11.4, 11.9, 12.7, 15.7, 15.8, 17.3, 17.8, 18.3, 18.9 , 19.7, 19.9, 20.5, 21.0, 21.7, 22.2, 22.5, 23.1, 23.6, 24.7, 25.3, 27.0, 27 .2, 27.9, 28.1, 29.5, 32.6, 35.7 and 37.0 degrees 2 theta, where each peak is ± 0.2 degrees 2 theta. 26. Pharmaceutical composition according to claim 14, wherein the compound is (1 R ,2 S ,5 S ) -N -{(1 S )-1-cyano-2-[(3 S )-2-oxopyrrolidine-3 -yl]ethyl}-6,6-dimethyl-3-[3-methyl- N- (trifluoroacetyl)-L-valyl]-3-azabicyclo[3.1.0]hexane-2-carboxamide, solid form 4, which is characterized by an X-ray powder diffraction pattern essentially as shown in FIG. 6 and containing diffraction peaks at 7.6, 9.8, 10.8, 11.2, 11.4, 11.7, 12.0, 12.3, 12.7, 13.7, 14.9 , 15.1, 15.9, 17.5, 18.0, 18.2, 18.5, 18.8, 20.0, 20.4, 20.7, 21.1, 21.6, 21 .8, 22.3, 23.1, 23.4, 24.2, 24.9, 25.2, 26.1, 27.0, 27.2, 28.1, 28.9, 29.4 , 29.5, 29.8, 30.0, 30.6, 30.8, 31.3, 31.8, 32.5, 32.8, 33.2, 33.4, 35.5, 35 .6, 36.0, 36.4, 37.1, 38.7, 39.4, 39.5 and 39.8 degrees 2 theta, where each peak is ± 0.2 degrees 2 theta.

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