Perspectives on SARS-CoV-2 Main Protease Inhibitors

Abstract

The main protease (M^pro) plays a crucial role in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication and is highly conserved, rendering it one of the most attractive therapeutic targets for SARS-CoV-2 inhibition. Currently, although two drug candidates targeting SARS-CoV-2 M^pro designed by Pfizer are under clinical trials, no SARS-CoV-2 medication is approved due to the long period of drug development. Here, we collect a comprehensive list of 817 available SARS-CoV-2 and SARS-CoV M^pro inhibitors from the literature or databases and analyze their molecular mechanisms of action. The structure-activity relationships (SARs) among each series of inhibitors are discussed. Additionally, we broadly examine available antiviral activity, ADMET (absorption, distribution, metabolism, excretion, and toxicity), and animal tests of these inhibitors. We comment on their druggability or drawbacks that prevent them from becoming drugs. This Perspective sheds light on the future development of M^pro inhibitors for SARS-CoV-2 and future coronavirus diseases.

Figure 1.

3D conformation alignment (a) and 2D sequence alignment (b) of SARS-CoV-2 and SARS-CoV main proteases (M^pro). The sequences and structures are from the Protein Data Bank (PDB) ID 7C6S (SARS-CoV-2 M^pro, cyan in part a) and 2A5I (SARS-CoV M^pro, green in part a).

Figure 2.

Overall scheme of SARS-CoV/SARS-CoV-2 M^pro catalytic mechanism by Cys145 and His41 at the active site.

Figure 3.

Subsite nomenclature for proteolytic enzymes. Amino acid residues to the left of the polypeptide scissile amide bond are numbered sequentially, beginning with P₁ and increasing toward the N-terminus. Amino acid residues to the right of the scissile bond are numbered sequentially, beginning with P₁′ and increasing toward the C-terminus. Complementary regions of the protease active site employ the corresponding S numbering.

Figure 4.

P₂-modified hydroxymethylketone (HMK) covalent inhibitors.

Figure 5.

Cocrystal structure of the covalent adduct of 2 bound to SARS-CoV-2 M^pro (PDB ID 6XHM) (a) and its corresponding 2D interaction diagram (b).

Figure 6.

Acyloxymethylketone covalent inhibitors against SARS-CoV M^pro.

Figure 7.

Cocrystal structure of the covalent adduct of 9 bound to SARS-CoV M^pro (PDB ID 6XHN) (a) and its corresponding 2D interaction diagram (b).

Figure 8.

P₃- and P₂-modified HMK and alkoxymethylketone covalent inhibitors.

Figure 9.

α-Ketoamide covalent inhibitors.

Figure 10.

Cocrystal structures of the covalent adducts of 29 (a, b) and 32 (c, d) bound to SARS-CoV or SARS-CoV-2 M^pro (PDB ID 5N5O and 6Y2G).

Figure 11.

Benzothiazole ketone-containing inhibitors 33–39.

Figure 12.

Benzothiazole ketone-containing inhibitors 40–52.

Figure 13.

Cocrystal structures of the covalent adducts of 40 bound to SARS-CoV-2 M^pro (PDB ID 7E18) (a) and its corresponding 2D interaction diagram (b).

Figure 14.

Thiazole ketone-containing inhibitors from Konno et al. and Thanigaimalai et al.

Figure 15.

Nitrile-containing drug candidate from Pfizer (PF-07321332).

Figure 16.

Bicycloproline-containing SARS-CoV-2 M^pro inhibitors derived from telaprevir.

Figure 17.

Bicycloproline-containing SARS-CoV-2 M^pro inhibitors derived from boceprevir.

Figure 18.

Crystal structure of 60 covalently bound to SARS-CoV-2 M^pro (PDB ID 7D3I) (a) and its corresponding 2D interaction diagram (b).

Figure 19.

Other aldehyde-based inhibitors.

Figure 20.

Cocrystal structures of the covalent adducts of 92 (a, b) and 105 (c, d) bound to SARS-CoV or SARS-CoV-2 M^pro (PDB ID 6M0K and 6LO0).

Figure 21.

Covalent inhibitors with aldehyde bisulfite warhead. The IC₅₀ values of 106–109 and 111 were reported by Rathnayake et al. For 110 (GC376), the IC₅₀ values of 0.62 μM and 2.2 μM for SARS-CoV-2 and SARS-CoV M^pro were given by Rathnayake et al. The IC₅₀ values 0.19 μM and 0.05 μM to SARS-CoV-2 and SARS-CoV M^pro were reported by Vuong et al. The IC₅₀ value 0.03 μM against SARS-CoV-2 M^pro was given by Ma et al.

Figure 22.

Cocrystal structures of the covalent adduct of 110 (GC376) bound to SARS-CoV-2 M^pro (PDB ID 6WTJ) (a) and its corresponding 2D interaction diagram (b).

Figure 23.

Michael acceptor-based inhibitors.

Figure 24.

Cocrystal structures of the covalent adducts of 112 (a, b), 113 (c, d), and 114 (e, f) bound to SARS-CoV M^pro or SARS-CoV-2 M^pro (PDB ID 2ZU4, 7JT7, and 2ZU5).

Figure 25.

Calpain inhibitors.

Figure 26.

Crystal structures of the covalent adducts of 118 (a, b) and 119 (c, d) bound to SARS-CoV-2 M^pro (PDB ID 6XFN and 6XA4).

Figure 27.

5-Chloropyridine ester and benzotriazole ester derived non-peptidomimetic covalent inhibitors from Ghosh et al. and Wu et al. NI represents no inhibition.

Figure 28.

5-Chloropyridyl or 5-chloropyridine ester derived non-peptidomimetic covalent inhibitors from Zhang et al. and Niu et al.

Figure 29.

Proposed mechanism of the SARS-CoV M^pro inhibition by acylation with ester-based inhibitors.

Figure 30.

Crystal structure of the covalent complex of 152 (MAC-5576) with SARS-CoV-2 M^pro (PDB ID 7JT0) through the acylation of Cys145 (a) and its corresponding 2D interaction diagram (b).

Figure 31.

Ebselen derived non-peptidomimetic covalent inhibitors.

Figure 32.

Ebsulfur derived non-peptidomimetic covalent inhibitors.

Figure 33.

Most potent noncovalent M^pro inhibitor among aryl boronic acid derivatives.

Figure 34.

N-Substituted isatin derivatives as noncovalent SARS-CoV-2 and SARS-CoV M^pro inhibitors.

Figure 35.

Noncovalent M^pro inhibitors containing benzotriazole.

Figure 36.

Most potent anilide based inhibitor.

Figure 37.

Noncovalent aldehyde inhibitors.

Figure 38.

Crystal structures of 193 (a, b) and 194 (c, d) bound to SARS-CoV M^pro (PDB ID 3AVZ and 3ATW).

Figure 39.

Symmetric peptides and molecules.

Figure 40.

Aromatic-disulfide based inhibitors.

Figure 41.

Other noncovalent M^pro inhibitors.

Figure 42.

Crystal structures of 209 (a, b) and 211 (c, d) bound to SARS-CoV-2 M^pro (PDB ID 2GZ7 and 6M2N).

Figure 43.

Metal conjugated SARS-CoV-2 or SARS-CoV M^pro inhibitors.

Figure 44.

Crystal structures of 212 (a, b), 213 (c, d), and 214 (e, f) bound to SARS-CoV-2 M^pro (PDB ID 2Z9L, 2Z9K and 2Z9G).