SARS-CoV-2 Mpro
February 2022
SARS-CoV-2 Main protease
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (Mpro) is an attractive drug target for coronavirus disease (COVID-19) since it plays a critical role in viral replication and transcription, and is less vulnerable to mutations than other SARS-CoV-2 proteins. Additionally, it is a virus selective cysteine protease whose inhibitors have low toxic effects in human. Moreover, targeting proteases in other viruses such as HIV and hepatitis C lead to the development of drugs that successfully blocked the spread of these viruses.
Antiviral therapeutics are highly needed to fight COVID-19, namely, to prevent more severe symptoms and hospitalization of infected patients.
To target SARS-CoV-2 Mpro, scientists at Pfizer successfully developed a small-molecule oral drug called Paxlovid which got the FDA authorization for emergency use in December 2021.
In the race to find a drug against COVID-19, they had a head start thanks to previous research efforts to contain the SARS-CoV-1 outbreak in 2002. They identified that the lead (PF-00835231) from previous work could be used against the new virus. In fact, the Mpro SARS-CoV-1 and SARS-CoV-2 have identical binding sites (100% sequence similarity). After confirming this hypothesis in antiviral assays they adopted a lead optimization strategy aiming to increase the oral bioavailability of the compound. To do so, they intended to reduce the number of hydrogen bond (HB) donors in PF-00835231.
Each compound alteration was supported by the production of X-Ray co-crystal structures (now available in the PDB: 7RFR, 7RFU, 7RFS, 7RFW).
One of the HB donors the scientists decided to eliminate was the NH group on the ligand leucine moiety which creates an H-bond with the glutamine residue in the main protease hydrophobic pocket S2. One of the design ideas then was to introduce a cyclic leucine mimetic group (6,6-dimethyl-3-azabicyclo [3.1.0]hexane)). Computational studies predicted that this new compound (compound 3) would fit in the active site and keep a similar binding mode. The assays showed that compound 3 was active (although with a slightly lower potency) and the scientists decided to go forward with the optimization cycle. The binding mode of compound 3 was confirmed by a co-crystallized X-ray structure (Image 1).
Image 1: The hydrogen bond between the ligand leucin moiety (in blue) and the glutamine residue in the main protease (Q189), was lost when the Pfizer scientists designed compound 3 (in orange) with increased oral bioavailability. This image is captured in the 3decision 3D viewer.
The next step in the lead optimization cycle was to restore the interaction between the ligand and the glutamine and thus increase the antiviral potency. The X-ray co-crystallized structure of compound 3 (in orange, image 1) shows that the indole moiety does not fully occupy the back pocket S3.
In other words, there is room to grow here. One can imagine that the scientists used computational studies again to test ideas with different functional groups that could create new interactions with the protein. The final compound (Nirmatrelvir, part of the drug Paxlovid) contains a trifluoroacetamide moiety that reaches further into the back pocket S3 and creates several polar interactions with the protein (in purple, image 2).
Image 2: Comparison between the indole moiety in compound 3 (in blue) with the trifluoroacetamide moiety in the final compound (in purple). The trifluoroacetamide group protrudes into the S3 pocket and restores several polar interactions with the protein. This image is captured in the 3decision 3D viewer.
References:
How Pfizer scientists transformed an old drug lead into a COVID-19 antiviral, Chemical & Engineering News, Volume 100, Issue 3
Owen DR, Allerton CMN, Anderson AS, Aschenbrenner L, Avery M, Berritt S, Boras B, Cardin RD, Carlo A, Coffman KJ, Dantonio A, Di L, Eng H, Ferre R, Gajiwala KS, Gibson SA, Greasley SE, Hurst BL, Kadar EP, Kalgutkar AS, Lee JC, Lee J, Liu W, Mason SW, Noell S, Novak JJ, Obach RS, Ogilvie K, Patel NC, Pettersson M, Rai DK, Reese MR, Sammons MF, Sathish JG, Singh RSP, Steppan CM, Stewart AE, Tuttle JB, Updyke L, Verhoest PR, Wei L, Yang Q, Zhu Y. An oral SARS-CoV-2 Mpro inhibitor clinical candidate for the treatment of COVID-19. Science. 2021 Dec 24;374(6575):1586-1593. doi: 10.1126/science.abl4784. Epub 2021 Nov 2. PMID: 34726479.