- This study identified the key differences in the binding pocket of the main protease in the SARS-CoV viruses, the Human corona virus, and several enteroviruses. 
- 11r and 11u were the two most promising broad spectrum inhibitors tested for the viruses.
- 11r was is the most promising in its effectiveness against human coronaviruses and the  SARS-CoV because of the similarity between the human corona virus and the SARS-CoV virus, this could lead to promising results with further testing of this inhibitor. 
- 11u is the most broad-spectrum against the viruses tested. It proved to be the most equipotent in all the viral proteases. 
- 11s and 11n proved to be less promising because of their lower potencies in other viruses however it was very effective at binding to the human corona virus binding pocket. 


Discussion/Conclusion - alpha-ketoamide's effectiveness as broad-spectrum inhibitors against SARS-CoV viruses

In the human coronavirus, they observed an IC50 of 12.5 μM but a surprisingly effective EC50 of 1.8 μM. But it was more potent in the enteroviruses. Because of the similarity between the SARS-CoV and the current human coronavirus proteases, the effectiveness the 11r inhibitor on the SARS-CoV (IC50 = 0.7 μM and EC50 = 1.8-2.1 μM) indicates that is could have a potential viral inhibition effectiveness in further studies. 

Grand Valley State University

Four successful alpha-ketoamides were observed in this study: 
- 11n
- 11r
- 11s
- 11u

List of successful alpha-ketoamide broad spectrum inhibitors

Zhang, L., Lin, D., Kusov, Y., Nian, Y., Ma, Q., Wang, J., von Brunn, A., Leyssen, P., Lanko, K., Neyts, J., de Wilde, A., Snijder, E. J., Liu, H., & Hilgenfeld, R. (2020). α-Ketoamides as Broad-Spectrum Inhibitors of Coronavirus and Enterovirus Replication: Structure-Based Design, Synthesis, and Activity Assessment. Journal of medicinal chemistry, 63(9), 4562–4578. https://doi.org/10.1021/acs.jmedchem.9b01828

α‑Ketoamides as Broad-Spectrum Inhibitors of Coronavirus and Enterovirus Replication: Structure-Based Design, Synthesis, and Activity Assessment 

The IC50 of this inhibitor was 1.08 μM which was much lower than 11a, and the EC50 was 0.6 μM which are both positive readings showing that this was a successful inhibitor. It provided much better results compared to 11a because of the isobutyl group in replacement of the butyl group which allowed it to interact with the ”lid” and the and sufficiently bind. 

However, it had very high EC50 values in the SARS-CoV virus, leading to believe it might not be as promising as a broad-spectrum inhibitor and with the human coronavirus. 

Proposed alpha-ketoamide Inhibitor: 11n 

Proposed alpha-ketoamide Inhibitor: 11r

The IC50 of the inhibitor was 1.4 μM and the EC50 was 1.3 μM. It had a high affinity for the main protease pocket because of the near-perfect size of this inhibitor in the pocket. 
Albeit promising in IC50, the binding in the enteroviruses was very weak showing it would not be a braod-spectrum inhibitor.

Proposed alpha-ketoamide Inhibitor: 11s

11u was the only inhibitor that had acceptable IC50 values for all of the six viral proteases they tested on across the enteroviruses and the coronaviruses because it was nearly equipotent in all of them. Also, their unpublished data discussing their toxicity in mice showed no toxicity with this inhibitor. 

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