“Alkyne as a Latent Warhead to Covalently Target SARS-CoV‑2 Main Protease”. Article review

We are all aware of the impact of the COVID-19 pandemic generated by the SARS-CoV-2 virus on the pharmaceutical sector. It appears to be a thing of the past, with essentially no new information about SARS-CoV-2 available in the public domain.

However, issues continue to occur as a result of novel viral varieties and limits in existing immunization techniques. Despite tremendous progress in vaccine research, questions about their efficacy against novel viral varieties and the vulnerability of specific populations, such as immunocompromised people, remain.

As a result, the emphasis is on the development of antiviral medications that target the SARS-CoV-2 main protease (M_pro), a critical enzyme in the virus's reproduction cycle. M_pro is a protein that is involved in the processing of viral polyproteins and is thought to be a prospective therapeutic target.

The article titled "Alkyne as a Latent Warhead to Covalently Target SARS-CoV-2 Main Protease" discusses the structure and function of SARS-CoV-2 M_pro, emphasizing its unique characteristics, including a consensus substrate cleavage sequence, which makes it an attractive target for drug development. The presence of the catalytic residue Cys₁₄₅ in the active site makes M_pro susceptible to covalent inhibition by small molecule inhibitors.

Рic.1 Proposed irreversible thiol−alkyne addition of alkyne-containing inhibitors withMpro to form a vinyl sulfide linkage (doi.org/10.1021/acs.jmedchem.3c00810).

The article mentions the development of various covalent inhibitors, including Paxlovid, which has shown efficacy in reducing the risk of severe disease and hospitalizations. However, limitations and resistance issues with existing inhibitors emphasize the need for new drugs targeting M_pro.

The following article introduces the concept of electrophilic warheads, with a particular focus on latent warheads that offer specificity and reduce off-target effects. The authors describe their efforts to identify M_pro inhibitors using novel electrophilic warheads, particularly emphasizing the use of terminal alkyne as a latent warhead.

The researchers tested several peptidomimetic scaffolds and discovered that alkyne-containing inhibitors inhibited M_pro strongly. Biochemical and structural investigations were used to better describe these inhibitors and their potential as "clickable" probes for monitoring target interaction was investigated.

Importantly, alkyne-containing inhibitors have shown antiviral action in COVID-19 infection cell models, highlighting their promise as a new class of medications for the disease's treatment.

Biochemical and structural analyses showed that the alkyne forms an irreversible vinyl sulfide bond with the catalytic cysteine M_pro, indicating its potential as an effective inhibitor. The best alkyne-containing inhibitors were found to prevent SARS-CoV-2 infection in cellular models, indicating that alkynes can be used as latent warheads to target cysteine proteases not only in viruses but potentially in other contexts.

The advantage of latent warheads, as explained by the authors, is their minimal internal reactivity. They become active for covalent inhibition only in the active site of the respective enzymes. This selectivity is crucial for reducing off-target effects. The excerpt notes that compounds with latent alkynes as warheads tend to have fewer unintended covalent interactions compared to conventional warheads.

However, the researchers noticed that the rate of inactivation of M_pro by inhibitors containing latent alkyne warheads was relatively slow, requiring a long incubation period for potent in vitro inhibition. This slow inactivation may affect the effectiveness of these compounds in fighting SARS-CoV-2 infection, which is rapidly evolving in real-time. To overcome this limitation, the researchers increased the inactivation rate of M_pro by modifying the alkene warhead with an electron acceptor group CF₃.

Pic.1 Tuning up alkyne reactivity through a chemical modification. Alkyne substitution with an electron withdrawing group, trifluoromethyl, to tune its warhead reactivity (doi.org/10.1021/acs.jmedchem.3c00810).

The article concludes by acknowledging that further research is needed to determine whether terminal or appropriately substituted alkynes can indeed serve as effective latent warheads for targeted covalent inhibition in therapeutic development. In essence, the study focuses on the potential of alkynes as a new approach to develop inhibitors of M_pro, a key target in the fight against SARS-CoV-2.