AT-7519

The main proteinase (M^pro) of SARS-CoV-2 plays a critical role in cleaving viral polyproteins into functional proteins required for viral replication and assembly, making it a prime drug target for COVID-19. It is well known that noncompetitive inhibition offers potential therapeutic options for treating COVID-19, which can effectively reduce the likelihood of cross-reactivity with other proteins and increase the selectivity of the drug. Therefore, the discovery of allosteric sites of M^pro has both scientific and practical significance. In this study, we explored the binding characteristics and inhibiting process of M^pro activity by two recently reported allosteric inhibitors, pelitinib and AT7519 which were obtained by the X-ray screening experiments, to probe the allosteric mechanism via molecular dynamic (MD) simulations. We found that pelitinib and AT7519 can stably bind to M^pro far from the active site. The binding affinity is estimated to be -24.37 ± 4.14 and - 26.96 ± 4.05 kcal/mol for pelitinib and AT7519, respectively, which is considerably stable compared with orthosteric drugs. Furthermore, the strong binding caused clear changes in the catalytic site of M^pro, thus decreasing the substrate accessibility. The community network analysis also validated that pelitinib and AT7519 strengthened intra- and inter-domain communication of M^pro dimer, resulting in a rigid M^pro, which could negatively impact substrate binding. In summary, our findings provide the detailed working mechanism for the two experimentally observed allosteric sites of M^pro. These allosteric sites greatly enhance the 'druggability' of M^pro and represent attractive targets for the development of new M^pro inhibitors.

Small molecules that induce protein degradation hold the potential to overcome several limitations of the currently available inhibitors. Monovalent or molecular glue degraders, in particular, enable the benefits of protein degradation without the disadvantages of high molecular weight and the resulting challenge in drug development that are associated with bivalent molecules like Proteolysis Targeting Chimeras. One key challenge in designing monovalent degraders is how to build in the degrader activity─how can we convert an inhibitor into a degrader? If degradation activity requires very specific molecular features, it will be difficult to find new degraders and challenging to optimize those degraders toward drugs. Herein, we demonstrate that an unexpectedly wide range of modifications to the degradation-inducing group of the cyclin K degrader CR8 are tolerated, including both aromatic and nonaromatic groups. We used these findings to convert the pan-CDK inhibitors dinaciclib and AT-7519 to Cyclin K degraders, leading to a novel dinaciclib-based compound with improved degradation activity compared to CR8 and confirm the mechanism of degradation. These results suggest that general design principles can be generated for the development and optimization of monovalent degraders.