| dc.description.abstract | Structure-based drug design is a drug discovery strategy where rational design of drug molecules take place based on the structural information of therapeutic targets. With the development of structural biology technologies such as protein crystallography and cryo-electron microscopy, which results in the availability of more and more proteins in a higher and higher resolution, structure-based drug design has become one of the most useful drug discovery strategy in both academia and pharmaceutical industry. This dissertation discusses applying structure-based drug design strategies in inhibitor development targeting ENL (eleven-nienteen leukemia) protein, which is an important protein in the mix lineage leukemia (MLL)-rearranged leukemia, and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease, a vital viral enzyme for its replication.
Chapter I is a brief introduction to the topics of this dissertation. Starting with a short introduction of the concept of structure-based drug design, it then mainly discusses the molecular mechanism of the pathogenesis and potential therapeutic targets of the following two diseases: MLL-rearranged leukemia and COVID-19.
Chapter II describes the development of a series of selective ENL YEATS domain inhibitors. ENL is a histone acetylation reader essential for disease maintenance in acute leukemias, especially the MLL-rearranged leukemia. The function of ENL is dependent on the recognition of histone acetylation by its YEATS domain, suggesting that inhibition of the ENL YEATS domain is a potential strategy to treat MLL-rearranged leukemia. In our study, high-throughput screening of a small molecule library was carried out to identify inhibitors for the ENL YEATS domain. Structureactivity relationship studies of the hits and structure-based inhibitor design led to two compounds with IC50 values below 100 nM in inhibiting the ENL-acetyl-H3 interaction. Both compounds and their precursor displayed strong selectivity toward the ENL YEATS domain over all other human YEATS domains. One of these compounds also exhibited on-target inhibition of ENL in cultured leukemia cells and a synergistic effect with the BET bromodomain inhibitor JQ1 in killing leukemia cells. Together, we have developed selective inhibitors for the ENL YEATS domain, providing the basis for further medicinal chemistry-based optimization to advance both basic and translational research of ENL.
Chapter III and IV describes the development of SARS-CoV-2 main protease inhibitors and the assessment of their selectivity against host proteases. The COVID-19 pathogen, SARS-CoV-2, requires its main protease (SC2Mpro) to digest two of its translated long polypeptides to form mature viral proteins that are essential for viral replication and pathogenesis. Inhibition of this vital proteolytic process is effective in preventing the virus from replicating in infected cells and therefore provides a potential COVID-19 treatment option. Guided by previous medicinal chemistry studies about SARS-CoV main protease (SC1Mpro), we designed and synthesized a series of peptidyl aldehyde inhibitors that reversibly covalently bind to the active cysteine of SC2Mpro. The most potent compound has an IC50 of 8.3 nM. Crystallographic analysis confirmed the covalent linkage between the aldehyde inhibitors and active cysteine and showed structural rearrangement of the apoenzyme to accommodate the inhibitors. Two inhibitors completely prevented the SARS-CoV-2-induced cytopathogenic effect in Vero E6 cells at 2.5–5 μM and A549/ACE2 cells at 0.16–0.31 μM.
Even though a number of inhibitors have been developed for the SARS-CoV-2 main protease as potential COVID-19 medications, little is known about their selectivity. Using enzymatic assays, we characterized inhibition of TMPRSS2, furin, and cathepsin B/K/L by 11 previously developed Mpro inhibitors. Our data revealed that all these inhibitors are inert toward TMPRSS2 and furin. Diaryl esters also showed low inhibition of cathepsins. However, all aldehyde inhibitors displayed high potency in inhibiting three cathepsins. A cellular analysis indicated high potency of MPI5 and MPI8 in inhibiting lysosomal activity, which is probably attributed to their inhibition of cathepsins. Among all aldehyde inhibitors, MPI8 shows the best selectivity toward cathepsin L. With respect to cathepsin B and K. MPI8 is the most potent compound among all aldehyde inhibitors in inhibiting SARS-CoV-2 in Vero E6 cells. Cathepsin L has been demonstrated to play a critical role in the SARS-CoV-2 cell entry. By selectively inhibiting both SARS-CoV-2 MPro and the host cathepsin L, MPI8 potentiates dual inhibition effects to synergize its overall antiviral potency and efficacy. Due to its high selectivity toward cathepsin L that reduces potential toxicity toward host cells and high cellular and antiviral potency, we urge serious consideration of MPI8 for preclinical and clinical investigations for treating COVID-19. | en |
