Page 153 - SRPSKO DRUŠTVO ISTRAŽIVAČA RAKA

SDIRSACR Oncology Insights

paradoxical role of USP7 in p53 regulation together with its importance in ubiquitination processes, it is a critical target
for the development of novel therapeutic strategies in cancer.
Materials and Methods: Within the scope of this study, we aimed to explore the possible structural impacts of missense
variants in USP7 on the coordination of MDM2-p53 binding together with the selection of effective inhibitors. For this
purpose, we utilized TCGA database to select the missense variants in USP7 with a pathogenity score >70%, already
associated with the solid cancer. By using VMD tool, we prepared mutant and native USP7 systems that were further
subjected to classical Molecular Dynamic simulations with CHARMM parameters. Molecular Dynamic simulations
were collected as NpT ensemble for 100 ns. Then, the impacts of the pathogenic missense variants on USP7-ubiquitin
aldehyde and USP7-inhibitor binding were evaluated via protein-protein and protein-small molecule docking studies.
Results: Depending on the results of 100 ns MD simulations, we have reported that 10 out of 24 variants were leading
to significant change in the structural properties of USP7 by causing thermodynamic instability. Also, the presence
of several variants was leading to competitive binding of specific USP7 inhibitors towards the ubiquitin aldehyde,
known as a natural ligand, to keep USP7 in active conformation. Therefore, the required structural stability for its
effective function was not achieved by USP7 due to the lack of, or partial interaction with ubiquitin aldehyde. Within
the literature, 15 different small molecules serving as USP7 inhibitors were selected to evaluate their binding potential
and effectiveness towards USP7-mutant complexes via AutoDock Vina docking tool. Among many, the binding
performances of XL177A, XL188, XL203C, ALM4, ALM5, FLX4, L55, USP7-055 and USP7-797 inhibitors towards USP7
were superior even in the presence of many missense variants in USP7. Among these, the best performance was
reported for ALM4, ALM5, FLX4 and L55 small molecules in terms of effective inhibition of USP7 even in the presence
of many missense variants. We also reported the impacts of missense variants existing in USP7-TRAF domain on the
dynamics of MDM2-p53. It was reported that R66C variant resulted in the loss of p53 coordination, by up to 30% while
maintaining MDM2 coordination due to the loss of structural flexibility within TRAF domain.
Conclusion: The data obtained in this study highlight the importance of constituting theoretical background about the
mutation-specific structural and functional changes for the effective inhibitor selection for USP7.

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