Investigating Wee1 and Myt1 combined inhibition as a potential cancer therapeutic strategy
Creators: Sokhi, Sargun
Publisher: name: University of Alberta Library
Year: 2024

<p>The genomic integrity of a cell is under constant threat of damage due to exposure to endogenous or exogenous sources, resulting in surveillance by cell cycle checkpoints. These checkpoints are present at G1/S, Intra-S, G2/M and intra-M (mitotic phase); and put a pause to the proliferative signaling until any metabolic, genetic, or physical damage is resolved. Wee1 and Myt1 kinases are two checkpoint kinases that prevent premature entry into mitosis at the G2/M checkpoint by adding inhibitory phosphorylation on Cdk1/cyclin B1 (main mitosis driving complex). While Wee1 phosphorylates Cdk1 on tyrosine 15, Myt1 is a dual kinase that phosphorylates Cdk1 on tyrosine 15 as well as threonine 14. Due to partial redundancy in Myt1 and Wee1’s function, there has been a lack in research needed on Myt1 on the basis of it being a kinase of less importance. Although there are conflicting results on Myt1’s role in maintaining G2/M checkpoint through Cdk1 inhibition, many reports in the literature have highlighted the importance of Myt1 as an independent and required player in controlling cell cycle progression. Many research groups, including ours, have shown how Myt1 can be providing a selective advantage to the cancer cells. Myt1 upregulation has been observed to be a common mechanism by which cancer cells buy enough time to repair DNA damage thereby allowing them to proliferate and expand in number. In light of all this recent evidence, we undertook to investigate the role of Myt1 as a potential cancer therapeutic target. We found out that Myt1 overexpression not only mediates resistance to Adavosertib but also to other cell cycle checkpoint kinase inhibitor (PD166285) and DNA damage checkpoint kinase inhibitors (AZD6738 and UCN-01). These inhibitors prevent inhibitory phosphorylation on Cdk1 leading to abrogation of G2/M checkpoint and premature entry into mitosis. High Cdk1 activity also leads to prolonged mitotic arrest with cells dying as a result of mitotic catastrophe. We demonstrate that Myt1 confers resistance by compensating Cdk1 inhibition in the presence of these different kinase inhibitors. Myt1 overexpression leads to reduced premature mitotic entry and decreased length of mitosis eventually leading to increased survival rates in Adavosertib treated cells. Elevated Myt1 levels also conferred resistance to inhibitors of ATR or Chk1 inhibitor. <br>
To overcome Myt1 overexpression mediated resistance, we investigated Myt1 and Wee1 combined inhibition using as a potential cancer therapeutic strategy. We found that MK-1775 and RP-6306 combination treatment shows synergistic cell killing in a panel of cancer cell lines. Wee1 inhibition shows synergistic cell killing with Myt1 inhibition with effect also pronounced in inducible Myt1 overexpressing cells. Wee1/Myt1 combined inhibition resists the increase in clonogenic potential of the cancer cells transiently overexpressing Myt1. RP-6306 and MK-1775 combination treatment prevents Cdk1 inhibition on both tyrosine 15 and threonine 14 and abrogates G2/M checkpoint. This leads to premature entry into mitosis and also promotes prolonged mitotic arrest leading to cell death. We found that one of the prominent mechanisms of cell death in the cells treated with the combination treatment is centromere fragmentation leading to mitotic catastrophe. My research explores the mechanistic understanding of the synthetic lethality observed with the Wee1 and Myt1 small molecule inhibitors while contributing to the optimization of a novel cancer therapeutic strategy.</p>