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Every form of tobacco-related lung cancer has at least one thing in common: deregulated E2F. E2F is a transcription factor, a protein responsible for turning on and off the cell’s ability to read, or transcribe, genes. The genes that E2F controls are affiliated with promoting cell proliferation. Too much of this will lead to tumor growth and cancer. In a healthy cell, E2F spends most of its time in an inactive state, where it is naturally inhibited by the retinoblastoma protein, Rb. Rb prevents E2F activity throughout what is called the “gap 1” phase, or G1, of the cell’s life cycle. As the cell transitions to its next phase, the synthesis phase, an enzyme named Cdk interacts with Rb, causing Rb to release E2F which then regains its activity. In all tobacco-related lung cancers, E2F is found to be overactive which leads to a constant read-me mode of the genes it is responsible for, and in turn an over-proliferation of cells. Despite this universal trigger of cancer, there is currently no therapy that directly targets E2F.
I propose the development of a novel strategy that targets E2F directly, not through inhibition, but by degrading it entirely. The approach employs proteolysis-targeting chimera (PROTAC) technology, wherein a bifunctional molecule contains one motif designed to bind E2F and another to recruit an E3 ubiquitin ligase. E3 ubiquitin ligases are enzymes that facilitate the transfer of ubiquitin to a target. Ubiquitin is a molecule that acts as a signal to recruit degradation machinery to its attached target. PROTACs force the interaction of a target with E3 ubiquitin ligase, sentencing the target to a degradative end. My overall hypothesis is that selective degradation of active E2F will lead to cell cycle arrest and/or cell death of tobacco-related lung cancer cells. To achieve the goal of creating and testing an E2F-degrading PROTAC, the project has two main aims:
- Test the hypothesis using a model cell-permeable peptide PROTAC in a panel of lung adenocarcinoma cancer cells
- Identify a molecule that binds an E2F regulatory domain and can be developed into an E2F PROTAC.
Transcription factors like E2F are historically undruggable targets; their geometry is unapproachable to traditional drugs that need a snug fit to ensure long-lasting inhibition. A PROTAC could overcome this challenge since its function is not to provide long-lasting inhibition, but to facilitate interaction between its target and E3 ligase, after which the target is destined for degradation. By designing an E2F-targeting PROTAC, I will create the first PROTAC aimed at a transcription factor, thus opening a door to the world of undruggables.
By targeting the Rb-binding interface of an E2F regulatory domain, yet designing the E2F-binding motif of the PROTAC to have a weaker binding affinity than Rb, I will specifically target active E2F. This gives the PROTAC the therapeutic advantage of specificity and potency. For the same reason, this PROTAC would also give researchers unprecedented control over the presence of E2F in vivo, allowing new ventures in cell cycle studies.
The outcomes of this research would not only provide a novel strategy that can later be developed into a tobacco-related lung cancer therapeutic, but will empower future research with a unique tool for probing the cell cycle. |
