In my Chordoma Open Lab Notebook entries, I will tell you about a project that is very important to me, targeting the transcription factor brachyury (also known as TBXT) for the rare cancer chordoma. In the coming weeks and months, the amount of science strategy, science details, and science progress shared here will rise. Today’s entry will simply introduce the project to you, tell you how we got here, and share my motivation. By the way, it will help me and the team if you let me (us!) know what you think, what questions you have, and what you’d like to know. Please feel free to share these posts with others via email links or social media. In addition, a long career has taught me that I don’t have all the answers! If you have ideas, please share them with us!
Working on Chordoma is personal for me. Eighteen years ago, my daughter Carolyn was in first grade and a bright and joyful little boy in her class named Justin (pictured below, graphic from Chordoma Foundation) was diagnosed with chordoma. Of course, I had never heard about this rare cancer which strikes only one in a million people. In the US your chances of getting hit by lightning are higher. Justin and his wonderful parents shared his difficult journey with his classmates and their parents. Justin wanted to talk about this. Justin drew his friends in and inspired them to find joy and strength and motivation in his journey. Their enthusiasm in turn inspired me. As a scientist I wanted to help for Justin, Justin’s parents, my daughter Carolyn, and all of Justin’s classmates who gave what they could of their time, talents, and treasure.
A number of years after Justin’s diagnosis I met Josh Sommer, a Chordoma patient himself, and Co-Founder and Executive Director of the Chordoma Foundation. Josh attended a number of fund raisers that my daughter and her classmates were involved in, and one thing led to another and I found ways to get involved. Josh inspired me with his positive attitude, drive, and ability to band people together. He is a connector, and science thrives on connections. We are all better when we work together. Josh introduced me to Dr. Adrienne Flanagan, a leader in chordoma research, a key thought leader on the links between brachyury and chordoma, and a real blessing to the chordoma community for her hard work, insights, and passion. I was able to get involved by leveraging an open science project I was working on at the pharmaceutical company GlaxoSmithKline (GSK) sharing inhibitors of an important class of proteins called kinases. Adrienne and her team tested them, and we were able to make some interesting connections between kinase inhibition and chordoma cell line growth. I believe there is more to be done with kinase inhibition and chordoma, but that’s a story for another day.
I have since moved from GSK to the Eshelman School of Pharmacy at the University of North Carolina at Chapel Hill. Our SGC-UNC team (https://www.sgc-unc.org/) is the first US site of the Structural Genomics Consortium (SGC, https://www.thesgc.org/) a worldwide public private partnership committed to supporting the discovery of new medicines through open science. We generate tools and reagents and share them with the scientific community openly and without restrictions. We want to move quickly, we want to facilitate research programs in other groups, and we want to help patients.
TBXT (brachyury): a key target for chordoma
Emerging data suggests that the transcription factor TBXT is a key vulnerability of chordoma and a prime candidate to target in order to create medicines to treat chordoma. For example, TBXT knockdown stops the proliferation of chordoma cell lines and reduces their viability. TBXT is the top selectively essential gene in chordoma. A great place to start if you want to know more about TBXT and chordoma is this paper by Tanaz Sharifnia and colleagues (link here: https://pubmed.ncbi.nlm.nih.gov/30664779/). As a transcription factor, it falls in the “difficult target” category. There is no active site tailor made for small molecules. Details of downstream signaling via protein-protein interactions are poorly understood. On the plus side, TBXT is expressed in development but almost nonexistent (or very low expression) in most adult human tissues, suggesting that if we are successful, compounds may have a useful therapeutic index. The high unmet need and weight of evidence convince us that TBXT is an Achilles heel of chordoma that needs to be addressed through a targeted drug discovery program.
Our initial thought was to see if we could identify small molecule binders for TBXT using x-ray crystallography. To get there we needed a number of things to fall into place. I am grateful to the Mark Foundation and Chordoma Foundation for funding our initial proposal that allowed us to generate enough data to convince ourselves that this strategy holds great promise.
As part of the SGC, I was well aware of the SGC Target Enabling Package (TEP) program and realized it was an ideal mechanism to facilitate study of brachyury. You can read more about the TEP program here: https://www.thesgc.org/tep. I was excited to nominate TBXT as a TEP target, and absolutely thrilled when SGC scientist Opher Gileadi (Oxford University, SGC) agreed to work on the project. Opher and his talented team started the project by expressing and purifying the TBXT protein and then in short order they identified conditions to reproducibly grow crystals of the protein. They then collaborated with the team at Diamond to do an x-ray crystallography based high throughput fragment screen using the XChem platform (more information on XChem here: https://www.diamond.ac.uk/Instruments/Mx/Fragment-Screening.html#). This resulted in more than 40 fragments binding to a number of different sites on the transcription factor. As a medicinal chemist, it is an exciting day when you see a range of small molecules bound to your target of interest! With x-ray evidence that there are hot spots for binding small molecules to TBXT, we have now set our sights on a more challenging goal and bigger prize, optimization of these fragments into molecules that modulate TBXT biology and kill chordoma cells. In coming posts, we will provide more detail on the initial fragment screen and start walking through the steps we are taking.
Some of our project goals
- Develop and provide reagents the community can use to explore TBXT biology (eg: compounds, protein constructs, biophysical assays, cell assays, antibodies)
- Develop assays to measure affinity (thermal shift, SPR)
- Synthesize analogues of fragments at each site in order to optimize binding potency
- Generate crystal structures of new and improved compounds
- Generate Structure Activity Relationships that allow us or others to generate reagents that can lead to the degradation of TBXT
- Understand impact potent fragments (and degraders) have on TBXT biology
Project Summary – things you will hear more about as the project progresses
To date we have exemplified reproducible crystallography of TBXT and demonstrated the utility of fragment screening as a means of identifying TBXT hotspots for binding small molecules. The primary aim of the current project is to develop small molecule inhibitors of TBXT or of its interaction partners and identify at least one tractable lead molecule targeting TBXT that has potential for final optimization into a candidate compound. As we progress towards this end, we also aim to understand the structural basis of the activities of TBXT and contribute to the understanding of the mechanism of the disease. At the end of this two-year project, we will have the infrastructure, assays, reagents, and compounds in place that form the basis for a drug discovery project to move TBXT modulators into the clinic.
In the coming weeks different members of the project team will provide updates. Here is a very brief team introduction. Currently our team is located at three sites.
- SGC-UNC at the UNC Eshelman School of Pharmacy: David Drewry, Carrow Wells
- SGC at Oxford University: Opher Gileadi, Joseph Newman, Angeline Gavard
- Institute for Cancer Research: Paul Workman, Paul Clarke, Hadley Sheppard
We also have guidance and support from the teams at our generous funders, the Mark Foundation for Cancer Research (Becky Bish, Ryan Schoenfeld, Michele Cleary) and the Chordoma Foundation (Josh Sommer, Joan Levy). Thank you!
You can find out more about these organizations here:
Mark Foundation for Cancer Research: https://themarkfoundation.org/
The Chordoma Foundation: https://www.chordomafoundation.org/