If you’ve spent any time working in or around Fred Hutch Cancer Center, you might convince yourself that researchers are well on their way to treating or even curing the majority of human cancers. However, despite record levels of funding for cancer research and fundamental advances in our collective understanding of the disease, the sheer diversity of cancer types and behaviors means that there are some cancers which continue to fly under the radar. As their name suggests, these rare cancers are distinguished by their relatively low population incidence (not a low aggressiveness or mortality rate) and for this reason do not generally attract the same amount of research funding or attention as more common cancer types. If rare cancers had a poster child, it may well be cholangiocarcinoma, a cancer of the bile ducts (which transport bile acids from the liver to the small intestine). While cholangiocarcinoma is far from the most prevalent cancer that people get, its incidence is increasing, and its prognosis is dismal—we’re talking a five-year survival rate of roughly 20%, which drops to 2% for metastatic disease.
“Rare cancers are not only difficult to study from a financial perspective,” notes Dr. Sita Kugel, whose lab in the Human Biology Division at Fred Hutch studies this cancer type, “but also from a scientific perspective—clinical data and patient samples are sparse, and the model systems which are so crucial to understanding the disease are often underdeveloped or completely absent.” For the longest time, these problems hampered research progress on one particular subtype of cholangiocarcinoma called intrahepatic cholangiocarcinoma (ICC); however, the tide began to turn thanks in no small part to Dr. Supriya ‘Shoop’ Saha, a former principal investigator and clinician in the Human Biology Division who established one of the first labs in the world dedicated to the study of cholangiocarcinoma. One of Dr. Saha’s focuses was a specific subtype of ICC characterized by mutations in the enzyme isocitrate dehydrogenase (IDH1/2), which plays a crucial role in central carbon metabolism. When mutated in cancer, IDH changes its normal enzymatic activity to instead produce an ‘oncometabolite’ called 2-hydroxyglutarate (2-HG), which accumulates in cells and promotes tumorigenesis via multiple emerging mechanisms.
During his postdoctoral studies, Dr. Saha not only created the first mouse models of IDH-mutant ICC—he also used high-throughput drug screens and cutting-edge gene editing techniques to identify a drug—dasatinib—that selectively killed IDH-mutant ICC cells by inhibiting a kinase called SRC (‘Sarc’). Tragically, Dr. Saha passed away before his lab could figure out how exactly dasatinib targeted these cancer cells; in his absence, a team including Dr. Kugel, postdoctoral scholar Dr. Iris Luk, and technician Caroline Bridgwater are leading the charge to understand how exactly this precision therapy works. Reporting their results in Science Translational Medicine, the team are well on their way to putting the puzzle pieces together.
To understand how dasatinib kills IDH-mutant ICC tumors while sparing their IDH-wild type counterparts, Luk and colleagues treated a battery of ICC cell lines with dasatinib and looked for changes in pathways downstream of SRC (the kinase which dasatinib inhibits). Sure enough, dasatinib abolished phosphorylation of one of these pathways—a protein called S6—and its partner kinase, fittingly called S6 kinase, or S6K. As Dr. Kugel notes, “S6/S6K are well known targets of the mTOR pathway—a master growth regulatory pathway—so we were convinced for a long time that dasatinib was inhibiting S6/S6K phosphorylation via the mTOR pathway.” Surprisingly, however, the team found the opposite; while dasatinib reduced S6/S6K phosphorylation (which in turn inhibited protein synthesis and caused cell death), it did so without the involvement of the mTOR pathway.
So, if dasatinib ‘turned off’ S6/S6K without using mTOR, what was the missing link between SRC and S6/S6K? As it turns out, the simplest assumption—that SRC itself phosphorylates S6/S6K—was incorrect, since SRC is unable to phosphorylate the specific amino acids which are regulated on S6/S6K. Instead, by employing high throughput phosphoproteomics, Dr. Luk and team found their culprit: a mysterious protein called MAGI1, which they showed is a SRC target and required for dasatinib to alter S6/S6K phosphorylation. So, SRC phosphorylates MAGI1, which phosphorylates S6/S6K—mission accomplished, right? Not so fast. “This finding was very striking but also threw us for a bit of a loop,” noted Luk, “because MAGI1 is just a scaffold protein—it has no intrinsic kinase or phosphatase activity, so it couldn’t be the one actually modulating S6/S6K phosphorylation.” After another series of intricate, hypothesis-driven experiments, the team found the final missing puzzle piece: a phosphatase called PP2A, which interacts with SRC and MAGI1 and de-phosphorylates S6K. Finally, the team had a more-or-less complete model: under normal conditions, SRC phosphorylates MAGI1, which prevents it from playing matchmaker with PP2A and S6K; inhibiting SRC with dasatinib gets rid of this phosphorylation on MAGI1 and allows PP2A to bind, whereupon is ‘turns off’ S6K, eventually killing the cells (see below).
If we know that dasatinib works to kill IDH-mutant ICC cells, why should we bother figuring out the reasons why it does this? “Because if we know the mechanism of action of dasatinib, we can design strategies to increase this effect and circumvent resistance,” replied Dr. Kugel. To cap off an already impressive study, the team did just that. After finding that ICC cells could become resistant to dasatinib by simply increasing their baseline levels of phosphorylated-S6K (the ultimate target of the entire dasatinib-induced signaling cascade, as noted above), Luk and team tested a combination therapy which consisted of dasatinib and M2698, an S6K inhibitor. Strikingly, the dasatinib/M2698 combination therapy reduced tumor growth and improved survival of a patient-derived xenograft model of IDH-mutant ICC compared to dasatinib treatment alone. As Dr. Luk noted, “Many precision therapies for these cancer types are cytostatic (i.e. they prevent the tumors from growing any further), but we were surprised to see that dasatinib/M2698 actually shrunk the tumors, which neither therapy was able to do on its own.”
From a new molecular mechanism controlling S6K phosphorylation and protein synthesis independently of the mTOR pathway, to a mechanism of action for dasatinib in IDH-mutant ICC and a rationally designed combination therapy for this rare and aggressive cancer type, this tour de force truly has something for everyone. Where is the team going next? “If you look closely,” noted Dr. Kugel, “you’ll notice that our model doesn’t really account for why dasatinib would have all of these effects in IDH-mutant but not IDH-wild type ICC cells.” Thus, the team’s ongoing work is focused on figuring out IDH’s role in this complex web of molecular interactions, as well as bringing their new combination therapy ‘from the bench to the bedside’ of ICC patients whose gruesome disease has been overlooked, until now. Shoop would be proud.
The spotlighted work was funded by the National Institutes of Health, the Evening for Maria Fund, a Cholangiocarcinoma Foundation Postdoctoral Fellowship, and the Spanish Ministry of Science and Innovation. It was supported by the Flow Cytometry, Preclinical Modeling, Comparative Medicine, Experimental Histopathology, and Proteomics/Metabolomics Shared Resources at Fred Hutch.
Fred Hutch/University of Washington/Seattle Children’s Cancer Consortium members Drs. Lucas Sullivan, Paul Lampe, Jonathan Cooper, Raymond Yeung, and Sita Kugel contributed to this study.
Luk, I. S.*, Bridgwater, C. M.*, Yu, A., Boila, L. D., Yáñez-Bartolomé, M., Lampano, A. E., Hulahan, T. S., Boukhali, M., Kathiresan, M., Macarulla, T., Kenerson, H. L., Yamamoto, N., Sokolov, D., Engstrom, I. A., Sullivan, L. B., Lampe, P. D., Cooper, J. A., Yeung, R. S., Tian, T. V., … Kugel, S. (2024). SRC inhibition enables formation of a growth suppressive MAGI1-PP2A complex in isocitrate dehydrogenase-mutant cholangiocarcinoma. Science Translational Medicine, 16(747), eadj7685.
*these authors contributed equally