Finding a rare bile duct cancer’s weaknesses

Fred Hutch Cancer Center postdoctoral fellow Liberalis (Debraj) Boila, PhD, has received two awards to support his studies of an aggressive bile duct cancer called intrahepatic cholangiocarcinoma, or ICC. Funding from the Department of Defense Rare Cancers Research Program and the Cholangiocarcinoma Foundation (CCF) will allow Boila, mentored by Fred Hutch pancreatic and liver cancer researcher Sita Kugel, PhD, to study how two of the most common ICC mutations drive cancer and contribute to each subtype’s unique treatment vulnerabilities.

“Standard first-line treatment for ICC hasn’t changed since the last decade. Patients need more targeted therapies,” Boila said.

His two-year, $100,000 DOD Rare Cancers Program Concept Award will enable Boila to study a subtype of ICC that currently has no targeted treatment. The DOD Rare Cancers Research Program was developed to address the burden of rare cancers in military personnel and veterans. About 16% of the cancers affecting military vets are rare cancers, including ICC. A deeper understanding of this subtype’s genetic underpinnings and potential drug susceptibilities will lay the groundwork, Boila hopes, for clinical trials of new, more tailored treatments for ICC.

Boila’s one-year, $50,000 Linda Blum Memorial Research Fellowship from the CCF will allow him to discover what underlies a different ICC subtype’s drug sensitivities. These findings could help scientists develop strategies to stave off drug resistance or more effective combination therapies. The CCF provides seed funding to creative early-career investigators whose promising projects are likely to have significant clinical impact.

ICC: Rare but aggressive

Cholangiocarcinoma, or CCA, is cancer of the bile ducts. ICC is intra-hepatic or intra-liver CCA. It arises in the bile ducts within the liver and makes up about 10-20% of all cases of CCA. While the overall incidence of CCA is quite rare — no more than six people per 100,000 in most countries — the disease comes with a heavy cost. Globally, about 2% of cancer-related deaths every year can be attributed to CCA, which predominantly occurs in older people. Vietnam veterans, who have a median age of 71 years and are the U.S.’ largest veteran cohort, have an abnormally high incidence of CCA.

“If ICC is diagnosed early, the prognosis is much better,” Boila said. “But that almost never happens.”

Because ICC often produces no symptoms, it is usually diagnosed at a late stage when surgery is no longer an option. According to the American Cancer Society, only 9% as many patients with intra-liver bile duct cancer will survive five years, compared to the percentage of people without the same disease who will survive five years. This relative survival rate is 23% for patients whose ICC is diagnosed early, but can be as low as 3% if the tumor is diagnosed after it has spread through the body.

Only about half of patients diagnosed with ICC will live a year past treatment. More effective and targeted treatments are desperately needed, Boila said.

Using innovative preclinical models to understand ICC

Liver cancer incidence, including ICC, is rising, but ICC’s rarity makes it hard to study. With so few patients, it’s difficult to gather enough tissue samples to make confident inferences about what’s driving the cancer and where it may be vulnerable.

“It’s true of most rare cancers: small cohorts make it difficult to get statistically significant observations from patient data. It’s why we depend more on experimental models,” Boila said.

He has several novel preclinical models he can use to better understand ICC. The late Supriya “Shoop” Saha, MD, PhD, a Fred Hutch ICC expert, developed genetically engineered mouse models (or GEMMs) that grow ICC tumors in their bile ducts. These include GEMMs that model the top two mutated genes seen in ICC. One GEMM has a liver-specific cancer-associated mutation in the gene encoding the enzyme isocitrate dehydrogenase, or IDH. The other is a completely novel GEMM strain with a liver-specific knockout of a gene called ARID1A.

After Saha passed away, Kugel, his wife, took over his lab and projects, continuing his work and legacy. She now leads a combined lab focused on both liver and pancreatic cancer.

Discovering how ARID1A mutations cause cancer

About 20% of human ICC cases have a mutation in the ARID1A gene. ARID1A is part of a protein complex that restructures DNA packaging to influence expression of certain genes. Its loss may shift which genes are turned on or off in a way that promotes tumor formation. Boila’s DOD funding will allow him to begin untangling this mechanism.

“Using this ARID1A-knockout model, we’ll be able to gather data showing the driving factors promoting ICC in ARID1A-mutated bile duct cells,” Boila said. “We’re trying to find the mechanisms that regulate tumor formation in this ICC subtype.”

In addition to the new mouse model, he’ll use cell lines and organoids — which recapture in a lab dish some degree of a tumor's 3D nature — to reveal how ARID1A mutations promote ICC development.

Boila’s suite of preclinical models will also aid his efforts to discover potential new therapies for patients with ARID1A-mutated ICC. Currently the only targeted treatments for patients with ICC are aimed at tumors with mutations in IDH or a growth factor gene, FGFR.

To give patients more options, Boila will use his DOD funding to support the hunt for potential new drugs. He will screen a compendium of FDA-approved drugs for efficacy against ICC with mutated ARID1A, testing them first in his cell line and organoid models. Those that seem most promising will be tested in “preclinical” trials run in patient-derived xenograft (PDX) models, in which human tumor tissue is implanted into mice. PDX models can give scientists a better idea of how human tumors may respond to a potential treatment than a genetically manipulated mouse tumor would.

Exploring the drug sensitivities of IDH1-mutant ICC

Boila’s CCF award will allow him to better understand another ICC subtype, driven by mutations in a gene involved in metabolism. This gene encodes the enzyme isocitrate dehydrogenase, or IDH, which creates an important metabolite in our cells’ energy-generating cycle. Unlike cancer-promoting mutations in ARID1A, which deactivate its protein, cancer-promoting mutations in the IDH1 gene give the IDH enzyme a new function.

Usually, IDH breaks down a molecule called isocitrate, turning it into CO₂ and α-ketoglutarate. Alpha-ketoglutarate is an important co-factor in a wide array of enzymatic reactions within our cells. Mutant IDH gains the ability to create a new, toxic metabolite: D-2-hydroxyglutarate, or D-2HG. High concentrations of D-2HG and low levels of α-ketoglutarate alter the activity of many enzymes that regulate gene expression, shifting the pattern of genes that are turned on and off.

Tumor cells tend to be less “differentiated” than normal cells, which means they can’t perform the specialized functions that are unique to various cell types. The gene expression shifts in IDH1-mutant cells help promote cancer by blocking the liver stem cells’ ability to differentiate into functional liver cells.

Boila's CCF funding will support his studies of an enzyme called PP2A, which regulates sensitivity to chemotherapies called nucleoside analogs. These anti-cancer drugs damage DNA by mimicking its components. Standard treatments for many types of cancer, nucleoside analogs include drugs like gemcitabine, cytarabine, cladribine and clofarabine.

Whether IDH mutation status influences PP2A’s ability to promote sensitivity to gemcitabine is unknown. Kugel’s team revealed that a new signaling complex, of which PP2A is a part, removes activating modifications from growth-promoting proteins. Boila will work to better understand the critical role that PP2A plays in IDH1-mutant ICC biology, and how modifying PP2A function may influence tumor cell growth, survival and drug sensitivities.

Boila recently attended and presented their latest findings at the CCF annual conference. It was an opportunity to connect with pioneers in the CCA research community as well as patients and patient advocates. He was able to interact with a global community of scientists who presented clinical and research data from around the world, including regions where CCA is more prevalent than in the U.S.

He also learned about how regulatory bodies work to help make drugs more accessible to patients and spoke with interested patients and caregivers.

“It was a wonderful experience,” Boila said. “It reminds me how my work impacts patients’ lives. The experience was very enriching.”

Leading by example

Boila also commended Kugel and his other Fred Hutch mentors for their guidance and support.

“Sita has been an incredibly motivating and supportive mentor,” he said. “She leads by example.”

Having trained in leukemia research during his PhD, Boila wanted to utilize his expertise in cancer biology and passion for understanding how cells regulate modifications of DNA and DNA packaging in the study of rare cancers. Kugel’s expertise in gastrointestinal cancers and careful mentorship allowed Boila to dive into the intricacies of ICC research. And her guidance doesn’t end at the lab bench: Kugel has designed a dynamic training plan, which she and Boila regularly update, to ensure Boila achieves his long-term goal of leading his own research group, he said.

Kugel has also modeled the perseverance and resilience needed to weather — and grow from — scientific setbacks, Boila said. He pointed to her recent McDougall Mentoring Award as further testament to her outstanding support for trainees.

Fred Hutch prostate and bladder cancer physician-scientist Andrew Hsieh, MD, co-mentored Boila for his DOD award. Fred Hutch’s Jonathan Cooper, PhD, an expert in the critical enzyme inhibited by dasatanib, acted as co-mentor for Boila’s CCF grant.

“Debraj is a fantastic scientist — intellectually curious and rigorous,” Kugel said. “I am so happy and proud to support him for this project and into the future as he grows into an independent scientist and establishes his own lab.”