Research Area: Cancer Biology

Discovery of how cancer drugs find their targets could lead to a new toolset for drug development
Whitehead Institute
June 17, 2020

In the watery inside of a cell, complex processes take place in tiny functional compartments called organelles. Energy-producing mitochondria are organelles, as is the frilly golgi apparatus, which helps to transport cellular materials. Both of these compartments are bound by thin membranes.

But in the past few years, research at Whitehead Institute and elsewhere has shown that there are other cellular organelles held together without a membrane. These organelles, called condensates, are tiny droplets which keep certain proteins close together amidst the chaos of the cell, allowing complex functions to take place within. “We know of about 20 types of condensate in the cell so far,” says Isaac Klein, a postdoc in Richard Young’s lab at Whitehead Institute and oncologist at the Dana-Farber Cancer Institute.

Now, in a paper published in Science on June 19, Klein and Ann Boija, another postdoc in Young’s lab, show the mechanism by which small molecules, including cancer drugs, are concentrated in these cellular droplets — a finding that could have implications for the development of new cancer therapeutics. If researchers could tailor a chemical to seek out and concentrate in one kind of droplet in particular, it might have a positive effect on the delivery efficiency of the drug. “We thought, maybe that’s an avenue by which we can improve cancer treatments and discover new ones,” says Klein.

“This [research] is part of a revolutionary new way of looking at the organization within cells,” says Phillip Sharp, a professor at the Massachusetts Institute of Technology’s Koch Institute for Integrative Cancer Research and a co-author on the study. “Cells are not little pools of soup, all mixed together. They are actually highly organized, compartmentalized units, and that organization is important in their function and in their diseases. We’ve just started to understand that, and this new paper is a really important step, using that insight, to understand how to potentially treat diseases differently.”

CONDENSATES AND DRUG DELIVERY

To explore how different properties of condensates inside the cell’s nucleus affected the delivery of cancer drugs, Boija and Klein selected a few example condensates to study. These included splicing speckles, which store cellular materials needed for RNA splicing, nucleoli, where ribosomes are formed, and a new kind of droplet Young’s lab discovered in 2018 called a transcriptional condensate. These new condensates bring together all the different proteins needed to successfully transcribe a gene.

The researchers created their own suite of four different fluorescently-labeled condensates by adding glowing tags to marker proteins specific to each kind of droplet. For example, transcriptional condensates are marked by the droplet-forming protein MED1, splicing speckles by a protein called SRSF2, and nucleoli by FIB1 and NPM1.

Now that they could tell individual droplets apart by their cellular purpose, the team, along with the help of Nathanael Gray, a chemical biologist at Harvard University and the Dana-Farber Cancer Institute, created fluorescent versions of clinically important drugs. The tested drugs included cisplatin and mitoxantrone, two anti-tumor medicines commonly used in chemotherapy. These therapeutics were the perfect test subjects, because they both target proteins that lie within nuclear condensates.

The researchers added the cancer drugs to a mixture containing various droplets (and only droplets, none of the actual drug targets), and found that the drugs sorted themselves into specific condensates. Mitoxantrone concentrated in condensates marked by MED1, FIB1 and NPM1, selectively avoiding the others. Cisplatin, too, showed a particular affinity for droplets held together by MED1.

“The big discovery with these in vitro studies is that a drug can concentrate within transcriptional condensate independent of its target,” Boija says. “We used to think that drugs come to the right place because their targets are there, but in our in vitro system, the target is not there. That’s really informative — it shows the drug is actually being concentrated in a different way than we thought.”

To understand why some drugs were drawn into transcriptional condensates, they screened a panel of chemically-modified dyes and found that the important part of many drugs — the part that led them to concentrate in transcriptional condensates  —  is the molecules’ aromatic ring structure. Aromatic rings are stable, ring-shaped groupings of carbon atoms. The aromatic ring in some drugs are thought to stack with rings in MED1’s amino acids, leading the drug to concentrate in transcriptional condensates.

Being able to tailor a drug to enter a certain condensate is a powerful tool for drug developers. “We found that if we add an aromatic group to a molecule, it becomes concentrated within the transcriptional condensate,” Boija says. “It’s that type of interaction that is important when we design new drugs to enter transcriptional condensates — and maybe we can improve existing drugs by modifying their structure. This will be very exciting to look into.”

WHERE DRUGS CONCENTRATE AFFECTS HOW WELL THEY FIGHT CANCER

In order for this tool to be practically useful in drug development, the researchers had to make sure that concentration in specific droplets would actually impact the drugs’ performance. Boija and Klein decided to test this using cisplatin, which is drawn to transcriptional condensates by MED1 and works to fight cancer by adding clunky platinum molecules to DNA strands. This damages tumor cells’ genetic material. When the researchers administered cisplatin to a mixture of different condensates, both in the test tube and in cells, the drug preferentially altered DNA that lay within transcriptional condensates.

This could explain why cisplatin and other platinum drugs are effective against so many diverse cancers, says Young, who is also a professor of biology at MIT; cancer-causing genes often carry regions of DNA called super enhancers, which are extremely active in transcription, leading to very large transcriptional condensates. “We now think the reason that drugs like cisplatin can work well in patients with diverse cancers is because they’re becoming selectively concentrated at the cancer-causing genes, where these large transcriptional condensates occur,” he said. “The effect is to have the drug home in on the gene that’s causing each cancer to be so deadly.”

A DRUG RESISTANCE MYSTERY, SOLVED

The new insights in condensate behavior also provided some answers to another question in cancer research: why people become immune to the breast cancer drug tamoxifen.Tamoxifen works by attaching itself to estrogen receptors in the cancer cells, preventing them from getting the hormones they need to grow and eventually slowing or stopping the formation of new cancer cells altogether. The drug is one of the most effective treatments for the disease, reducing recurrence rates for ER+ breast cancers by around 50%.

Unfortunately, many patients quickly develop a resistance to tamoxifen — sometimes as soon as a few months after they start taking it. This happens in a variety of ways — for example, sometimes the cancer cells will mutate to be able to kick the tamoxifen out of the cells, or simply produce fewer estrogen receptors for the drug to bind. One form of resistance was associated with an overproduction of the protein MED1, but scientists didn’t know why.

With their newfound knowledge of how a drug’s activity is affected by where it concentrates, Boija and Klein had a hypothesis: the extra MED1 might increase the size of the droplets, effectively diluting the concentration of tamoxifen and making it more difficult for the drug to bind its targets. When they tested this in the laboratory, the team found that more MED1 did indeed cause larger droplets, leading to lower concentrations of tamoxifen.

A NEW TOOLSET FOR DRUG DESIGNERS

The ability to better understand the behavior of drugs in cancer cells — how they concentrate, and why the cancer could become resistant to them — may provide drug developers with a new arsenal of tools to craft efficient therapeutics. “This study suggests that we should be exploring whether we can design or isolate drugs that are concentrated in a given condensate, and to understand how existing drugs are concentrated in the cell,” says Phil Sharp. “I think this is really important for drug development — and I think [figuring it out] is going to be fun.”

Chimeras offer a new way to study childhood cancer in mice
Eva Frederick | Whitehead Institute
March 5, 2020

In a new paper published March 5 in the journal Cell Stem Cell, researchers in Whitehead Institute Member Rudolf Jaenisch’s lab introduce a new way to model human neuroblastoma tumors in mice using chimeras — in this case, mice that have been modified to have human cells in parts of their nervous systems. “This may serve as a unique model that you can use to study the dynamic of immune cells within human tumors,” says Malkiel Cohen, a postdoc in Jaenisch’s lab and the first author of the paper.

Neuroblastoma is a rare and unpredictable form of childhood cancer that affects around 800 young children in the US each year. Neuroblastoma tumors often occur in parts of the sympathetic nervous system, which includes the nerves that run parallel to the spinal cord and the adrenal medulla, part of the glands that produce hormones such as adrenaline. Neuroblastoma is notoriously hard to study primarily because of its disparate behavior: the tumors often shrink spontaneously in infants, while in toddlers they are highly aggressive and often fatal. “The seeds for the cancer are sown during fetal life,” says Rani George, MD, PhD, an associate professor of pediatrics at Harvard Medical School and a neuroblastoma researcher and physician at Dana-Farber Cancer Institute and Boston Children’s Hospital, and a co-senior author on the paper. “For obvious reasons, you can’t really study the development of these tumors in humans.”

Until now, researchers didn’t have many realistic ways to study these tumors in animal models, either. They could create transgenic mice with cancer-causing genes, but the resulting tumors were mouse tumors, not human ones, and had some key differences. Another method involved taking human tumor cells and implanting them in a mouse — a process called xenotransplantation — but that only worked in mice with compromised immune systems, and didn’t allow researchers to study how the tumors formed in the first place or how they interacted with a fully functioning immune system. “This is where we think the new model is a perfect fit,” said Stefani Spranger, PhD, an assistant professor of Biology at the Massachusetts Institute of Technology (MIT) and the Koch Institute for Integrative Cancer Research at MIT and a co-senior author on the paper.

Human-mouse chimeras have been used in the past to study Alzheimer’s disease and brain development. Jaenisch, who is also a professor of biology at MIT, and his lab had been working for years to create chimeric mice with human cells in the neural crest — the group of developing cells that go on to form parts of the sympathetic nervous system — and published their findings in 2016. “In this study, we hoped to use these mice with human neural crest cells to study how neuroblastoma tumors form and respond to immune system attacks,” Jaenisch says.

To create these chimeric mice, Cohen and coauthors at MIT’s Koch Institute and the Dana-Farber Cancer Institute first engineered human pluripotent stem cells to express two genes known to be abnormal in neuroblastoma, MYCN and mutated ALK, and modified them so they became neural crest cells, from which human neuroblastomas are derived. The genes could be turned on and off with the addition of doxycycline, an antibiotic. They also inserted the gene for eGFP, a brightly glowing fluorescent protein originally isolated from jellyfish. This would allow the team to tell whether the cells were spreading correctly through the bodies of the mice, and would cause any tumors originating from these human cells to be luminous under fluorescent light.

The researchers injected mouse embryos with these cells, and watched over the course of embryonic development as the cells proliferated and human tissues crept into the developing peripheral nervous systems of the tiny mice. To activate the two cancer-causing genes, researchers spiked the pregnant mice’ water with doxycycline, and over the next few days in utero — and in the weeks and months after the pups were born — the researchers inspected the chimeras to see whether tumors would appear.

Over the course of the next 15 months, 14% of the mice developed tumors — 29 mice out of 198 total. The tumors mostly appeared in the space behind the abdominal cavity close to the nerves along the spinal cord, although one mouse developed a tumor in its adrenal gland. Both locations are common places for human children to develop neuroblastoma. The researchers took samples of the tumors and found that they contained the glowing protein eGFP, which confirmed that they were of human origin.

When the team examined the growth patterns of the cancerous cells, they found that the tumors were remarkably similar to human neuroblastomas: they contained cell markers typical of human tumors, and some grew in characteristic rosette shapes — features that did not often appear in tumors implanted in immunocompromised mice through xenotransplantation.

Having successfully induced neuroblastoma tumors in the chimeric mice, the researchers took the opportunity to examine the communication between immune cells and tumors — and specifically, how the tumors evaded destruction by anti-cancer immune cells called T cells. One factor that makes human neuroblastomas and many other cancers dangerous is their sophisticated strategy for avoiding being destroyed by T cells. “The cancer tricks the immune system,” Cohen says.  By activating chemical signals that exhaust the T cells, the tumors effectively weaken their attack. The tumors in the chimeric mice, Cohen found, use a similar method to human neuroblastomas to evade immune responses.

Cohen and others plan to test the new system’s potential for modeling other cancers such as melanoma, and to use it to investigate potential treatments for neuroblastoma patients. “The obvious next step is to study how treatment of these tumors will allow these chimeric mice to be cured,” he says. “This is a model that will allow us to approach not only how to get rid of the tumor, but also to fix the immune system and recover those exhausted T cells, allowing them to fight back and deplete the tumor.”

This research was funded by the National Institutes of Health, as well as grants from the Emerald Foundation, the LEO Foundation, the Melanoma Research Foundation, and the St. Baldrick’s Foundation.

Citation: Cohen, M., et al. Formation of Human Neuroblastoma in Mouse-Human Neural Crest Chimeras. Cell Stem Cell. March 5, 2020. DOI: https://doi.org/10.1016/j.stem.2020.02.001

***

Written by Eva Frederick

***

Singing for joy and service

After surgery to correct childhood hearing loss, Swarna Jeewajee discovered a a desire to be a physician-scientist, and a love of a cappella music.

Shafaq Patel | MIT News correspondent
February 3, 2020

Swarna Jeewajee grew up loving music — she sings in the shower and blasts music that transports her to a happy state. But until this past year, she never felt confident singing outside her bedroom.

Now, the senior chemistry and biology major spends her Saturdays singing around the greater Boston area, at hospitals, homes for the elderly, and rehabilitation centers, with the a cappella group she co-founded, Singing For Service.

Jeewajee says she would not have been able to sing in front of people without the newfound confidence that came after she had transformative ear surgery in the spring of 2018.

Jeewajee grew up in Mauritius, a small island off the east coast of Madagascar, where she loved the water and going swimming. When she was around 8 years old, she developed chronic ear infections as a result of a cholesteatoma, which caused abnormal skin growth in her middle ear.

It took five years and three surgeries for the doctors in Mauritius to diagnose what had happened to Jeewajee’s ear. She spent some of her formative years at the hospital instead of leading a normal childhood and swimming at the beach.

By the time Jeewajee was properly diagnosed and treated, she was told her hearing could not be salvaged, and she had to wear a hearing aid.

“I sort of just accepted that this was my reality,” she says. “People used to ask me what the hearing aid was like — it was like hearing from headphones. It felt unnatural. But it wasn’t super hard to get used to it. I had to adapt to it.”

Eventually, the hearing aid became a part of Jeewajee, and she thought everything was fine. During her first year at MIT, she joined Concourse, a first-year learning community which offers smaller classes to fulfill MIT’s General Institute Requirements, but during her sophomore year, she enrolled in larger lecture classes. She found that she wasn’t able to hear as well, and it was a problem.

“When I was in high school, I didn’t look at my hearing disability as a disadvantage. But coming here and being in bigger lectures, I had to acknowledge that I was missing out on information,” Jeewajee says.

Over the winter break of her sophomore year, her mother, who had been living in the U.S. while Jeewajee was raised by her grandmother in Mauritius, convinced Jeewajee to see a specialist at Massachusetts Eye and Ear Hospital. That’s when Jeewajee encountered her role model, Felipe Santos, a surgeon who specializes in her hearing disorder.

Jeewajee had sought Santos’ help to find a higher-performing hearing aid, but instead he recommended a titanium implant to restore her hearing via a minimally invasive surgery. Now, Jeewajee does not require a hearing aid at all, and she can hear equally well from both ears.

“The surgery helped me with everything. I used to not be able to balance, and now I am better at that. I had no idea that my hearing affected that,” she says.

These changes, she says, are little things. But it’s the little things that made a large impact.

“I gained a lot more confidence after the surgery. In class, I was more comfortable raising my hand. Overall, I felt like I was living better,” she says.

This feeling is what brought Jeewajee to audition for the a cappella group. She never had any formal training in singing, but in January, during MIT’s Independent Activities Period, her friend mentioned that she wanted to start an a cappella group and convinced Jeewajee to help her launch Singing For Service.

Jeewajee describes Singing For Service as her “fun activity” at MIT, where she can just let loose. She is a soprano singer, and the group of nine to 12 students practices for about three hours a week before their weekly performances. They prepare three songs for each show; a typical lineup is a Disney melody, Josh Groban’s “You Raise Me Up,” and a mashup from the movie “The Greatest Showman.”

Her favorite part is when they take song requests from the audience. For example, Singing For Service recently went to a home for patients with multiple sclerosis, who requested songs from the Beatles and “Bohemian Rhapsody.” After the performance, the group mingles with the audience, which is one of Jeewajee’s favorite parts of the day.

She loves talking with patients and the elderly. Because Jeewajee was a patient for so many years growing up, she now wants to help people who are going through that type of experience. That is why she is going into the medical field and strives to earn an MD-PhD.

“When I was younger, I kind of always was at the doctor’s office. Doctors want to help you and give you a treatment and make you feel better. This aspect of medicine has always fascinated me, how someone is literally dedicating their time to helping you. They don’t know you, they’re not family, but they’re here for you. And I want to be there for someone as well,” she says.

Jeewajee says that because she grew up with a medical condition that was poorly understood, she wants to devote her career to search for answers to tough medical problems. Perhaps not surprisingly, she has gravitated toward cancer research.

She discovered her passion for this field after her first year at MIT, when she spent the summer conducting research in a cancer hospital in Lyon, through MISTI-France. There, she experienced an “epiphany” as she watched scientists and physicians come together to fight cancer, and was inspired to do the same.

She cites the hospital’s motto, “Chercher et soigner jusqu’à la guérison,” which means “Research and treat until the cure,” as an expression of what she will aspire to as a physician-scientist.

Last summer, while working at The Rockefeller University investigating mechanisms of resistance to cancer therapy, she developed a deeper appreciation for how individual patients can respond differently to a particular treatment, which is part of what makes cancer so hard to treat. Upon her return at MIT, she joined the Hemann lab at the Koch Institute for Integrative Cancer Research, where she conducts research on near-haploid leukemia, a subtype of blood cancer. Her ultimate goal is to find a vulnerability that may be exploited to develop new treatments for these patients.

The Koch Institute has become her second home on MIT’s campus. She enjoys the company of her labmates, who she says are good mentors and equally passionate about science. The walls of the lab are adorned with science-related memes and cartoons, and amusing photos of the team’s scientific adventures.

Jeewajee says her work at the Koch Institute has reaffirmed her motivation to pursue a career combining science and medicine.

“I want to be working on something that is challenging so that I can truly make a difference. Even if I am working with patients for whom we may or may not have the right treatment, I want to have the capacity to be there for them and help them understand and navigate the situation, like doctors did for me growing up,” Jeewajee says.

New pathway for lung cancer treatment

MIT researchers identify pyrimidine biosynthesis as a target for the treatment of small cell lung cancer.

Bendta Schroeder | Koch Institute
November 11, 2019

MIT cancer biologists have identified a new therapeutic target for small cell lung cancer, an especially aggressive form of lung cancer with limited options for treatment.

Lung cancer is the leading cause of cancer-associated mortality in the United States and worldwide, with a five-year survival rate of less than 20 percent. But of the two major sub-types of lung cancer, small cell and non-small cell, small cell is more aggressive and has a much poorer prognosis. Small cell lung cancer tumors grow quickly and metastasize early, resulting in a five-year survival rate of about 6 percent.

“Unfortunately, we haven’t seen the same kinds of new treatments for small cell lung cancer as we have for other lung tumors,” says Tyler Jacks, director of the Koch Institute for Integrative Cancer Research at MIT. “In fact, patients are treated today more or less the same way they were treated 40 or 50 years ago, so clearly there is a great need for the development of new treatments.”

A study appearing in the Nov. 6 issue of Science Translational Medicine shows that small cell lung cancer cells are especially reliant on the pyrimidine biosynthesis pathway and that an enzyme inhibitor called brequinar is effective against the disease in cell lines and mouse models.

Jacks is the senior author of this study. Other MIT researchers include Associate Professor of Biology and Koch Institute member Matthew Vander Heiden, and co-lead authors postdoc researcher Leanne Li and graduate student Sheng Rong Ng.

Roadblock for cell replication

Researchers in the Jacks lab used CRISPR to screen small cell lung cancer cell lines for genes that already have drugs targeting them, or that are likely to be druggable, in order to find therapeutic targets that can be tested more quickly and easily in a clinical setting.

The group found that small cell lung cancer tumors are particularly sensitive to the loss of a gene encoding dihydroorotate dehydrogenase (DHODH), a key enzyme in the de novo pyrimidine biosynthesis pathway. Upon discovering that the sensitivity involved a metabolic pathway, the researchers sought the collaboration of the Vander Heiden lab, experts in normal and cancer cell metabolism who were already conducting studies on the role of pyrimidine metabolism and DHODH inhibitors in other cancers.

Pyrimidine is one of the major building blocks of DNA and RNA. Unlike healthy cells, cancer cells are constantly dividing and need to synthesize new DNA and RNA to support the production of new cells. The investigators found that small cell lung cancer cells have an unexpected vulnerability: Despite their dependence on the availability of pyrimidine, this synthesis pathway is much less active in small cell lung cancer cells than in other types of cancer cells examined in the study. Through inhibiting DHODH, they found that small cell lung cancer cells were not able to produce enough pyrimidine to keep up with demand.

When researchers treated a genetically engineered mouse model of small cell lung cancer tumors with the DHODH inhibitor brequinar, tumor progression slowed down and the mice survived longer than untreated mice. Similar results were observed for small cell lung cancer tumors in the liver, a frequent site of metastasis in patients.

In addition to mouse model studies, the researchers tested four patient-derived small cell lung cancer tumor models and found that brequinar worked well for two of these models — one of which does not respond to the standard platinum-etoposide regimen for this disease.

“These findings are noteworthy because second-line treatment options are very limited for patients whose cancers no longer respond to the initial treatment, and we think that this could potentially represent a new option for these patients,” says Ng.

Shorter pathway to the clinic

Brequinar has already been approved for use in patients as an immunosuppressant, and there has been some preclinical research showing that brequinar and other DHODH inhibitors may be effective for other types of cancers.

“We’re excited because our findings could provide a new way to help small cell lung cancer patients in the future,” says Li. “While we still have a lot of work to do before brequinar can be tested in the clinic as a therapy for small cell lung cancer, we’re hopeful that this might happen more quickly now that we’re starting with a drug that is known to be safe in humans.”

Next steps for the researchers include optimizing the therapeutic efficacy of DHODH inhibitors and combining them with other currently available treatment options for small cell lung cancer, such as chemotherapy and immunotherapy. To help clinicians tailor treatments to individual patients, researchers will also work to identify biomarkers for tumors that are susceptible to this therapy, and investigate resistance mechanisms in tumors that do not respond to this treatment.

The research was funded, in part, by the MIT Center for Precision Cancer Medicine and the Ludwig Center for Molecular Oncology at MIT.

Researchers discover new source of drug resistance in pancreatic cancer
Lucy Jackub
October 17, 2019

The best available treatments for pancreatic cancer are highly toxic, and, as chemotherapies go, not very effective. The drug gemcitabine has been used for decades to extend the life of patients, but very high doses are required to combat the tumor, which grows in the pancreas surrounded by stiff, fibrous, noncancerous tissue called stroma. This hallmark of pancreatic cancer makes it unusually difficult to treat: the more stromal tissue accumulates, the less the drug works, while patients still endure brutal side effects. Only 8.5 percent of pancreatic cancer patients survive five years beyond their diagnosis, so there’s an urgent need to figure out why existing treatments are failing.

Scientists have known for a long time that gemcitabine fights cancer by killing cells during replication, though why it works for pancreatic cancer in particular is a bit of a mystery. The drug is a small molecule that masquerades as the nucleoside deoxycytidine, one unit in the nucleic acids that make up DNA. Once gemcitabine is integrated into a replicating strand of DNA, additional nucleosides can’t be joined to it. The new DNA strand can’t be completed, and the cell dies. Now, researchers from MIT have discovered that non-cancer cells in the pancreatic stromal tissue secrete astonishing quantities of deoxycytidine. They found that competition with deoxycytidine makes its imposter, gemcitabine, less effective, explaining why higher doses of the drug are needed as more stromal tissue grows around the tumor.

“That was an answer we were looking for — what is making pancreatic tumors resistant to gemcitabine?” says Michael Hemann, associate professor of biology, a member of MIT’s Koch Institute for Integrative Cancer Research, and co-senior author of the study. “Understanding the basic mechanisms of these drugs allows us to return to the clinic with improved strategies to treat patients with cancer.”

Douglas Lauffenburger, a professor of biological engineering, is also a co-senior author of the study, which represents a collaboration between the Hemann lab, the Lauffenburger lab, and the Vander Heiden lab, and appeared online in Cancer Research on September 4. Hemann lab graduate student Simona Dalin is the lead author.

The mystery ingredient

For years, researchers at MIT have been investigating different sources of chemotherapy resistance in stromal tissue. When Dalin took up the study two years ago, she was building on the findings of a former postdoc in the Hemann lab, Emanuel Kreidl. Kreidl had found that stellate cells, one type of cell in the pancreatic stromal tissue surrounding the tumor, were releasing something into the microenvironment of the pancreas that disrupted the function of gemcitabine.

Cells secrete all sorts of things — micro RNAs, fatty acids, proteins — that may be taken up and used by neighboring cells. Biologists call these ambient materials around the cell its “media.”  Kreidl had tried boiling, digesting, and filtering the stellate cell media, but nothing he did made gemcitabine any more effective against the cancer cells. The usual suspects commonly implicated in drug resistance caused by neighboring cells, like proteins, would break down under such tests. “That’s when we knew there was something new here,” says Dalin. Her challenge was to figure out what that mystery ingredient was.

Mark Sullivan PhD ‘19, then a graduate student and biochemist in Vander Heiden lab, was enlisted to help separate the stellate cell media into its molecular components and identify them. After doing so, Dalin says, “it was fairly obvious that deoxycytidine was the thing that we were looking for.” Because gemcitabine works by taking deoxycytidine’s place in DNA replication, it made sense that the presence of a lot of deoxycytidine could make it difficult for gemcitabine to fulfill its function.

Molecules pass in and out of cells through gates in the cell membrane, called transporters. Using a drug that blocks certain transporters, Dalin was able to shut the gate in the stellate cells through which deoxycytidine is released. With less deoxycytidine around, the gemcitabine was effective at lower doses, confirming her hypothesis. Now, the researchers just needed to figure out how and where deoxycytidine was getting in the way of the drug.

Once inside the cell, a nucleoside must have one or more phosphate groups added to it by several enzymes in order to become a nucleotide that can be used to build DNA. Gemcitabine goes through the same process. The researchers determined that gemcitabine was competing with deoxycytidine for the first of those enzymes, deoxycytidine kinase. When they flooded the cell with that enzyme, gemcitabine didn’t have to wait in line for its phosphate groups — and could get into the DNA to work its fatal subterfuge.

Upending Assumptions

Going forward, the Hemann lab aims to identify drugs that could inhibit the production of deoxycytidine and restore the tumor’s sensitivity to gemcitabine. Senthil Muthuswamy, an associate professor of medicine at Beth Israel Deaconess Medical Center who was not involved in the research, says this study provides “new and important insights” into how and why tumors develop resistance to gemcitabine. The findings, he adds, are “likely to have important implications for developing ways to overcome gemcitabine resistance in pancreatic cancer.”

The study’s findings may shed light on other cancer treatments that work similarly to gemcitabine. For every nucleoside, there are look-alike molecules, or analogs, that are used in cancer therapies. For example, the purine analog fludarabine is used to treat acute myeloid leukemia, another tenacious carcinoma. These generic drugs have been adopted through trial and error in the clinic, but scientists don’t fully understand why they are effective at the molecular level.

In theory, nucleoside analog drugs should work interchangeably; every nucleoside is necessary in either the replication of DNA or RNA. In practice, though, these drugs are only effective for certain cancers. The MIT researchers speculate that the sheer amount of deoxycytidine being produced in the pancreas could suggest that pancreatic cells have a particular need for deoxycytidine that also makes them more responsive to its analogs — perhaps explaining why gemcitabine targets pancreatic cancer cells effectively.

“Understanding more about nucleoside biology, and more about which organs have high levels of which nucleosides, might help us understand when to use which chemotherapies,” Dalin says.

This study leaves the researchers with many questions about how and why nucleosides are produced in the body, a realm of basic biology that is still poorly understood. It’s generally assumed that cells only make nucleosides for their own internal use in DNA replication. But pancreatic stellate cells produce a lot of deoxycytidine, far more than they need for themselves, suggesting the excess nucleosides may serve some unknown purpose in neighboring cells. Although more experiments are needed to determine this mysterious purpose, the MIT researchers have some ideas.

“These extra nucleosides introduce a possibility that perhaps making deoxycytidine is a normal function of stellate cells in the pancreas, in order to provide building blocks for the cells around them,” says Hemann. “And that’s a real surprise.”

This work was funded in part by a David H. Koch Fellowship and the MIT Center for Precision Cancer Medicine.

Image: Deoxycytidine and gemcitabine, its look-alike molecule, enter a cancer cell through the same gate in the cell membrane and are altered by the same enzyme (dCK) before they are integrated into DNA. Credit: Courtesy of the researchers.

Citation:
“Deoxycytidine Release from Pancreatic Stellate Cells Promotes Gemcitabine Resistance.”
Cancer Research, online Sept. 4, 2019, DOI: 10.1158/0008-5472.CAN-19-0960.
Dalin, S., Sullivan, M.R., Lau, A.N., Grauman-Boss, B., Mueller, H.S., Kreidl, E., Fenoglio, S., Luengo, A., Lees, J.A., Vander Heiden, M.G. and Lauffenburger, D.A.

Study links certain metabolites to stem cell function in the intestine

Molecules called ketone bodies may improve stem cells’ ability to regenerate new intestinal tissue.

Anne Trafton | MIT News Office
August 22, 2019

MIT biologists have discovered an unexpected effect of a ketogenic, or fat-rich, diet: They showed that high levels of ketone bodies, molecules produced by the breakdown of fat, help the intestine to maintain a large pool of adult stem cells, which are crucial for keeping the intestinal lining healthy.

The researchers also found that intestinal stem cells produce unusually high levels of ketone bodies even in the absence of a high-fat diet. These ketone bodies activate a well-known signaling pathway called Notch, which has previously been shown to help regulate stem cell differentiation.

“Ketone bodies are one of the first examples of how a metabolite instructs stem cell fate in the intestine,” says Omer Yilmaz, the Eisen and Chang Career Development Associate Professor of Biology and a member of MIT’s Koch Institute for Integrative Cancer Research. “These ketone bodies, which are normally thought to play a critical role in energy maintenance during times of nutritional stress, engage the Notch pathway to enhance stem cell function. Changes in ketone body levels in different nutritional states or diets enable stem cells to adapt to different physiologies.”

In a study of mice, the researchers found that a ketogenic diet gave intestinal stem cells a regenerative boost that made them better able to recover from damage to the intestinal lining, compared to the stem cells of mice on a regular diet.

Yilmaz is the senior author of the study, which appears in the Aug. 22 issue of Cell. MIT postdoc Chia-Wei Cheng is the paper’s lead author.

An unexpected role

Adult stem cells, which can differentiate into many different cell types, are found in tissues throughout the body. These stem cells are particularly important in the intestine because the intestinal lining is replaced every few days. Yilmaz’ lab has previously shown that fasting enhances stem cell function in aged mice, and that a high-fat diet can stimulate rapid growth of stem cell populations in the intestine.

In this study, the research team wanted to study the possible role of metabolism in the function of intestinal stem cells. By analyzing gene expression data, Cheng discovered that several enzymes involved in the production of ketone bodies are more abundant in intestinal stem cells than in other types of cells.

When a very high-fat diet is consumed, cells use these enzymes to break down fat into ketone bodies, which the body can use for fuel in the absence of carbohydrates. However, because these enzymes are so active in intestinal stem cells, these cells have unusually high ketone body levels even when a normal diet is consumed.

To their surprise, the researchers found that the ketones stimulate the Notch signaling pathway, which is known to be critical for regulating stem cell functions such as regenerating damaged tissue.

“Intestinal stem cells can generate ketone bodies by themselves, and use them to sustain their own stemness through fine-tuning a hardwired developmental pathway that controls cell lineage and fate,” Cheng says.

In mice, the researchers showed that a ketogenic diet enhanced this effect, and mice on such a diet were better able to regenerate new intestinal tissue. When the researchers fed the mice a high-sugar diet, they saw the opposite effect: Ketone production and stem cell function both declined.

Stem cell function

The study helps to answer some questions raised by Yilmaz’ previous work showing that both fasting and high-fat diets enhance intestinal stem cell function. The new findings suggest that stimulating ketogenesis through any kind of diet that limits carbohydrate intake helps promote stem cell proliferation.

“Ketone bodies become highly induced in the intestine during periods of food deprivation and play an important role in the process of preserving and enhancing stem cell activity,” Yilmaz says. “When food isn’t readily available, it might be that the intestine needs to preserve stem cell function so that when nutrients become replete, you have a pool of very active stem cells that can go on to repopulate the cells of the intestine.”

The findings suggest that a ketogenic diet, which would drive ketone body production in the intestine, might be helpful for repairing damage to the intestinal lining, which can occur in cancer patients receiving radiation or chemotherapy treatments, Yilmaz says.

The researchers now plan to study whether adult stem cells in other types of tissue use ketone bodies to regulate their function. Another key question is whether ketone-induced stem cell activity could be linked to cancer development, because there is evidence that some tumors in the intestines and other tissues arise from stem cells.

“If an intervention drives stem cell proliferation, a population of cells that serve as the origin of some tumors, could such an intervention possibly elevate cancer risk? That’s something we want to understand,” Yilmaz says. “What role do these ketone bodies play in the early steps of tumor formation, and can driving this pathway too much, either through diet or small molecule mimetics, impact cancer formation? We just don’t know the answer to those questions.”

The research was funded by the National Institutes of Health, a V Foundation V Scholar Award, a Sidney Kimmel Scholar Award, a Pew-Stewart Trust Scholar Award, the MIT Stem Cell Initiative, the Koch Institute Frontier Research Program through the Kathy and Curt Marble Cancer Research Fund, the Koch Institute Dana Farber/Harvard Cancer Center Bridge Project, and the American Federation of Aging Research.

MIT “Russian Doll” tech lands $7.9M international award to fight brain tumors

Researchers from MIT will work with teams in the U.K. and Europe to use nanoparticles to carry multiple drug therapies to treat glioblastoma.

Koch Institute
July 30, 2019

Tiny “Russian doll-like” particles that deliver multiple drugs to brain tumors, developed by researchers at MIT and funded by Cancer Research UK, are at the center of a new international collaboration.

Professor Paula Hammond from the Department of Chemical Engineering developed the nanoparticle technology, which will be used in an effort to treat glioblastoma — the most aggressive and deadly type of brain tumor.

Hammond will be working with Professor Michael Yaffe from the Department of Biological Engineering to determine the combinations of drugs placed within the particles, and the order and timing in which the drugs are released.

The nanoparticles — 1,000 times smaller than a human hair — are coated in a protein called transferrin, which helps them cross the blood-brain barrier. This is a membrane that keeps a tight check on anything trying to get in to the brain, including drugs.

Not only are the nanoparticles able to access hard-to-reach areas of the brain, they have also been designed to carry multiple cancer drugs at once by holding them inside layers, similarly to the way Russian dolls fit inside one another.

To make the nanoparticles even more effective, they will carry signals on their surface so that they are only taken up by brain tumor cells. This means that healthy cells should be left untouched, which will minimize the side effects of treatment.

The researchers, who are based at the Koch Institute for Integrative Cancer Research, are also working with Professor Forest White from the Department of Biological Engineering. The group are one of three international teams to have been given Cancer Research UK Brain Tumor Awards — in partnership with The Brain Tumour Charity — receiving $7.9 million of funding. The awards are designed to accelerate the pace of brain tumor research. Altogether, teams were awarded a total of $23 million.

Just last year, around 24,200 people in the United States were diagnosed with brain tumors. With around 17,500 deaths from brain tumors in the same year, survival remains tragically low.

Brain tumors represent one of the hardest types of cancer to treat because not enough is known about what starts and drives the disease, and current treatments are not effective enough.

The researchers from MIT will now work with teams in the U.K. and Europe to use the nanoparticles to carry multiple drug therapies to treat glioblastoma.

Early research carried out in the lab has already shown that nanoparticles loaded with two different drugs were able to shrink glioblastomas in mice. The team has also demonstrated that the nanoparticles can kill lymphoma cells grown in the lab, and they are also exploring their use in ovarian cancer.

The Cancer Research UK Brain Tumor Award will now allow the researchers and their collaborators to use different drug combinations to find the best parameters to tackle glioblastomas.

Drugs that have already been approved, as well as experimental drugs that have passed initial safety testing in people, will be used. Because of this, if an effective drug combination is found, the team won’t have to navigate the initial regulatory hurdles needed to get them into clinical testing, which could help get promising treatments to patients faster.

“Glioblastoma is particularly challenging because we want to get highly effective but toxic drug combinations safely across the blood-brain barrier, but also want our nanoparticles to avoid healthy brain cells and only target the cancer cells,” Hammond says. “We are very excited about this alliance between the MIT Koch Institute and our colleagues in Edinburgh to address these critical challenges.”

Unmasking mutant cancer cells

A new dosing regimen for an old cancer drug shows new promise as an immunotherapy.

Bendta Schroeder | Koch Institute
July 16, 2019

As cancer cells progress, they accumulate hundreds and even thousands of genetic and epigenetic changes, resulting in protein expression profiles that are radically different from that of healthy cells. But despite their heavily mutated proteome, cancer cells can evade recognition and attack by the immune system.

Immunotherapies, particularly checkpoint inhibitors that reinvigorate exhausted T cells, have revolutionized the treatment of certain forms of cancer. These breakthrough therapies have resulted in unprecedented response rates for some patients. Unfortunately, most cancers fail to respond to immunotherapies and new strategies are therefore needed to realize their full potential.

A team of cancer biologists including members of the laboratories of David H. Koch Professor of Biology Tyler Jacks, director of the Koch Institute for Integrative Cancer Research at MIT, and fellow Koch Institute member Forest White, the Ned C. and Janet Bemis Rice Professor and member of the MIT Center for Precision Cancer Medicine, took a complementary approach to boosting the immune system.

Although cancer cells are rife with mutant proteins, few of those proteins appear on a cell’s surface, where they can be recognized by immune cells. The researchers repurposed a well-studied class of anti-cancer drugs, heat shock protein 90 (HSP90) inhibitors, that make cancer cells easier to recognize by revealing their mutant proteomes.

Many HSP90 inhibitors have been studied extensively for the past several decades as potential cancer treatments. HSP90 protects the folded structure of a number of proteins when cells undergo stress, and in cancer cells plays an important role in stabilizing protein structure undermined by pervasive mutations. However, despite promising preclinical evidence, HSP90 inhibitors have produced discouraging outcomes in clinical trials, and none have achieved FDA approval.

In a study appearing in Clinical Cancer Research, the researchers identified a potential reason behind those disappointing results. HSP90 inhibitors have only been clinically tested at bolus doses — intermittent, large doses — that often result in unwanted side effects in patients.

RNA profiling of human clinical samples and cultured cancer cell lines revealed that this bolus-dosing schedule results in the profound suppression of immune activity as well as the activation of heat shock factor 1 protein (HSF1). Not only does HSF1 activate the cell’s heat shock response, which counteracts the effect of the HSP90 inhibitor, but it is known to be a powerful enabler of cancer cell malignancy.

In striking contrast, the researchers used cancer mouse models with intact immune systems to show that sustained, low-level dosing of HSP90 inhibitors avoids triggering both the heat shock response and the immunosuppression associated with high doses.

Using a method devised by the White lab that combines mass spectrometry-based proteomics and computational modeling, the researchers discovered that the new dosing regimen increased the number and diversity of peptides (protein fragments) on the cell surface. These peptides, which the team found to be released by HSP90 during sustained low-level inhibition, were then free to be taken up by the cell’s antigen-presenting machinery and used to flag patrolling immune cells.

“These results connect a fundamental aspect of cell biology — protein folding — to anti-tumor immune responses” says lead author Alex Jaeger, a postdoctoral fellow in the Jacks lab and a former member of the laboratory of the late MIT biologist and Professor Susan Lindquist, whose work inspired the study’s HSP90 dosing scheule. “Hopefully, our findings can reinvigorate interest in HSP90 inhibition as a complementary approach for immunotherapy.”

Using the new dosing regimen, the researchers were able to clear tumors in mouse models at drug concentrations that are 25-50 times lower than those used in clinical trials, significantly reducing the risk for toxic side effects in patients. Importantly, because several forms of HSP90 inhibitors have already undergone extensive clinical testing, the new dosing regimen can be tested in patients quickly.

This work was supported in part by the Damon Runyon Cancer Research Foundation, the Takeda Pharmaceuticals Immune Oncology Research Fund, and an MIT Training Grant in Environmental Science; foundational work on HSF1 was supported by the Koch Institute Frontier Research Program.

Cancer biologists identify new drug combo

Two drugs that block cell division synergize to kill tumor cells.

Anne Trafton | MIT News Office
July 10, 2019

When it comes to killing cancer cells, two drugs are often better than one. Some drug combinations offer a one-two punch that kills cells more effectively, requires lower doses of each drug, and can help to prevent drug resistance.

MIT biologists have now found that by combining two existing classes of drugs, both of which target cancer cells’ ability to divide, they can dramatically boost the drugs’ killing power. This drug combination also appears to largely spare normal cells, because cancer cells divide differently than healthy cells, the researchers say. They hope a clinical trial of this combination can be started within a year or two.

“This is a combination of one class of drugs that a lot of people are already using, with another type of drug that multiple companies have been developing,” says Michael Yaffe, a David H. Koch Professor of Science and the director of the MIT Center for Precision Cancer Medicine. “I think this opens up the possibility of rapid translation of these findings in patients.”

The discovery was enabled by a new software program the researchers developed, which revealed that one of the drugs had a previously unknown mechanism of action that strongly enhances the effect of the other drug.

Yaffe, who is also a member of the Koch Institute for Integrative Cancer Research, is the senior author of the study, which appears in the July 10 issue of Cell Systems. Koch Institute research scientists Jesse Patterson and Brian Joughin are the first authors of the paper.

Unexpected synergy

Yaffe’s lab has a longstanding interest in analyzing cellular pathways that are active in cancer cells, to find how these pathways work together in signaling networks to create disease-specific vulnerabilities that can be targeted with multiple drugs. When the researchers began this study, they were looking for a drug that would amplify the effects of a type of drug known as a PLK1 inhibitor. Several PLK1 inhibitors, which interfere with cell division, have been developed, and some are now in phase 2 clinical trials.

Based on their previous work, the researchers knew that PLK1 inhibitors also produce a type of DNA and protein damage known as oxidation. They hypothesized that pairing PLK1 inhibitors with a drug that prevents cells from repairing oxidative damage could make them work even better.

To explore that possibility, the researchers tested a PLK1 inhibitor along with a drug called TH588, which blocks MTH1, an enzyme that helps cells counteract oxidative damage. This combination worked extremely well against many types of human cancer cells. In some cases, the researchers could use one-tenth of the original doses of each drug, given together, and achieve the same rates of cell death of either drug given on its own.

“It’s really striking,” Joughin says. “It’s more synergy than you generally see from a rationally designed combination.”

However, they soon realized that this synergy had nothing to do with oxidative damage. When the researchers treated cancer cells missing the gene for MTH1, which they thought was TH588’s target, they found that the drug combination still killed cancer cells at the same high rates.

“Then we were really stuck, because we had a good combination, but we didn’t know why it worked,” Yaffe says.

To solve the mystery, they developed a new software program that allowed them to identify the cellular networks most affected by the drugs. The researchers tested the drug combination in 29 different types of human cancer cells, then fed the data into the software, which compared the results to gene expression data for those cell lines. This allowed them to discover patterns of gene expression that were linked with higher or lower levels of synergy between the two drugs.

This analysis suggested that both drugs were targeting the mitotic spindle, a structure that forms when chromosomes align in the center of a cell as it prepares to divide. Experiments in the lab confirmed that this was correct. The researchers had already known that PLK1 inhibitors target the mitotic spindle, but they were surprised to see that TH588 affected the same structure.

“This combination that we found was very nonobvious,” Yaffe says. “I would never have given two drugs that both targeted the same process and expected anything better than just additive effects.”

“This is an exciting paper for two reasons,” says David Pellman, associate director for basic science at Dana-Farber/Harvard Cancer Center, who was not involved in the study. “First, Yaffe and colleagues make an important advance for the rational design of drug therapy combinations. Second, if you like scientific mysteries, this is a riveting example of molecular sleuthing. A drug that was thought to act in one way is unmasked to work through an entirely different mechanism.”

Disrupting mitosis

The researchers found that while both of the drugs they tested disrupt mitosis, they appear to do so in different ways. TH588 binds to microtubules, which form the mitotic spindle, and slows their assembly. Many similar microtubule inhibitors are already used clinically to treat cancer. The researchers showed that some of those microtubule inhibitors also synergize with PLK1 inhibitors, and they believe those would likely be more readily available for rapid use in patients than TH588, the drug they originally tested.

While the PLK1 protein is involved in multiple aspects of cell division and spindle formation, it’s not known exactly how PLK1 inhibitors interfere with the mitotic spindle to produce this synergy. Yaffe said he suspects they may block a motor protein that is necessary for chromosomes to travel along the spindle.

One potential benefit of this drug combination is that the synergistic effects appear to specifically target cancer cell division and not normal cell division. The researchers believe this could be because cancer cells are forced to rely on alternative strategies for cell division because they often have too many or too few chromosomes, a state known as aneuploidy.

“Based on the work we have done, we propose that this drug combination targets something fundamentally different about the way cancer cells divide, such as altered cell division checkpoints, chromosome number and structure, or other structural differences in cancer cells,” Patterson says.

The researchers are now working on identifying biomarkers that could help them to predict which patients would respond best to this drug combination. They are also trying to determine the exact function of PLK1 that is responsible for this synergy, in hopes of finding additional drugs that would block that interaction.

The research was funded by the National Institutes of Health, the Charles and Marjorie Holloway Foundation, the Ovarian Cancer Research Fund, the MIT Center for Precision Cancer Medicine, the Koch Institute Dana Farber/Harvard Cancer Center Bridge Project, an American Cancer Society Postdoctoral Fellowship, the Koch Institute Support (core) Grant from the National Cancer Institute, and the Center for Environmental Health Support Grant.