Research Area: Cancer Biology

Tracing a cancer’s family tree to its roots reveals how tumors grow

Family trees of lung cancer cells reveal how cancer evolves from its earliest stages to an aggressive form capable of spreading throughout the body.

Greta Friar | Whitehead Institute
May 5, 2022

Over time, cancer cells can evolve to become resistant to treatment, more aggressive, and metastatic — capable of spreading to additional sites in the body and forming new tumors. The more of these traits that a cancer evolves, the more deadly it becomes. Researchers want to understand how cancers evolve these traits in order to prevent and treat deadly cancers, but by the time cancer is discovered in a patient, it has typically existed for years or even decades. The key evolutionary moments have come and gone unobserved.

MIT Professor Jonathan Weissman and collaborators have developed an approach to track cancer cells through the generations, allowing researchers to follow their evolutionary history. This lineage-tracing approach uses CRISPR technology to embed each cell with an inheritable and evolvable DNA barcode. Each time a cell divides, its barcode gets slightly modified. When the researchers eventually harvest the descendants of the original cells, they can compare the cells’ barcodes to reconstruct a family tree of every individual cell, just like an evolutionary tree of related species. Then researchers can use the cells’ relationships to reconstruct how and when the cells evolved important traits. Researchers have used similar approaches to follow the evolution of the virus that causes Covid-19, in order to track the origins of variants of concern.

Weissman and collaborators have used their lineage-tracing approach before to study how metastatic cancer spreads throughout the body. In their latest work, Weissman; Tyler Jacks, the Daniel K. Ludwig Scholar and David H. Koch Professor of Biology at MIT; and computer scientist Nir Yosef, associate professor at the University of California at Berkeley and the Weizmann Institute of Science, record their most comprehensive cancer cell history to date. The research, published today in Cell, tracks lung cancer cells from the very first activation of cancer-causing mutations. This detailed tumor history reveals new insights into how lung cancer progresses and metastasizes, demonstrating the wealth of understanding that lineage tracing can provide.

“This is a new way of looking at cancer evolution with much higher resolution,” says Weissman, who is a professor of biology at MIT, a member of the Whitehead Institute for Biomedical Research, and an investigator with Howard Hughes Medical Institute. “Previously, the critical events that cause a tumor to become life-threatening have been opaque because they are lost in a tumor’s distant past, but this gives us a window into that history.”

In order to track cancer from its very beginning, the researchers developed an approach to simultaneously trigger cancer-causing mutations in cells and start recording the cells’ history. They engineered mice such that when their lung cells were exposed to a tailor-made virus, that exposure activated a cancer-causing mutation in the Kras gene and deactivated tumor suppressing gene Trp53 in the cells, as well as activating the lineage tracing technology. The mouse model, developed in Jacks’ lab, was also engineered so that lung cancer would develop in it very similarly to how it would in humans.

“In this model, cancer cells develop from normal cells and tumor progression occurs over an extended time in its native environment. This closely replicates what occurs in patients,” Jacks says. Indeed, the researchers’ findings closely align with data about disease progression in lung cancer patients.

The researchers let the cancer cells evolve for several months before harvesting them. They then used a computational approach developed in their previous work to reconstruct the cells’ family trees from their modified DNA barcodes. They also measured gene expression in the cells using RNA sequencing to characterize each individual cell’s state. With this information, they began to piece together how this type of lung cancer becomes aggressive and metastatic.

“Revealing the relationships between cells in a tumor is key to making sense of their gene expression profiles and gaining insight into the emergence of aggressive states,” says Yosef, who is a co-corresponding author on both the current work and the previous lineage tracing paper.

The results showed significant diversity between subpopulations of cells within the same tumor. In this model, cancer cells evolved primarily through inheritable changes to their gene expression, rather than through genetic mutations. Certain subpopulations had evolved to become more fit — better at growth and survival — and more aggressive, and over time they dominated the tumor. Genes that the researchers identified as commonly expressed in the fittest cells could be good candidates for possible therapeutic targets in future research. The researchers also discovered that metastases originated only from these groups of dominant cells, and only late in their evolution. This is different from what has been proposed for some other cancers, in which cells may gain the ability to metastasize early in their evolution. This insight could be important for cancer treatment; metastasis is often when cancers become deadly, and if researchers know which types of cancer develop the ability to metastasize in this stepwise manner, they can design interventions to stop the progression.

“In order to develop better therapies, it’s important to understand the fundamental principles that tumors adopt to develop,” says co-first author Dian Yang, a Damon Runyon Postdoctoral Fellow in Weissman’s lab. “In the future, we want to be able to look at the state of the cancer cells when a patient comes in, and be able to predict how that cancer’s going to evolve, what the risks are, and what is the best treatment to stop that evolution.”

The researchers also figured out important details of the evolutionary paths that cancer subpopulations take to become fit and aggressive. Cells evolve through different states, defined by key characteristics that the cell has at that point in time. In this cancer model the researchers found that early on, cells in a tumor quickly diversified, switching between many different states. However, once a subpopulation landed in a particularly fit and aggressive state, it stayed there, dominating the tumor from that stable state. Furthermore, the ultimately dominant cells seemed to follow one of two distinct paths through different cell states. Either of those paths could then lead to further progression that enabled cancers to enter aggressive “mesenchymal” cell states, which are linked to metastasis.

After the researchers thoroughly mapped the cancer cells’ evolutionary paths, they wondered how those paths would be affected if the cells experienced additional cancer-linked mutations, so they deactivated one of two additional tumor suppressors. One of these affected which state cells stabilized in, while the other led cells to follow a completely new evolutionary pathway to fitness.

The researchers hope that others will use their approach to study all kinds of questions about cancer evolution, and they already have a number of questions in mind for themselves. One goal is to study the evolution of therapeutic resistance, by seeing how cancers evolve in response to different treatments. Another is to study how cancer cells’ local environments shape their evolution.

“The strength of this approach is that it lets us study the evolution of cancers with fine-grained detail,” says co-first author Matthew Jones, a graduate student in the Weissman and Yosef labs. “Every time there is a shift from bulk to single-cell analysis in a technology or approach, it dramatically widens the scope of the biological insights we can attain, and I think we are seeing something like that here.”

A spectrum of cancer cells
Greta Friar | Whitehead Institute
April 11, 2022

Cancer is at its most deadly when it spreads and forms tumors in new tissues. This process, called metastasis, is responsible for the vast majority of cancer deaths, and yet there is still a lot that researchers do not know about how and when it happens. Whitehead Institute Founding Member Robert Weinberg, also the Daniel K. Ludwig Professor for Cancer Research at the Massachusetts Institute of Technology, studies the mechanisms behind metastasis. One such mechanism is a process called the epithelial-mesenchymal transition (EMT), which causes epithelial cells, which normally stick tightly together, to lose their cohesion, enabling them to move around and even invade nearby tissue. This EMT program also operates during embryonic development. Cancer cells can co-opt this process and use it travel from their original tumor site to distant tissues throughout the body. Some of the cancer cells that spread are able, on rare occasions, to form new tumors in these tissues—metastases—while the great majority of these cells remain dormant after entering the distant tissues.

New research from Weinberg and postdoc Yun Zhang shows that cells change in diverse ways through the actions of the EMT, which can influence whether cells are able to form new tumors after they spread. The work, published in Nature Cell Biology on April 11, 2022, also identifies two regulators of the EMT and shows that loss of each regulator leads to a different metastatic risk profile.

“Using triple negative breast cancer as a model, we are trying to go a bit deeper into understanding the molecular mechanisms that regulate the EMT, how cells enter into different EMT intermediate states, and which of these states contribute to metastasis,” Zhang says.

The EMT was originally imagined as a sort of binary switch, in which cells start out epithelial and become mesenchymal, much like a light switch being flicked from off to on. However, researchers are learning that the EMT works more like a dimmer switch that can be shifted along a spectrum of brightness. Cells that undergo the EMT usually end up in hybrid states between the epithelial and mesenchymal extremes. These cells in the middle of the spectrum, which have some characteristics of each extreme, are called “quasi-mesenchymal” cells, and it turns out that they–rather than cells that become fully mesenchymal–are the most capable of metastasizing and forming new tumors throughout the body.

Protected versus plastic cells

Weinberg and Zhang set out to better understand the EMT spectrum and what controls cells’ movement along it. First, they compared epithelial cells to each other and found that some were more plastic or prone to transitioning along the EMT spectrum than others. They also used the CRISPR gene editing tool to screen for genes that might be regulating the cells’ plasticity. If researchers can learn what makes a cell become quasi-mesenchymal—posing a high risk for metastasis—they might be able use this information, at some time in the future, to develop strategies to prevent cells from entering this high-risk state.

The CRISPR gene screen turned up a number of molecules that seemed to influence cells’ epithelial-mesenchymal plasticity. Two groups of these molecules had especially strong effects: PRC2, a complex that operates in chromosomes to silence or inactivate genes, and KMT2D-COMPASS, a complex that helps activate genes. Both complexes help to keep cells in a stable epithelial state. Loss of either complex makes cells more prone to moving along the EMT spectrum.

The researchers then determined how the loss of either complex enables the EMT. PRC2 normally silences several key EMT-related genes. When PRC2 is lost, those genes activate, which in turn sensitizes the cell to a signal that can trigger the EMT. The loss of KMT2D-COMPASS affects how well PRC2 can bind its targets, leading to the same signal sensitivity. In spite of the similar mechanisms at play, the loss of PRC2 versus KMT2D-COMPASS leads cells to transition to end up in different EMT states, an exciting finding for the researchers. Cells without KMT2D-COMPASS became fully mesenchymal, while cells without PRC2 became hybrid or quasi-mesenchymal. Consequently, cells without PRC2 were much more capable of metastasis than cells without KMT2D-COMPASS (or cells in which both complexes were active) in mouse models. When the researchers looked at historical data from breast cancer patients, they observed the same pattern: people with faulty PRC2 component genes had worse outcomes. These findings provide further evidence that cells in the middle of the EMT spectrum are most likely to metastasize.

This work supports the understanding of the EMT as a spectrum rather than a simple switch, and shows that different EMT regulators can program cells to transition to different parts of the EMT spectrum. Additionally, the finding that loss of PRC2 is linked to metastasis has implications for cancer drugs currently in development that work by inactivating PRC2. Benefits of the drugs may outweigh risks for patients with certain types of cancer for which PRC2 is an effective target. However, Weinberg and Zhang caution that researchers leading clinical trials of PRC2-targeting drugs should be careful about selecting patients and monitoring outcomes. In the types of cancer cells that the researchers looked at, even temporary PRC2 inactivation, such as from a therapy trial, was sufficient to trigger cells to become EMT hybrids with increased metastatic capacity.

Weinberg and Zhang intend to continue exploring the genes identified in their CRISPR screen to see if they can identify other hybrid states along the EMT spectrum, in which cells have different combinations of epithelial and mesenchymal features. They hope that by deepening their understanding of the gene expression profiles of cancer cells associated with different EMT trajectories, they can contribute to the development of therapies for people with potentially metastatic cancers.

“Understanding when and how cancer cells become able to form life-threatening metastases is crucial in order to help the many patients for whom this is a risk,” Weinberg says. “This work provides new insights into the mechanisms that enable cells to metastasize and the roles that different EMT programs can play.”

Mentorship and medicine

MIT senior Daniel Zhang aims to provide hope for young patients and support to young students.

Celina Zhao | Department of Biology
February 24, 2022

During the virtual spring 2020 semester, Daniel Zhang, a senior majoring in biology, put his time at home to good use. In the garage of his home in San Diego, California, Zhang helped his 13-year-old brother build a lab to study dry eye disease.

This combination of mentorship and medicine feels like second nature to Zhang. When his parents opened a family-run optometry clinic, Zhang was their first patient and then their receptionist. And after a close family member passed away from leukemia, he remembers thinking, “Humans are susceptible to so many diseases — why don’t we have better cures?”

That question propelled him to spend his high school summers studying biomarkers for the early detection of leukemia at the University of California at San Diego. He was invited to present his research at the London International Youth Science Forum, where he spoke to scientists from almost 70 countries. Afterward, he was hooked on the idea of scientific research as a career.

“Research is like standing on the shoulders of giants,” he says. “My experience at the forum was when I knew I loved science and wanted to continue using it to find common ground with others from completely different cultures and backgrounds.”

Exploring the forefront of cancer research

As soon as he arrived at MIT as a first-year undergraduate, Zhang began working under the guidance of postdoc Peter Westcott in professor Tyler Jacks’ lab. The lab focuses on developing better mouse and organoid models to study cancer progression — in Zhang’s case, metastatic colorectal cancer.

One of the ways to model colorectal cancer is by injecting an engineered virus directly into the colons of mice. The viruses, called lentiviral agents, “knock out” tumor suppressor genes and activate the so-called oncogenes that drive cancer forward. However, the imprecise nature of this injection also unintentionally transforms many “off-target” cells into cancer cells, producing a cancer that’s far too widespread and aggressive. Additionally, rare tumors called sarcomas are often initiated rather than adenocarcinomas, the type of tumor found in 95 percent of human cases. As a result, these mouse models are limited in their ability to accurately model colorectal cancer.

To address this problem, Zhang and Westcott designed a method using CRISPR/Cas9 to target a special stem cell called LGR5+, which researchers believe are the types of cells that, when mutated, grow into colorectal cancer. His technique modifies only the LGR5+ cells, which would allow researchers to control the rate at which adenocarcinomas grow. Therefore, it generates a model that is not only much more similar to human colorectal cancer than other models, but also allows researchers to quickly test for other potential cancer driver genes with CRISPR/Cas9. Designing an accurate model is crucial for developing and testing effective new therapies for patients, Zhang says.

During MIT’s virtual spring and fall semesters of 2020, Zhang shifted his focus from benchwork in the lab to computational biology. Using patient data from the Cancer Genome Atlas, Zhang analyzed mutation rates and discovered three genes potentially involved in colorectal cancer tumor suppression. He plans to test their function in his new mouse model to further validate how the dysfunction of these genes drives colorectal cancer progression.

For his work on organoid modeling of colorectal cancer, a third project he’s worked on during his time at the Jacks lab, he also won recognition from the American Association for Cancer Research (AACR). As one of 10 winners of the Undergraduate Scholar Award, he had the opportunity to present his research at the virtual AACR conference in 2021 and again at the next AACR Conference in New Orleans in April 2022.

He credits MIT’s “mens et manus” philosophy, encouraging the hands-on application of knowledge, as a large part of his early success in research.

“I’ve found that, at MIT, a lot of people are pursuing projects and asking questions that have never been thought of before,” Zhang says. “No one has ever been able to develop a late-stage model for colorectal cancer that’s amenable to gene editing. As far as I know, other than us, no one in the world is even working on this.”

Inspiring future generations to pursue STEM

Outside of the lab, Zhang devotes a substantial amount of time to sharing the science he’s so passionate about. Not only has he been awarded the Gene Brown Prize for undergraduate teaching for his time as a teaching assistant for the lab class 7.002 (Fundamentals of Experimental Molecular Biology), but he’s also taken on leadership roles in science outreach activities.

During the 2020-21 academic year, he served as co-director of DynaMIT, an outreach program that organizes a two-week STEM program over the summer for underserved sixth to ninth graders in the greater Boston area. Although the program is traditionally held in-person, in summer 2021 it was held virtually. But Zhang and the rest of the board didn’t let the virtual format deter them from maximizing the fun and interactive nature of the program. They packed and shipped nearly 120 science kits focused on five major topics — astronomy, biology, chemistry, mechanical engineering, and math — allowing the students to explore everything from paper rockets to catapults and trebuchets to homemade ice cream.

“At first, we were worried that most of the students wouldn’t turn on their cameras, since we saw that trend all over MIT classes during the semester,” Zhang says. “But almost everyone had their cameras on the entire time. It was really gratifying to see students come in on Monday really shy, but by Friday be actively participating, making jokes with the mentors, and being really excited about STEM.”

To investigate the long-term impacts of the program, he also helped kick-start a project that followed up with DynaMIT alumni, some of whom have already graduated from college. Zhang says: “We were happy to see that 80-90 percent of DynaMIT alumni enjoyed the program, rating it four or five out of five, and close to 70 percent of them said that DynaMIT had a really positive impact on their trajectory toward a career in STEM.”

Zhang has also served as president of the MIT Pre-medical Society, with the goals of fostering an encouraging environment for premed undergraduates, and providing guidance and resources to first- and second-year students still undecided about the premed path. To achieve these objectives, he pioneered an MIT-hosted mixer with the premedical societies of other Boston colleges, including Wellesley College, Boston University, Tufts University, and Harvard University. At the mixer, students were able to network with each other and listen to guest speakers from the different universities talk about their experiences in medicine. He also started a “big/little” initiative that paired third- and fourth-year mentors with first- and second-year students.

Providing new opportunity and hope

The wealth of activities Zhang has participated in at MIT has inspired his choices for the future. After graduation, he plans to take a gap year and work as a research technician in pediatric oncology before applying to MD/PhD programs.

On the mentorship side, he’s currently working to establish a nonprofit organization called Future African Scientist with his former Ugandan roommate, Martin Lubowa, whom he met at a study abroad program during MIT’s Independent Activities Period in 2020. The organization will teach high schoolers in Africa professional skills and expose them to different STEM topics — a project Zhang plans to work on post-MIT and into the long term.

Ultimately, he hopes to lead his own lab at the intersection of CRISPR-Cas9 technology and cancer biology, and to serve as a mentor to future generations of researchers and physicians.

As he puts it: “All of the experiences I’ve had so far have solidified my goal of conducting research that impacts patients, especially young ones. Being able to provide new opportunity and hope to patients suffering from late-stage metastatic diseases with no current cures is what inspires me every day.”

Advocating for vaccine equity

Postdoc Dig Bijay Mahat became a cancer researcher to improve healthcare in Nepal, but the COVID-19 pandemic exposed additional resource disparities.

Raleigh McElvery
February 17, 2022

When Dig Bijay Mahat arrived at MIT in 2017 to begin his postdoctoral studies, he had one very clear goal: to become an expert in cancer research and diagnostics so he could improve healthcare in Nepal, where he was born. In 2020, when the COVID-19 pandemic laid bare additional discrepancies in resource equity around the world, his goal did not waiver. But it did expand to fill a more immediate need — help Nepal find the best way to navigate widespread COVID testing requirements and vaccine rollouts.

Mahat was born in the western region of Nepal, where his family has owned a large swath of land for generations. Before Mahat was born, his grandfather passed away unexpectedly. And, as the eldest son, Mahat’s father assumed responsibility for his five of siblings at the age of 21. As a result, Mahat’s father missed his chance to pursue the education he’d envisioned. Perhaps because of this, he made it his mission to give Mahat the education he never received. However, no school was quite good enough, and he shuffled Mahat between nine different institutions before the age of 18.

While his father wished him all the success and prestige that would come with pursing a medical career, Mahat had other plans. Toward the end of high school, he became captivated by song writing, and even secretly used his school tuition money one semester to record an album. “It was a disastrous flop,” he now recalls with a smile.

Although his foray into the music industry provides comic relief today, at the time Mahat was dismayed to be back on the medical track. However, he did convince his father to let him go to the US for college. He ended up at Towson University in Maryland, living with his aunt and uncle and delivering pizzas to support his nuclear family back in Nepal. Some weeks, he clocked in over 100 hours of deliveries.

As a molecular biology, biochemistry, and bioinformatics major, he took every research opportunity he could get, and became enthralled by breast cancer research. Shortly thereafter, his mother was diagnosed with the same disease, which further strengthened his conviction to learn as much as he could in the US, and return to Nepal to help as many patients as he could.

“The state of cancer diagnostics is very poor in Nepal,” he explains. Patient biopsies must be sent to other countries such as India — a costly practice at the mercy of politics and travel restrictions. “The least we can do is become self-sufficient and provide these vital molecular diagnostics tools to our own people,” Mahat says.

He went on to earn his PhD in molecular biology and genetics from Cornell University, and by the fall of 2017 he had secured his dream job: a postdoctoral position in the lab of MIT Professor of Biology Susan Lindquist. Mahat had spent much of his time at Cornell studying a protein known as heat shock factor 1, and Lindquist had conducted seminal work showing that this same protein enables healthy cells to suddenly turn into cancer cells. Just as he had finalized his new apartment lease and was preparing to start his new job, Lindquist wrote from the hospital to tell him she had late-stage ovarian cancer, and suggested he complete his postdoctoral studies elsewhere.

Gutted, he scrambled to find another position, and built up the courage to contact MIT professor, Koch Institute member, and Nobel laureate Phil Sharp. Mahat put together a formal research proposal and presented it to Sharp. A few days later, he became the lab’s newest member.

“From the beginning, the things that struck me about Phil were his humility, his attention to experimental detail, and his inexplicable reservoir of insight,” Mahat says. “If I could carry even just some of that same humility with me for the rest of my life, I would be a good human being.”

In 2018, Mahat and Sharp filed a patent with the potential to revolutionize disease diagnostics. Widely-available single-cell sequencing technologies reveal the subset of RNAs inside a cell that build proteins. But Mahat and his colleagues found a way to take a snapshot of all the RNA inside a single cell that is being transcribed from DNA — including RNAs that will never become proteins. Because many ailments arise from mutations in the “non-coding” DNA that gives rise to this “non-coding” RNA, the researchers hope their new method will help expose the function of non-coding variants in diseases like diabetes, autoimmune disorders, neurological diseases, and cancer.

Mahat was still immersed in this research in early 2020 when the COVID-19 pandemic began to escalate. As case numbers soared around the world, it became clear to him that the wealth of COVID testing resources available on MIT’s campus — and throughout the US in general — dwarfed the means available to his family back in Nepal. Polymerase chain reaction (PCR) testing remains the most popular and accurate means to detect the virus in patient samples. While PCR machines are quite common in molecular biology labs across the US, the entire country of Nepal owned just a few at the start of the pandemic, according to Mahat.

“Digbijay was focused intensely on developing our novel single-cell technology when he became aware of Nepal’s challenges to control the COVID-19 pandemic,” Sharp recalls. “While continuing his research in the lab, he spent several months contacting leaders in pharmaceutical companies in the US and leaders in public health in Nepal to help arrange access to vaccines and rapid tests.”

Mahat was already in contact with the Nepali Ministry of Health and Population regarding the state of the country’s cancer diagnostics, and so the government called on him to advise their COVID testing efforts. Given the high cost and limited availability of PCR machines and reagents, Mahat began discussions with MIT spinoff Sherlock Biosciences, in order to bring an alternative testing technology to Nepal. These COVID tests, which were developed at the Broad Institute of MIT and Harvard, use the CRISPR/Cas9 system — rather than PCR — to detect the SARS-CoV2 virus that causes COVID-19, making them cheaper and more readily available. Sherlock Biosciences ultimately donated $100,000-worth of testing kits, supplemented by an additional $100,000 grant from the Open Philanthropy Project to help purchase the equipment necessary to implement the tests. In December of 2020, Mahat and his wife Rupa Shah flew to Nepal to set up a testing center using these new resources.

Although this required Mahat to briefly pause his MIT research, Sharp was supportive of these extracurricular pursuits. “We are very proud of Jay’s effective work benefiting the people of Nepal,” Sharp says.

Around the same time, Mahat reached out to Institute professor and Moderna co-founder Robert Langer to help initiate vaccine talks with the Nepali government. Through Sharp’s contacts, Mahat was also able to connect the government with Johnson & Johnson. In addition, Mahat, Sharp, and Emeritus Professor Uttam RajBhandary wrote a letter to MIT president Rafael Reif, who joined other university leadership in urging the Biden administration to donate vaccines to low-income countries.

Nepal ultimately received its COVID-19 vaccines through the COVAX program, co-led by the Coalition for Epidemic Preparedness Innovations, GAVI Alliance, and the World Health Organization. Today, the country has begun administering boosters. There were also some funds left over from the Open Philanthropy Project grant, which went toward sending Nepal several thousand PCR kits designed to distinguish between the delta and omicron variants. Professor Tyler Jacks, the Koch Institute director at that time, also connected Mahat with the company Thermo Fisher Scientific to secure additional PCR reagents.

Roshan Pokhrel, the Secretary of Nepal’s Ministry of Health and Population, met Mahat prior to the pandemic, and relied on his expertise to begin establishing Nepal’s National Cancer Institute (NCI) in 2020. “It was his cooperation and coordination that helped us set up NCI,” Pokhrel says. “Mr. Mahat’s continuous support during the first two waves of our COVID-19 vaccine distribution was also highly appreciated. During the recent omicron outbreak, his support in our public laboratory helped us to monitor the variant.”

Bhagawan Koirala, chairman of the Nepal Medical Council, participated in the vaccine talks that Mahat organized between Nepal’s Ministry of Health and Johnson & Johnson. Koirala says he was impressed by Mahat’s exceptional credentials and his modesty, as well as his desire to promote cancer research and diagnostics. As the chairman of the Kathmandu Institute of Child Health, Koirala hopes to engage Mahat’s expertise in the future to help advance pediatric cancer research in Nepal.

“We have spoken extensively about the policies regarding cancer diagnostics in Nepal,” Koirala says. “Dr. Mahat and I are eager to work with the government to introduce policies that will help develop local diagnostic capacity and discourage sending patient samples out of the country. This will save costs, ensure patient privacy, and improve quality of care and research.”

These days, Mahat is nothing short of a local celebrity in Nepal. Despite his current drive for ensuring vaccine equity, his ultimate goal is still to work with individuals like Koirala and Pokhrel to bring cancer treatment resources to the country. He not only envisions setting up his own research center there, but also inspiring young people to pursue careers in research. “Before me, no one in my entire village had pursued a scientific career, so if I could motivate even a few young kids to follow that path, it would be a win for me.”

But, he adds, he’s not ready to leave MIT just yet; he still has more to learn. “I feel privileged and honored to be part of this compassionate community,” he says. “I’m also proud — proud that we’ve been able to come together in this time of need.”

A stealthy way to combat tumors

MIT biologists show that helper immune cells disguised as cancer cells can help rejuvenate T cells that attack tumors.

Anne Trafton | MIT News Office
November 18, 2021

Under the right circumstances, the body’s T cells can detect and destroy cancer cells. However, in most cancer patients, T cells become disarmed once they enter the environment surrounding a tumor.

Scientists are now trying to find ways to help treat patients by jumpstarting those lackluster T cells. Much of the research in this field, known as cancer immunotherapy, has focused on finding ways to stimulate those T cells directly. MIT researchers have now uncovered a possible new way to indirectly activate those T cells, by recruiting a population of helper immune cells called dendritic cells.

In a new study, the researchers identified a specific subset of dendritic cells that have a unique way of activating T cells. These dendritic cells can cloak themselves in tumor proteins, allowing them to impersonate cancer cells and trigger a strong T cell response.

“We knew that dendritic cells are incredibly important for the antitumor immune response, but we didn’t know what really constitutes the optimal dendritic cell response to a tumor,” says Stefani Spranger, the Howard S. and Linda B. Stern Career Development Professor at MIT and a member of MIT’s Koch Institute for Integrative Cancer Research.

The results suggest that finding ways to stimulate that specific population of dendritic cells could help to enhance the effectiveness of cancer immunotherapy, she says. In a study of mice, the researchers showed that stimulating these dendritic cells slowed the growth of melanoma and colon tumors.

Spranger is the senior author of the study, which appears today in the journal Immunity. The lead author of the paper is MIT graduate student Ellen Duong.

Spontaneous regression

When tumors begin to form, they produce cancerous proteins that T cells recognize as foreign. This sometimes allows T cells to eliminate tumors before they get very large. In other cases, tumors are able to secrete chemical signals that deactivate T cells, allowing the tumors to continue growing unchecked.

Dendritic cells are known to help activate tumor-fighting T cells, but there are many different subtypes of dendritic cells, and their individual roles in T cell activation are not fully characterized. In this study, the MIT team wanted to investigate which types of dendritic cells are involved in T cell responses that successfully eliminate tumors.

To do that, they found a tumor cell line, from a type of muscle tumor, that has been shown to spontaneously regress in mice. Such cell lines are difficult to find because researchers usually don’t keep them around if they can’t form tumors, Spranger says.

Studying mice, they compared tumors produced by that regressive cell line with a type of colon carcinoma, which forms tumors that grow larger after being implanted in the body. The researchers found that in the progressing tumors, the T cell response quickly became exhausted, while in the regressing tumors, T cells remained functional.

The researchers then analyzed the dendritic cell populations that were present in each of these tumors. One of the main functions of dendritic cells is to take up debris from dying cells, such as cancer cells or cells infected with a pathogen, and then present the protein fragments to T cells, alerting them to the infection or tumor.

The best-known type of dendritic cells required for antitumor immunity  are DC1 cells, which interact with T cells that are able to eliminate cancer cells. However, the researchers found that DC1 cells were not needed for tumor regression. Instead, using single-cell RNA sequencing technology, they identified a previously unknown activation state of DC2 cells, a different type of dendritic cell, that was driving T cell activation in the regressing tumors.

The MIT team found that instead of ingesting cellular debris, these dendritic cells swipe proteins called MHC complexes from tumor cells and display them on their own surfaces. When T cells encounter these dendritic cells masquerading as tumor cells, the T cells become strongly activated and begin killing the tumor cells.

This specialized population of dendritic cells appears to be activated by type one interferon, a signaling molecule that cells usually produce in response to viral infection. The researchers found a small population of these dendritic cells in colon and melanoma tumors that progress, but they were not properly activated. However, if they treated those tumors with interferon, the dendritic cells began stimulating T cells to attack tumor cells.

Targeted therapy

Some types of interferon have been used to help treat cancer, but it can have widespread side effects when given systemically. The findings from this study suggest that it could be beneficial to deliver interferon in a very targeted way to tumor cells, or to use a drug that would provoke tumor cells to produce type I interferon, Spranger says.

The researchers now plan to investigate just how much type I interferon is needed to generate a strong T cell response. Most tumor cells produce a small amount of type I interferon but not enough to activate the dendritic cell population that invigorates T cells. On the other hand, too much interferon can be toxic to cells.

“Our immune system is hardwired to respond to nuanced differences in type I interferon very dramatically, and that is something that is intriguing from an immunological perspective,” Spranger says.

The research was funded by the Koch Institute Support (core) Grant from the National Cancer Institute, a National Institutes of Health Pre-Doctoral Training Grant, a David H. Koch Graduate Fellowship, and the Pew-Steward Fellowship.

Stem Cell Research Zeroes in on Cancer

Collaborators investigate colon health with novel tools

Deborah Halber | Spectrum
November 9, 2021

In a building at the edge of the Massachusetts General Hospital (MGH) complex, Ömer Yilmaz, MD, and a group of pathology residents gather around a microscope. A resident reads from a chart: a growth was found in the intestine of a patient who had complained of abdominal pain.

Yilmaz, an MIT cancer researcher and a gastrointestinal pathologist, hoped a closer look at the tumor would reveal a noncancerous collection of fat cells or lymphoid cells.

It had taken a couple of days to prepare the biopsy. Somewhere in the hospital, the patient and her family were anxiously awaiting a diagnosis. Yilmaz leaned forward and adjusted the focus on the microscope.

On the tracks of cancer

If the long, twisting tube of the human digestive tract were stretched out straight, it would extend 30 feet, and its absorptive surface area is roughly comparable to the size of a tennis court. A significant chunk of that tube is the large intestine, an intricate place rife with microscopic structures called niches and crypts, evoking an underground cavern or the ocean floor. Besides the skin, the intestines are the body’s primary barrier against external invaders.

Yilmaz, an associate professor of biology at the Koch Institute for Integrative Cancer Research, believes certain cancers and diseases such as inflammatory bowel disease originate with a breakdown of the intestine’s protective barrier. Diet appears to affect intestinal stem cells; these cells can morph into a variety of cell types, and changes in stem cells can lead to cancer, but no one understands exactly how this occurs.

That’s where Yilmaz’s partnership with MIT biomedical engineer and chemist Alex Shalek comes in. Yilmaz and Shalek are both members of the MIT Stem Cell Initiative, which focuses on fundamental biological questions about benign and cancerous adult stem cells.

Shalek, a core member of the Institute for Medical Engineering and Science (IMES), a member of the Koch Institute, and an associate professor of chemistry, develops experimental and computational tools that provide researchers with detailed snapshots of what’s going on inside living cells at a moment in time. Some of these tools, Yilmaz hoped, would enable him to see how intestinal cells react when they encounter an influx of fat or are deprived of food for hours or days.

“In the past, people would have taken a piece of gut that had many different cell types and said, ‘What changes, on average, under different dietary conditions?’” Shalek says. His tools give him and Yilmaz more precise information, providing a window into the discrete molecular responses of individual cells within the colon.

The role of stem cells

Growing up in Battle Creek, Michigan, Yilmaz spent all his free time trailing after his father, a physician who had immigrated from Turkey. He’d make hospital rounds with his dad, visiting the pathology and radiology labs. As Yilmaz grew older, the two would talk about the mechanisms underlying disease.

After completing his MD/PhD at the University of Michigan, Yilmaz did his residency in pathology, the study of disease, at MGH. He began working at the Whitehead Institute with MIT biology professor David M. Sabatini, a pioneer in elucidating the mechanisms under-lying the regulation of growth and metabolism in mammals. Yilmaz had long been fascinated with stem cells’ seemingly miraculous ability to become any kind of cell the body needed. In adults, stem cells are relatively rare, best studied in bone marrow.

When scientists first found stem cells in the intestine in 2007, Yilmaz shifted his research focus. “As soon as intestinal stem cells were identified, I became interested in understanding how they are regulated by diet and aging,” he says.

“We know obesity elevates cancer risk in a wide range of tissues, including the colon, but we don’t know exactly how. And fasting regimens have been known to improve organ and tissue health, but this, too, is not well understood.”

To better study the transition from healthy to diseased cells in the colon, Yilmaz’s team generated colon tumors in mice that closely resemble human tumors. These colon tumors from mice or humans can be grown in culture, creating miniature three-dimensional tumors called organoids.

Subjecting the organoids to different conditions, Yilmaz and Sabatini found that in mice, age-related loss of stem cell function can be reversed by a 24-hour fast. Other studies looked at the type of high-fat diet leading to obesity. Yilmaz determined that a high-fat diet boosted the population of intestinal stem cells and generated even more cells that behaved like stem cells. These stem cells and stem-like cells are more likely to give rise to intestinal tumors.

What’s happening inside

In the microenvironment of the digestive system, the single layer of epithelial cells that line the colon die after only a few days of ferrying nutrients into the bloodstream and lymphatic system.

Stem cells sheltered in protected spaces with fanciful names like the crypts of Lieberkühn generate a hundred grams of new intestinal tissue every day. The source of all the epithelial cells as well as the cells of the villi, a velvety layer of fingerlike projections that line the intestine, stem cells repair and replace tissue continually assaulted by stomach acid, pancreatic enzymes, bile, fats, and bacteria.

Nearby cells guard the stem cells by secreting agents that fight off harmful bacteria, fungi, and viruses and help regulate the composition of the microbiome.

Most of the body’s stem cells, like those deep within bone marrow, are not nearly as prolific as intestinal stem cells, likely because there’s a risk associated with the stem cells’ ability to rapidly replace themselves: mutations.

At the heart of a cell’s behavior is its messenger RNA, or mRNA, the technology used in the Moderna and Pfizer Covid-19 vaccines. These mRNA vaccines teach cells how to make a protein that triggers an immune response to the virus. Each mRNA transcript, a single strand of RNA carrying a specific genetic instruction from the DNA in the nucleus to the cell’s protein-making machinery, determines which protein gets made to help support the cell’s activity.

“From a snapshot of all of the cell’s mRNA, its transcriptome, we can see how it is trying to respond to change,” Shalek says.

Shalek’s tools help him and Yilmaz measure the properties of multiple types of intestinal cells—immune cells, stem cells, and epithelial cells, to name a few—at once to see precisely how these otherwise invisible, minute features collectively orchestrate tissue-wide responses to external signals.

Sequencing a cell’s mRNA makeup requires smashing the cell open and collecting all of its transcripts. Shalek jokingly likened the process to an alien invader beaming human specimens up to a spaceship and investigating what’s happening inside them.

One of the methods Shalek helped develop tags each mRNA within a cell so that it can be traced back to its cell of origin even after it’s been ripped apart. The inexpensive, portable system, called Seq-Well, looks like an ice cube tray. Around the size of a stick of gum, it contains roughly a hundred thousand miniature wells, each approximately 50-by-50-by-50 microns.

Each cell is deposited into its own well, which contains a bead coated with uniquely barcoded DNA molecules; those DNA molecules are designed to latch onto mRNA and ignore the rest of the cell’s components. The wells are sealed and the cells broken apart. The beads are then extracted, processed, and analyzed, providing a record of each cell’s intentions in its last living moments.

The fact that the system can look simultaneously at thousands of individual cells of any type allows Shalek and Yilmaz to check the effect of nutrients on epithelial cells, immune cells, and stem cells all at once.

The Shalek lab is also developing screening tools that are particularly useful for exposing the Yilmaz lab’s organoids to hundreds of nutrients or drugs at one time, potentially reducing the effort needed to identify substances that boost or hinder stem cell function.

Already, Yilmaz and Shalek have used Seq-Well to identify an enzyme that could be a potential future target for a drug that would counter the negative effects of a high-fat diet on intestinal stem cells. More broadly, Yilmaz says, their collaboration is helping develop a very nuanced understanding of a very complex organ.

“Understanding that complexity is what has really driven our collaboration,” Yilmaz says. “Alex has developed the tools that enable us to dissect out individual cell populations and start to understand how environmental factors impact gene expression.”

“Scientists have spent the past 40 years delineating the genetic drivers of colon cancer, and we still have more to learn. But we’ve now entered the era in which we want to understand the impact of environmental and host factors,” Yilmaz says.

Yilmaz hopes to identify nutrients and metabolites that can enhance stem cell function to repair damage after injury, or to identify mechanisms that dampen tumor formation. In addition, biomarkers such as levels of certain substances in the blood could be a key to early intervention, he says.

“Can we identify which obese patients are more prone to developing colon cancer? If so, can we identify therapies that go after weaknesses in their tumors?”

Battling colon cancer

During the time Yilmaz spends at MGH, he looks at slide after slide of biopsied cells. Normal epithelial cells line up in a single, orderly row. After 15 years in medicine, the twisted appearance of diseased cells still shocks him. “You know, in most cases, the number one predictor of how bad a tumor is going to behave isn’t its genetic signature,” he says. “It’s how deep they invade into their organ of origin, whether they have spread to distant organs, and how bad they look under the microscope.” The cells of this patient’s tumor are misshapen, haphazardly stacked on top of each other.

The patient is in her forties. Yilmaz recalled that when he was a resident, colon cancer in a 40-year-old or 30-year-old was a rarity. He now sees such cases almost weekly. Colorectal cancer is among the top three leading causes of cancer-related deaths in the United States, according to the American Cancer Society. It’s expected to cause around 53,000 deaths during 2021. Yilmaz writes up his diagnosis: invasive cancer of the sigmoid colon. The patient’s oncologist will consult with Yilmaz, radiologists, and the surgical team to come up with a treatment plan.

Ultimately, Yilmaz wants to develop strategies to prevent and reduce the growth of tumors in the intestinal tract. The fact that increasingly younger patients are being diagnosed highlights, for him, the importance of diet. “It’s very worrisome,” he says. “We’re at the beginning of a trend where we’re going to see more and more young people afflicted with what can be a fatal disease if not caught early.” Diet could be an important place to start.

He says, “If you can prevent cancer, that’s the best treatment.”

Differences in T cells’ functional state determine resistance to cancer therapy

Researchers decipher when and why immune cells fail to respond to immunotherapy, suggesting that T cells need a different kind of prodding to re-engage the immune response.

Grace van Deelen
October 29, 2021

Non-small cell lung cancer (NSCLC) is the most common type of lung cancer in humans. Some patients with NSCLC receive a therapy called immune checkpoint blockade (ICB) that helps kill cancer cells by reinvigorating a subset of immune cells called T cells, which are “exhausted” and have stopped working. However, only about 35% of NSCLC patients respond to ICB therapy. Stefani Spranger’s lab at the MIT Department of Biology explores the mechanisms behind this resistance, with the goal of inspiring new therapies to better treat NSCLC patients. In a new study published on Oct. 29 in Science Immunology, a team led by Spranger lab postdoc Brendan Horton revealed what causes T cells to be non-responsive to ICB — and suggested a possible solution.

Scientists have long thought that the conditions within a tumor were responsible for determining when T cells stop working and become exhausted after being overstimulated or working for too long to fight a tumor. That’s why physicians prescribe ICB to treat cancer — ICB can invigorate the exhausted T cells within a tumor. However, Horton’s new experiments show that some ICB-resistant T cells stop working before they even enter the tumor. These T cells are not actually exhausted, but rather they become dysfunctional due to changes in gene expression that arise early during the activation of a T cell, which occurs in lymph nodes. Once activated, T cells differentiate into certain functional states, which are distinguishable by their unique gene expression patterns.

In order to determine why some tumors are resistant to ICB, Horton and the research team studied T cells in murine models of NSCLC. The researchers sequenced messenger RNA from the responsive and non-responsive T cells in order to identify any differences between the T cells. Supported in part by the Koch Institute Frontier Research Program, they used a technique called Seq-Well, developed in the lab of fellow Koch Institute member J. Christopher Love, the Raymond A. (1921) and Helen E. St. Laurent Professor of Chemical Engineering and a co-author of the study. The technique allows for the rapid gene expression profiling of single cells, which permitted Spranger and Horton to get a very granular look at the gene expression patterns of the T cells they were studying.

Seq-Well revealed distinct patterns of gene expression between the responsive and non-responsive T cells. These differences, which are determined when the T cells assume their specialized functional states, may be the underlying cause of ICB resistance.

Now that Horton and his colleagues had a possible explanation for why some T cells did not respond to ICB, they decided to see if they could help the ICB-resistant T cells kill the tumor cells. When analyzing the gene expression patterns of the non-responsive T cells, the researchers had noticed that these T cells had a lower expression of receptors for certain cytokines, small proteins that control immune system activity. To counteract this, the researchers treated lung tumors in murine models with extra cytokines. As a result, the previously non-responsive T cells were then able to fight the tumors — meaning that the cytokine therapy prevented, and potentially even reversed, the dysfunctionality.

Administering cytokine therapy to human patients is not currently safe, because cytokines can cause serious side effects as well as a reaction called a “cytokine storm,” which can produce severe fevers, inflammation, fatigue, and nausea. However, there are ongoing efforts to figure out how to safely administer cytokines to specific tumors. In the future, Spranger and Horton suspect that cytokine therapy could be used in combination with ICB.

“This is potentially something that could be translated into a therapeutic that could increase the therapy response rate in non-small cell lung cancer,” Horton says.

Spranger agrees that this work will help researchers develop more innovative cancer therapies, especially because researchers have historically focused on T cell exhaustion rather than the earlier role that T cell functional states might play in cancer.

“If T cells are rendered dysfunctional early on, ICB is not going to be effective, and we need to think outside the box,” she says. “There’s more evidence, and other labs are now showing this as well, that the functional state of the T cell actually matters quite substantially in cancer therapies.” To Spranger, this means that cytokine therapy “might be a therapeutic avenue” for NSCLC patients beyond ICB.

Jeffrey Bluestone, the A.W. and Mary Margaret Clausen Distinguished Professor of Metabolism and Endocrinology at the University of California-San Francisco, who was not involved with the paper, agrees. “The study provides a potential opportunity to ‘rescue’ immunity in the NSCLC non-responder patients with appropriate combination therapies,” he says.

This research was funded by the Pew-Stewart Scholars for Cancer Research, the Ludwig Center for Molecular Oncology, the Koch Institute Frontier Research Program through the Kathy and Curt Mable Cancer Research Fund, and the National Cancer Institute.

How diet affects tumors

A new study finds cutting off cells’ supplies of lipids can slow the growth of tumors in mice.

Anne Trafton | MIT News Office
October 20, 2021

In recent years, there has been some evidence that dietary interventions can help to slow the growth of tumors. A new study from MIT, which analyzed two different diets in mice, reveals how those diets affect cancer cells, and offers an explanation for why restricting calories may slow tumor growth.

The study examined the effects of a calorically restricted diet and a ketogenic diet in mice with pancreatic tumors. While both of these diets reduce the amount of sugar available to tumors, the researchers found that only the calorically restricted diet reduced the availability of fatty acids, and this was linked to a slowdown in tumor growth.

The findings do not suggest that cancer patients should try to follow either of these diets, the researchers say. Instead, they believe the findings warrant further study to determine how dietary interventions might be combined with existing or emerging drugs to help patients with cancer.

“There’s a lot of evidence that diet can affect how fast your cancer progresses, but this is not a cure,” says Matthew Vander Heiden, director of MIT’s Koch Institute for Integrative Cancer Research and the senior author of the study. “While the findings are provocative, further study is needed, and individual patients should talk to their doctor about the right dietary interventions for their cancer.”

MIT postdoc Evan Lien is the lead author of the paper, which appears today in Nature.

Metabolic mechanism

Vander Heiden, who is also a medical oncologist at Dana-Farber Cancer Institute, says his patients often ask him about the potential benefits of various diets, but there is not enough scientific evidence available to offer any definitive advice. Many of the dietary questions that patients have focus on either a calorie-restricted diet, which reduces calorie consumption by 25 to 50 percent, or a ketogenic diet, which is low in carbohydrates and high in fat and protein.

Previous studies have suggested that a calorically restricted diet might slow tumor growth in some contexts, and such a diet has been shown to extend lifespan in mice and many other animal species. A smaller number of studies exploring the effects of a ketogenic diet on cancer have produced inconclusive results.

“A lot of the advice or cultural fads that are out there aren’t necessarily always based on very good science,” Lien says. “It seemed like there was an opportunity, especially with our understanding of cancer metabolism having evolved so much over the past 10 years or so, that we could take some of the biochemical principles that we’ve learned and apply those concepts to understanding this complex question.”

Cancer cells consume a great deal of glucose, so some scientists had hypothesized that either the ketogenic diet or calorie restriction might slow tumor growth by reducing the amount of glucose available. However, the MIT team’s initial experiments in mice with pancreatic tumors showed that calorie restriction has a much greater effect on tumor growth than the ketogenic diet, so the researchers suspected that glucose levels were not playing a major role in the slowdown.

To dig deeper into the mechanism, the researchers analyzed tumor growth and nutrient concentration in mice with pancreatic tumors, which were fed either a normal, ketogenic, or calorie-restricted diet. In both the ketogenic and calorie-restricted mice, glucose levels went down. In the calorie-restricted mice, lipid levels also went down, but in mice on the ketogenic diet, they went up.

Lipid shortages impair tumor growth because cancer cells need lipids to construct their cell membranes. Normally, when lipids aren’t available in a tissue, cells can make their own. As part of this process, they need to maintain the right balance of saturated and unsaturated fatty acids, which requires an enzyme called stearoyl-CoA desaturase (SCD). This enzyme is responsible for converting saturated fatty acids into unsaturated fatty acids.

Both calorie-restricted and ketogenic diets reduce SCD activity, but mice on the ketogenic diet had lipids available to them from their diet, so they didn’t need to use SCD. Mice on the calorie-restricted diet, however, couldn’t get fatty acids from their diet or produce their own. In these mice, tumor growth slowed significantly, compared to mice on the ketogenic diet.

“Not only does caloric restriction starve tumors of lipids, it also impairs the process that allows them to adapt to it. That combination is really contributing to the inhibition of tumor growth,” Lien says.

Dietary effects

In addition to their mouse research, the researchers also looked at some human data. Working with Brian Wolpin, an oncologist at Dana-Farber Cancer Institute and an author of the paper, the team obtained data from a large cohort study that allowed them to analyze the relationship between dietary patterns and survival times in pancreatic cancer patients. From that study, the researchers found that the type of fat consumed appears to influence how patients on a low-sugar diet fare after a pancreatic cancer diagnosis, although the data are not complete enough to draw any conclusions about the effect of diet, the researchers say.

Although this study showed that calorie restriction has beneficial effects in mice, the researchers say they do not recommend that cancer patients follow a calorie-restricted diet, which is difficult to maintain and can have harmful side effects. However, they believe that cancer cells’ dependence on the availability of unsaturated fatty acids could be exploited to develop drugs that might help slow tumor growth.

One possible therapeutic strategy could be inhibition of the SCD enzyme, which would cut off tumor cells’ ability to produce unsaturated fatty acids.

“The purpose of these studies isn’t necessarily to recommend a diet, but it’s to really understand the underlying biology,” Lien says. “They provide some sense of the mechanisms of how these diets work, and that can lead to rational ideas on how we might mimic those situations for cancer therapy.”

The researchers now plan to study how diets with a variety of fat sources — including plant or animal-based fats with defined differences in saturated, monounsaturated, and polyunsaturated fatty acid content — alter tumor fatty acid metabolism and the ratio of unsaturated to saturated fatty acids.

The research was funded by the Damon Runyon Cancer Research Foundation, the National Institutes of Health, the Lustgarten Foundation, the Dana-Farber Cancer Institute Hale Family Center for Pancreatic Cancer Research, Stand Up to Cancer, the Pancreatic Cancer Action Network, the Noble Effort Fund, the Wexler Family Fund, Promises for Purple, the Bob Parsons Fund, the Emerald Foundation, the Howard Hughes Medical Institute, the MIT Center for Precision Cancer Medicine, and the Ludwig Center at MIT.