Research Area: Cancer Biology

A cysteine-rich diet may promote regeneration of the intestinal lining, study suggests

The findings from the Yilmaz Lab recently published in Nature, may offer a new way to help heal tissue damage from radiation or chemotherapy treatment.

Anne Trafton | MIT News
October 1, 2025

A diet rich in the amino acid cysteine may have rejuvenating effects in the small intestine, according to a new study from MIT. This amino acid, the researchers discovered, can turn on an immune signaling pathway that helps stem cells to regrow new intestinal tissue.

This enhanced regeneration may help to heal injuries from radiation, which often occur in patients undergoing radiation therapy for cancer. The research was conducted in mice, but if future research shows similar results in humans, then delivering elevated quantities of cysteine, through diet or supplements, could offer a new strategy to help damaged tissue heal faster, the researchers say.

“The study suggests that if we give these patients a cysteine-rich diet or cysteine supplementation, perhaps we can dampen some of the chemotherapy or radiation-induced injury,” says Omer Yilmaz, director of the MIT Stem Cell Initiative, an associate professor of biology at MIT, and a member of MIT’s Koch Institute for Integrative Cancer Research. “The beauty here is we’re not using a synthetic molecule; we’re exploiting a natural dietary compound.”

While previous research has shown that certain types of diets, including low-calorie diets, can enhance intestinal stem cell activity, the new study is the first to identify a single nutrient that can help intestinal cells to regenerate.

Yilmaz is the senior author of the study, which appears today in Nature. Koch Institute postdoc Fangtao Chi is the paper’s lead author.

Boosting regeneration

It is well-established that diet can affect overall health: High-fat diets can lead to obesity, diabetes, and other health problems, while low-calorie diets have been shown to extend lifespans in many species. In recent years, Yilmaz’s lab has investigated how different types of diets influence stem cell regeneration, and found that high-fat diets, as well as short periods of fasting, can enhance stem cell activity in different ways.

“We know that macro diets such as high-sugar diets, high-fat diets, and low-calorie diets have a clear impact on health. But at the granular level, we know much less about how individual nutrients impact stem cell fate decisions, as well as tissue function and overall tissue health,” Yilmaz says.

In their new study, the researchers began by feeding mice a diet high in one of 20 different amino acids, the building blocks of proteins. For each group, they measured how the diet affected intestinal stem cell regeneration. Among these amino acids, cysteine had the most dramatic effects on stem cells and progenitor cells (immature cells that differentiate into adult intestinal cells).

Further studies revealed that cysteine initiates a chain of events leading to the activation of a population of immune cells called CD8 T cells. When cells in the lining of the intestine absorb cysteine from digested food, they convert it into CoA, a cofactor that is released into the mucosal lining of the intestine. There, CD8 T cells absorb CoA, which stimulates them to begin proliferating and producing a cytokine called IL-22.

IL-22 is an important player in the regulation of intestinal stem cell regeneration, but until now, it wasn’t known that CD8 T cells can produce it to boost intestinal stem cells. Once activated, those IL-22-releasing T cells are primed to help combat any kind of injury that could occur within the intestinal lining.

“What’s really exciting here is that feeding mice a cysteine-rich diet leads to the expansion of an immune cell population that we typically don’t associate with IL-22 production and the regulation of intestinal stemness,” Yilmaz says. “What happens in a cysteine-rich diet is that the pool of cells that make IL-22 increases, particularly the CD8 T-cell fraction.”

These T cells tend to congregate within the lining of the intestine, so they are already in position when needed. The researchers found that the stimulation of CD8 T cells occurred primarily in the small intestine, not in any other part of the digestive tract, which they believe is because most of the protein that we consume is absorbed by the small intestine.

Healing the intestine

In this study, the researchers showed that regeneration stimulated by a cysteine-rich diet could help to repair radiation damage to the intestinal lining. Also, in work that has not been published yet, they showed that a high-cysteine diet had a regenerative effect following treatment with a chemotherapy drug called 5-fluorouracil. This drug, which is used to treat colon and pancreatic cancers, can also damage the intestinal lining.

Cysteine is found in many high-protein foods, including meat, dairy products, legumes, and nuts. The body can also synthesize its own cysteine, by converting the amino acid methionine to cysteine — a process that takes place in the liver. However, cysteine produced in the liver is distributed through the entire body and doesn’t lead to a buildup in the small intestine the way that consuming cysteine in the diet does.

“With our high-cysteine diet, the gut is the first place that sees a high amount of cysteine,” Chi says.

Cysteine has been previously shown to have antioxidant effects, which are also beneficial, but this study is the first to demonstrate its effect on intestinal stem cell regeneration. The researchers now hope to study whether it may also help other types of stem cells regenerate new tissues. In one ongoing study, they are investigating whether cysteine might stimulate hair follicle regeneration.

They also plan to further investigate some of the other amino acids that appear to influence stem cell regeneration.

“I think we’re going to uncover multiple new mechanisms for how these amino acids regulate cell fate decisions and gut health in the small intestine and colon,” Yilmaz says.

The research was funded, in part, by the National Institutes of Health, the V Foundation, the Koch Institute Frontier Research Program via the Kathy and Curt Marble Cancer Research Fund, the Bridge Project — a partnership between the Koch Institute for Integrative Cancer Research at MIT and the Dana-Farber/Harvard Cancer Center, the American Federation for Aging Research, the MIT Stem Cell Initiative, and the Koch Institute Support (core) Grant from the National Cancer Institute.

A more precise way to edit the genome

MIT researchers have dramatically lowered the error rate of prime editing, a technique that holds potential for treating many genetic disorders.

Anne Trafton | MIT News
September 17, 2025

A genome-editing technique known as prime editing holds potential for treating many diseases by transforming faulty genes into functional ones. However, the process carries a small chance of inserting errors that could be harmful.

MIT researchers have now found a way to dramatically lower the error rate of prime editing, using modified versions of the proteins involved in the process. This advance could make it easier to develop gene therapy treatments for a variety of diseases, the researchers say.

“This paper outlines a new approach to doing gene editing that doesn’t complicate the delivery system and doesn’t add additional steps, but results in a much more precise edit with fewer unwanted mutations,” says Phillip Sharp, an MIT Institute Professor Emeritus, a member of MIT’s Koch Institute for Integrative Cancer Research, and one of the senior authors of the new study.

With their new strategy, the MIT team was able to improve the error rate of prime editors from about one error in seven edits to one in 101 for the most-used editing mode, or from one error in 122 edits to one in 543 for a high-precision mode.

“For any drug, what you want is something that is effective, but with as few side effects as possible,” says Robert Langer, the David H. Koch Institute Professor at MIT, a member of the Koch Institute, and one of the senior authors of the new study. “For any disease where you might do genome editing, I would think this would ultimately be a safer, better way of doing it.”

Koch Institute research scientist Vikash Chauhan is the lead author of the paper, which appears today in Nature.

The potential for error

The earliest forms of gene therapy, first tested in the 1990s, involved delivering new genes carried by viruses. Subsequently, gene-editing techniques that use enzymes such as zinc finger nucleases to correct genes were developed. These nucleases are difficult to engineer, however, so adapting them to target different DNA sequences is a very laborious process.

Many years later, the CRISPR genome-editing system was discovered in bacteria, offering scientists a potentially much easier way to edit the genome. The CRISPR system consists of an enzyme called Cas9 that can cut double-stranded DNA at a particular spot, along with a guide RNA that tells Cas9 where to cut. Researchers have adapted this approach to cut out faulty gene sequences or to insert new ones, following an RNA template.

In 2019, researchers at the Broad Institute of MIT and Harvard reported the development of prime editing: a new system, based on CRISPR, that is more precise and has fewer off-target effects. A recent study reported that prime editors were successfully used to treat a patient with chronic granulomatous disease (CGD), a rare genetic disease that affects white blood cells.

“In principle, this technology could eventually be used to address many hundreds of genetic diseases by correcting small mutations directly in cells and tissues,” Chauhan says.

One of the advantages of prime editing is that it doesn’t require making a double-stranded cut in the target DNA. Instead, it uses a modified version of Cas9 that cuts just one of the complementary strands, opening up a flap where a new sequence can be inserted. A guide RNA delivered along with the prime editor serves as the template for the new sequence.

Once the new sequence has been copied, however, it must compete with the old DNA strand to be incorporated into the genome. If the old strand outcompetes the new one, the extra flap of new DNA hanging off may accidentally get incorporated somewhere else, giving rise to errors.

Many of these errors might be relatively harmless, but it’s possible that some could eventually lead to tumor development or other complications. With the most recent version of prime editors, this error rate ranges from one per seven edits to one per 121 edits for different editing modes.

“The technologies we have now are really a lot better than earlier gene therapy tools, but there’s always a chance for these unintended consequences,” Chauhan says.

Precise editing

To reduce those error rates, the MIT team decided to take advantage of a phenomenon they had observed in a 2023 study. In that paper, they found that while Cas9 usually cuts in the same DNA location every time, some mutated versions of the protein show a relaxation of those constraints. Instead of always cutting the same location, those Cas9 proteins would sometimes make their cut one or two bases further along the DNA sequence.

This relaxation, the researchers discovered, makes the old DNA strands less stable, so they get degraded, making it easier for the new strands to be incorporated without introducing any errors.

In the new study, the researchers were able to identify Cas9 mutations that dropped the error rate to 1/20th its original value. Then, by combining pairs of those mutations, they created a Cas9 editor that lowered the error rate even further, to 1/36th the original amount.

To make the editors even more accurate, the researchers incorporated their new Cas9 proteins into a prime editing system that has an RNA binding protein that stabilizes the ends of the RNA template more efficiently. This final editor, which the researchers call vPE, had an error rate just 1/60th of the original, ranging from one in 101 edits to one in 543 edits for different editing modes. These tests were performed in mouse and human cells.

The MIT team is now working on further improving the efficiency of prime editors, through further modifications of Cas9 and the RNA template. They are also working on ways to deliver the editors to specific tissues of the body, which is a longstanding challenge in gene therapy.

They also hope that other labs will begin using the new prime editing approach in their research studies. Prime editors are commonly used to explore many different questions, including how tissues develop, how populations of cancer cells evolve, and how cells respond to drug treatment.

“Genome editors are used extensively in research labs,” Chauhan says. “So the therapeutic aspect is exciting, but we are really excited to see how people start to integrate our editors into their research workflows.”

The research was funded by the Life Sciences Research Foundation, the National Institute of Biomedical Imaging and Bioengineering, the National Cancer Institute, and the Koch Institute Support (core) Grant from the National Cancer Institute.

Inflammation jolts “sleeping” cancer cells awake, enabling them to multiply again

A paper from the Weinberg Lab indicates that inflammation may be a factor in how metastatic cancer cells, those that have broken away from the original tumor, can erupt into a frenzy of growth and division months, years, or decades after initial treatment, seeding new, life-threatening tumors.

Shafaq Zia | Whitehead Institute
September 3, 2025

This migration of cancer cells, called metastasis, is especially common in breast cancer. For many patients, the disease can return months—or even decades—after initial treatment, this time in an entirely different organ.

Whitehead Institute Founding Member Robert Weinberg, also the Daniel K. Ludwig Professor for Cancer Research at Massachusetts Institute of Technology (MIT), has spent decades unraveling the complex biology of metastasis and pursuing research that could improve survival rates among patients with metastatic breast cancer—or prevent metastasis altogether.

In their latest study, Weinberg, postdoctoral fellow Jingwei Zhang, and colleagues ask a critical question: what causes these dormant cancer cells to erupt into a frenzy of growth and division? The group’s findings, published Sept. 1 in The Proceedings of the National Academy of Sciences (PNAS), point to a unique culprit.

This awakening of dormant cancer cells, they’ve discovered, isn’t a spontaneous process. Instead, the wake-up call comes from the inflamed tissue surrounding the cells. One trigger for this inflammation is bleomycin, a common chemotherapy drug that can scar and thicken lung tissue.

“The inflammation jolts the dormant cancer cells awake,” Weinberg says. “Once awakened, they start multiplying again, seeding new life-threatening tumors in the body.”

Decoding metastasis

There’s a lot that scientists still don’t know about metastasis, but this much is clear: cancer cells must undergo a long and arduous journey to achieve it. The first step is to break away from their neighbors within the original tumor.

Normally, cells stick to one another using surface proteins that act as molecular “velcro” but some cancer cells can acquire genetic changes that disrupt the production of these proteins and make them more mobile and invasive, allowing them to detach from the parent tumor.

Once detached, they can penetrate blood vessels and lymphatic channels, which act as highways to distant organs.

While most cancer cells die at some point during this journey, a few persist. These cells exit the bloodstream and invade different tissues—lungs, liver, bone, and even the brain—to give birth to new, often more aggressive tumors.

“Almost 90% of cancer-related deaths occur not from the original tumor but when cancer cells spread to other parts of the body,” says Weinberg. “This is why it’s so important to understand how these ‘sleeping’ cancer cells can wake up and start growing again.”

Setting up shop in new tissue comes with changes in surroundings—the “tumor microenvironment”—to which the cancer cells may not be well-suited. These cells face constant threats, including detection and attack by the immune system.

To survive, they often enter a protective state of dormancy that puts a pause on growth and division. This dormant state also makes them resistant to conventional cancer treatments, which often target rapidly dividing cells.

To investigate what makes this dormancy reversible months or years down the line, researchers in the Weinberg Lab injected human breast cancer cells into mice. These cancer cells were modified to produce a fluorescent protein, allowing the scientists to track their behavior in the body.

The group then focused on cancer cells that had lodged themselves in the lung tissue. By examining them for specific proteins—Ki67, ITGB4 and p63—that act as markers of cell activity and state, the researchers were able to confirm that these cells were in a non-dividing, dormant state.

Previous work from the Weinberg Lab had shown that inflammation in organ tissue can provoke dormant breast cancer cells to start growing again. In this study, the team tested bleomycin—a chemotherapy drug known to cause lung inflammation—that can be given to patients after surgery to lower the risk of cancer recurrence.

The researchers found that lung inflammation from bleomycin was sufficient to trigger the growth of large lung cancer colonies in treated mice—and to shift the character of these once dormant cells to those that are more invasive and mobile.

Zeroing in on the tumor microenvironment, the team identified a type of immune cells, called M2 macrophages, as drivers of this process. These macrophages release molecules called epidermal growth factor receptor (EGFR) ligands, which bind to receptors on the surface of dormant cancer cells. This activates a cascade of signals that provoke dormant cancer cells to start multiplying rapidly.

But EGFR signaling is only the initial spark that ignites the fire. “We found that once dormant cancer cells are awakened, they retain what we call an ‘awakening memory,’” Zhang says. “They no longer require ongoing inflammatory signals from the microenvironment to stay active [growing and multiplying]—they remember the awakened state.”

While signals related to inflammation are necessary to awaken dormant cancer cells, exactly how much signaling is needed remains unclear. “This aspect of cancer biology is particularly challenging because multiple signals contribute to the state change in these dormant cells,” Zhang says.

The team has already identified one key player in the awakening process but understanding the full set of signals and how each contributes is far more complex—a question they are continuing to investigate in their new work.

Studying these pivotal changes in the lives of cancer cells—such as their transition from dormancy to active growth—will deepen our scientific understanding of metastasis and, as researchers in the Weinberg Lab hope, lead to more effective treatments for patients with metastatic cancers.

Biologists identify targets for new pancreatic cancer treatments

Research from MIT and Dana-Farber Cancer Institute yielded hundreds of “cryptic” peptides that are found only on pancreatic tumor cells and could be targeted by vaccines or engineered T cells.

Anne Trafton | MIT News
May 7, 2025

Researchers from MIT and Dana-Farber Cancer Institute have discovered that a class of peptides expressed in pancreatic cancer cells could be a promising target for T-cell therapies and other approaches that attack pancreatic tumors.

Known as cryptic peptides, these molecules are produced from sequences in the genome that were not thought to encode proteins. Such peptides can also be found in some healthy cells, but in this study, the researchers identified about 500 that appear to be found only in pancreatic tumors.

The researchers also showed they could generate T cells targeting those peptides. Those T cells were able to attack pancreatic tumor organoids derived from patient cells, and they significantly slowed down tumor growth in a study of mice.

“Pancreas cancer is one of the most challenging cancers to treat. This study identifies an unexpected vulnerability in pancreas cancer cells that we may be able to exploit therapeutically,” says Tyler Jacks, the David H. Koch Professor of Biology at MIT and a member of the Koch Institute for Integrative Cancer Research.

Jacks and William Freed-Pastor, a physician-scientist in the Hale Family Center for Pancreatic Cancer Research at Dana-Farber Cancer Institute and an assistant professor at Harvard Medical School, are the senior authors of the study, which appears today in Science. Zackery Ely PhD ’22 and Zachary Kulstad, a former research technician at Dana-Farber Cancer Institute and the Koch Institute, are the lead authors of the paper.

Cryptic peptides

Pancreatic cancer has one of the lowest survival rates of any cancer — about 10 percent of patients survive for five years after their diagnosis.

Most pancreatic cancer patients receive a combination of surgery, radiation treatment, and chemotherapy. Immunotherapy treatments such as checkpoint blockade inhibitors, which are designed to help stimulate the body’s own T cells to attack tumor cells, are usually not effective against pancreatic tumors. However, therapies that deploy T cells engineered to attack tumors have shown promise in clinical trials.

These therapies involve programming the T-cell receptor (TCR) of T cells to recognize a specific peptide, or antigen, found on tumor cells. There are many efforts underway to identify the most effective targets, and researchers have found some promising antigens that consist of mutated proteins that often show up when pancreatic cancer genomes are sequenced.

In the new study, the MIT and Dana-Farber team wanted to extend that search into tissue samples from patients with pancreatic cancer, using immunopeptidomics — a strategy that involves extracting the peptides presented on a cell surface and then identifying the peptides using mass spectrometry.

Using tumor samples from about a dozen patients, the researchers created organoids — three-dimensional growths that partially replicate the structure of the pancreas. The immunopeptidomics analysis, which was led by Jennifer Abelin and Steven Carr at the Broad Institute, found that the majority of novel antigens found in the tumor organoids were cryptic antigens. Cryptic peptides have been seen in other types of tumors, but this is the first time they have been found in pancreatic tumors.

Each tumor expressed an average of about 250 cryptic peptides, and in total, the researchers identified about 1,700 cryptic peptides.

“Once we started getting the data back, it just became clear that this was by far the most abundant novel class of antigens, and so that’s what we wound up focusing on,” Ely says.

The researchers then performed an analysis of healthy tissues to see if any of these cryptic peptides were found in normal cells. They found that about two-thirds of them were also found in at least one type of healthy tissue, leaving about 500 that appeared to be restricted to pancreatic cancer cells.

“Those are the ones that we think could be very good targets for future immunotherapies,” Freed-Pastor says.

Programmed T cells

To test whether these antigens might hold potential as targets for T-cell-based treatments, the researchers exposed about 30 of the cancer-specific antigens to immature T cells and found that 12 of them could generate large populations of T cells targeting those antigens.

The researchers then engineered a new population of T cells to express those T-cell receptors. These engineered T cells were able to destroy organoids grown from patient-derived pancreatic tumor cells. Additionally, when the researchers implanted the organoids into mice and then treated them with the engineered T cells, tumor growth was significantly slowed.

This is the first time that anyone has demonstrated the use of T cells targeting cryptic peptides to kill pancreatic tumor cells. Even though the tumors were not completely eradicated, the results are promising, and it is possible that the T-cells’ killing power could be strengthened in future work, the researchers say.

Freed-Pastor’s lab is also beginning to work on a vaccine targeting some of the cryptic antigens, which could help stimulate patients’ T cells to attack tumors expressing those antigens. Such a vaccine could include a collection of the antigens identified in this study, including those frequently found in multiple patients.

This study could also help researchers in designing other types of therapy, such as T cell engagers — antibodies that bind an antigen on one side and T cells on the other, which allows them to redirect any T cell to kill tumor cells.

Any potential vaccine or T cell therapy is likely a few years away from being tested in patients, the researchers say.

The research was funded in part by the Hale Family Center for Pancreatic Cancer Research, the Lustgarten Foundation, Stand Up To Cancer, the Pancreatic Cancer Action Network, the Burroughs Wellcome Fund, a Conquer Cancer Young Investigator Award, the National Institutes of Health, and the National Cancer Institute.

New study reveals how cleft lip and cleft palate can arise

MIT biologists have found that defects in some transfer RNA molecules can lead to the formation of these common conditions.

Anne Trafton | MIT News
April 17, 2025

Cleft lip and cleft palate are among the most common birth defects, occurring in about one in 1,050 births in the United States. These defects, which appear when the tissues that form the lip or the roof of the mouth do not join completely, are believed to be caused by a mix of genetic and environmental factors.

In a new study, MIT biologists have discovered how a genetic variant often found in people with these facial malformations leads to the development of cleft lip and cleft palate.

Their findings suggest that the variant diminishes cells’ supply of transfer RNA, a molecule that is critical for assembling proteins. When this happens, embryonic face cells are unable to fuse to form the lip and roof of the mouth.

“Until now, no one had made the connection that we made. This particular gene was known to be part of the complex involved in the splicing of transfer RNA, but it wasn’t clear that it played such a crucial role for this process and for facial development. Without the gene, known as DDX1, certain transfer RNA can no longer bring amino acids to the ribosome to make new proteins. If the cells can’t process these tRNAs properly, then the ribosomes can’t make protein anymore,” says Michaela Bartusel, an MIT research scientist and the lead author of the study.

Eliezer Calo, an associate professor of biology at MIT, is the senior author of the paper, which appears today in the American Journal of Human Genetics.

Genetic variants

Cleft lip and cleft palate, also known as orofacial clefts, can be caused by genetic mutations, but in many cases, there is no known genetic cause.

“The mechanism for the development of these orofacial clefts is unclear, mostly because they are known to be impacted by both genetic and environmental factors,” Calo says. “Trying to pinpoint what might be affected has been very challenging in this context.”

To discover genetic factors that influence a particular disease, scientists often perform genome-wide association studies (GWAS), which can reveal variants that are found more often in people who have a particular disease than in people who don’t.

For orofacial clefts, some of the genetic variants that have regularly turned up in GWAS appeared to be in a region of DNA that doesn’t code for proteins. In this study, the MIT team set out to figure out how variants in this region might influence the development of facial malformations.

Their studies revealed that these variants are located in an enhancer region called e2p24.2. Enhancers are segments of DNA that interact with protein-coding genes, helping to activate them by binding to transcription factors that turn on gene expression.

The researchers found that this region is in close proximity to three genes, suggesting that it may control the expression of those genes. One of those genes had already been ruled out as contributing to facial malformations, and another had already been shown to have a connection. In this study, the researchers focused on the third gene, which is known as DDX1.

DDX1, it turned out, is necessary for splicing transfer RNA (tRNA) molecules, which play a critical role in protein synthesis. Each transfer RNA molecule transports a specific amino acid to the ribosome — a cell structure that strings amino acids together to form proteins, based on the instructions carried by messenger RNA.

While there are about 400 different tRNAs found in the human genome, only a fraction of those tRNAs require splicing, and those are the tRNAs most affected by the loss of DDX1. These tRNAs transport four different amino acids, and the researchers hypothesize that these four amino acids may be particularly abundant in proteins that embryonic cells that form the face need to develop properly.

When the ribosomes need one of those four amino acids, but none of them are available, the ribosome can stall, and the protein doesn’t get made.

The researchers are now exploring which proteins might be most affected by the loss of those amino acids. They also plan to investigate what happens inside cells when the ribosomes stall, in hopes of identifying a stress signal that could potentially be blocked and help cells survive.

Malfunctioning tRNA

While this is the first study to link tRNA to craniofacial malformations, previous studies have shown that mutations that impair ribosome formation can also lead to similar defects. Studies have also shown that disruptions of tRNA synthesis — caused by mutations in the enzymes that attach amino acids to tRNA, or in proteins involved in an earlier step in tRNA splicing — can lead to neurodevelopmental disorders.

“Defects in other components of the tRNA pathway have been shown to be associated with neurodevelopmental disease,” Calo says. “One interesting parallel between these two is that the cells that form the face are coming from the same place as the cells that form the neurons, so it seems that these particular cells are very susceptible to tRNA defects.”

The researchers now hope to explore whether environmental factors linked to orofacial birth defects also influence tRNA function. Some of their preliminary work has found that oxidative stress — a buildup of harmful free radicals — can lead to fragmentation of tRNA molecules. Oxidative stress can occur in embryonic cells upon exposure to ethanol, as in fetal alcohol syndrome, or if the mother develops gestational diabetes.

“I think it is worth looking for mutations that might be causing this on the genetic side of things, but then also in the future, we would expand this into which environmental factors have the same effects on tRNA function, and then see which precautions might be able to prevent any effects on tRNAs,” Bartusel says.

The research was funded by the National Science Foundation Graduate Research Program, the National Cancer Institute, the National Institute of General Medical Sciences, and the Pew Charitable Trusts.

Restoring healthy gene expression with programmable therapeutics

CAMP4 Therapeutics is targeting regulatory RNA, whose role in gene expression was first described by co-founder and MIT Professor Richard Young.

Zach Winn | MIT News
April 16, 2025

Many diseases are caused by dysfunctional gene expression that leads to too much or too little of a given protein. Efforts to cure those diseases include everything from editing genes to inserting new genetic snippets into cells to injecting the missing proteins directly into patients.

CAMP4 is taking a different approach. The company is targeting a lesser-known player in the regulation of gene expression known as regulatory RNA. CAMP4 co-founder and MIT Professor Richard Young has shown that by interacting with molecules called transcription factors, regulatory RNA plays an important role in controlling how genes are expressed. CAMP4’s therapeutics target regulatory RNA to increase the production of proteins and put patients’ levels back into healthy ranges.

The company’s approach holds promise for treating diseases caused by defects in gene expression, such as metabolic diseases, heart conditions, and neurological disorders. Targeting regulatory RNAs as opposed to genes could also offer more precise treatments than existing approaches.

“If I just want to fix a single gene’s defective protein output, I don’t want to introduce something that makes that protein at high, uncontrolled amounts,” says Young, who is also a core member of the Whitehead Institute. “That’s a huge advantage of our approach: It’s more like a correction than sledgehammer.”

CAMP4’s lead drug candidate targets urea cycle disorders (UCDs), a class of chronic conditions caused by a genetic defect that limits the body’s ability to metabolize and excrete ammonia. A phase 1 clinical trial has shown CAMP4’s treatment is safe and tolerable for humans, and in preclinical studies the company has shown its approach can be used to target specific regulatory RNA in the cells of humans with UCDs to restore gene expression to healthy levels.

“This has the potential to treat very severe symptoms associated with UCDs,” says Young, who co-founded CAMP4 with cancer genetics expert Leonard Zon, a professor at Harvard Medical School. “These diseases can be very damaging to tissues and causes a lot of pain and distress. Even a small effect in gene expression could have a huge benefit to patients, who are generally young.”

Mapping out new therapeutics

Young, who has been a professor at MIT since 1984, has spent decades studying how genes are regulated. It’s long been known that molecules called transcription factors, which orchestrate gene expression, bind to DNA and proteins. Research published in Young’s lab uncovered a previously unknown way in which transcription factors can also bind to RNA. The finding indicated RNA plays an underappreciated role in controlling gene expression.

CAMP4 was founded in 2016 with the initial idea of mapping out the signaling pathways that govern the expression of genes linked to various diseases. But as Young’s lab discovered and then began to characterize the role of regulatory RNA in gene expression around 2020, the company pivoted to focus on targeting regulatory RNA using therapeutic molecules known as antisense oligonucleotides (ASOs), which have been used for years to target specific messenger RNA sequences.

CAMP4 began mapping the active regulatory RNAs associated with the expression of every protein-coding gene and built a database, which it calls its RAP Platform, that helps it quickly identify regulatory RNAs to target  specific diseases and select ASOs that will most effectively bind to those RNAs.

Today, CAMP4 is using its platform to develop therapeutic candidates it believes can restore healthy protein levels to patients.

“The company has always been focused on modulating gene expression,” says CAMP4 Chief Financial Officer Kelly Gold MBA ’09. “At the simplest level, the foundation of many diseases is too much or too little of something being produced by the body. That is what our approach aims to correct.”

Accelerating impact

CAMP4 is starting by going after diseases of the liver and the central nervous system, where the safety and efficacy of ASOs has already been proven. Young believes correcting genetic expression without modulating the genes themselves will be a powerful approach to treating a range of complex diseases.

“Genetics is a powerful indicator of where a deficiency lies and how you might reverse that problem,” Young says. “There are many syndromes where we don’t have a complete understanding of the underlying mechanism of disease. But when a mutation clearly affects the output of a gene, you can now make a drug that can treat the disease without that complete understanding.”

As the company continues mapping the regulatory RNAs associated with every gene, Gold hopes CAMP4 can eventually minimize its reliance on wet-lab work and lean more heavily on machine learning to leverage its growing database and quickly identify regRNA targets for every disease it wants to treat.

In addition to its trials in urea cycle disorders, the company plans to launch key preclinical safety studies for a candidate targeting seizure disorders with a genetic basis, this year. And as the company continues exploring drug development efforts around the thousands of genetic diseases where increasing protein levels are can have a meaningful impact, it’s also considering collaborating with others to accelerate its impact.

“I can conceive of companies using a platform like this to go after many targets, where partners fund the clinical trials and use CAMP4 as an engine to target any disease where there’s a suspicion that gene upregulation or downregulation is the way to go,” Young says.

Helping the immune system attack tumors

Stefani Spranger is working to discover why some cancers don’t respond to immunotherapy, in hopes of making them more vulnerable to it.

Anne Trafton | MIT News
February 26, 2025

In addition to patrolling the body for foreign invaders, the immune system also hunts down and destroys cells that have become cancerous or precancerous. However, some cancer cells end up evading this surveillance and growing into tumors.

Once established, tumor cells often send out immunosuppressive signals, which leads T cells to become “exhausted” and unable to attack the tumor. In recent years, some cancer immunotherapy drugs have shown great success in rejuvenating those T cells so they can begin attacking tumors again.

While this approach has proven effective against cancers such as melanoma, it doesn’t work as well for others, including lung and ovarian cancer. MIT Associate Professor Stefani Spranger is trying to figure out how those tumors are able to suppress immune responses, in hopes of finding new ways to galvanize T cells into attacking them.

“We really want to understand why our immune system fails to recognize cancer,” Spranger says. “And I’m most excited about the really hard-to-treat cancers because I think that’s where we can make the biggest leaps.”

Her work has led to a better understanding of the factors that control T-cell responses to tumors, and raised the possibility of improving those responses through vaccination or treatment with immune-stimulating molecules called cytokines.

“We’re working on understanding what exactly the problem is, and then collaborating with engineers to find a good solution,” she says.

Jumpstarting T cells

As a student in Germany, where students often have to choose their college major while still in high school, Spranger envisioned going into the pharmaceutical industry and chose to major in biology. At Ludwig Maximilian University in Munich, her course of study began with classical biology subjects such as botany and zoology, and she began to doubt her choice. But, once she began taking courses in cell biology and immunology, her interest was revived and she continued into a biology graduate program at the university.

During a paper discussion class early in her graduate school program, Spranger was assigned to a Science paper on a promising new immunotherapy treatment for melanoma. This strategy involves isolating tumor-infiltrating T-cells during surgery, growing them into large numbers, and then returning them to the patient. For more than 50 percent of those patients, the tumors were completely eliminated.

“To me, that changed the world,” Spranger recalls. “You can take the patient’s own immune system, not really do all that much to it, and then the cancer goes away.”

Spranger completed her PhD studies in a lab that worked on further developing that approach, known as adoptive T-cell transfer therapy. At that point, she still was leaning toward going into pharma, but after finishing her PhD in 2011, her husband, also a biologist, convinced her that they should both apply for postdoc positions in the United States.

They ended up at the University of Chicago, where Spranger worked in a lab that studies how the immune system responds to tumors. There, she discovered that while melanoma is usually very responsive to immunotherapy, there is a small fraction of melanoma patients whose T cells don’t respond to the therapy at all. That got her interested in trying to figure out why the immune system doesn’t always respond to cancer the way that it should, and in finding ways to jumpstart it.

During her postdoc, Spranger also discovered that she enjoyed mentoring students, which she hadn’t done as a graduate student in Germany. That experience drew her away from going into the pharmaceutical industry, in favor of a career in academia.

“I had my first mentoring teaching experience having an undergrad in the lab, and seeing that person grow as a scientist, from barely asking questions to running full experiments and coming up with hypotheses, changed how I approached science and my view of what academia should be for,” she says.

Modeling the immune system

When applying for faculty jobs, Spranger was drawn to MIT by the collaborative environment of MIT and its Koch Institute for Integrative Cancer Research, which offered the chance to collaborate with a large community of engineers who work in the field of immunology.

“That community is so vibrant, and it’s amazing to be a part of it,” she says.

Building on the research she had done as a postdoc, Spranger wanted to explore why some tumors respond well to immunotherapy, while others do not. For many of her early studies, she used a mouse model of non-small-cell lung cancer. In human patients, the majority of these tumors do not respond well to immunotherapy.

“We build model systems that resemble each of the different subsets of non-responsive non-small cell lung cancer, and we’re trying to really drill down to the mechanism of why the immune system is not appropriately responding,” she says.

As part of that work, she has investigated why the immune system behaves differently in different types of tissue. While immunotherapy drugs called checkpoint inhibitors can stimulate a strong T-cell response in the skin, they don’t do nearly as much in the lung. However, Spranger has shown that T cell responses in the lung can be improved when immune molecules called cytokines are also given along with the checkpoint inhibitor.

Those cytokines work, in part, by activating dendritic cells — a class of immune cells that help to initiate immune responses, including activation of T cells.

“Dendritic cells are the conductor for the orchestra of all the T cells, although they’re a very sparse cell population,” Spranger says. “They can communicate which type of danger they sense from stressed cells and then instruct the T cells on what they have to do and where they have to go.”

Spranger’s lab is now beginning to study other types of tumors that don’t respond at all to immunotherapy, including ovarian cancer and glioblastoma. Both the brain and the peritoneal cavity appear to suppress T-cell responses to tumors, and Spranger hopes to figure out how to overcome that immunosuppression.

“We’re specifically focusing on ovarian cancer and glioblastoma, because nothing’s working right now for those cancers,” she says. “We want to understand what we have to do in those sites to induce a really good anti-tumor immune response.”

AI model deciphers the code in proteins that tells them where to go

Whitehead Institute and CSAIL researchers created a machine-learning model to predict and generate protein localization, with implications for understanding and remedying disease.

Greta Friar | Whitehead Institute
February 13, 2025

Proteins are the workhorses that keep our cells running, and there are many thousands of types of proteins in our cells, each performing a specialized function. Researchers have long known that the structure of a protein determines what it can do. More recently, researchers are coming to appreciate that a protein’s localization is also critical for its function. Cells are full of compartments that help to organize their many denizens. Along with the well-known organelles that adorn the pages of biology textbooks, these spaces also include a variety of dynamic, membrane-less compartments that concentrate certain molecules together to perform shared functions. Knowing where a given protein localizes, and who it co-localizes with, can therefore be useful for better understanding that protein and its role in the healthy or diseased cell, but researchers have lacked a systematic way to predict this information.

Meanwhile, protein structure has been studied for over half-a-century, culminating in the artificial intelligence tool AlphaFold, which can predict protein structure from a protein’s amino acid code, the linear string of building blocks within it that folds to create its structure. AlphaFold and models like it have become widely used tools in research.

Proteins also contain regions of amino acids that do not fold into a fixed structure, but are instead important for helping proteins join dynamic compartments in the cell. MIT Professor Richard Young and colleagues wondered whether the code in those regions could be used to predict protein localization in the same way that other regions are used to predict structure. Other researchers have discovered some protein sequences that code for protein localization, and some have begun developing predictive models for protein localization. However, researchers did not know whether a protein’s localization to any dynamic compartment could be predicted based on its sequence, nor did they have a comparable tool to AlphaFold for predicting localization.

Now, Young, also member of the Whitehead Institute for Biological Research; Young lab postdoc Henry Kilgore; Regina Barzilay, the School of Engineering Distinguished Professor for AI and Health at MIT’s Computer Science and Artificial Intelligence Laboratory (CSAIL); and colleagues have built such a model, which they call ProtGPS. In a paper published on Feb. 6 in the journal Science, with first authors Kilgore and Barzilay lab graduate students Itamar Chinn, Peter Mikhael, and Ilan Mitnikov, the cross-disciplinary team debuts their model. The researchers show that ProtGPS can predict to which of 12 known types of compartments a protein will localize, as well as whether a disease-associated mutation will change that localization. Additionally, the research team developed a generative algorithm that can design novel proteins to localize to specific compartments.

“My hope is that this is a first step towards a powerful platform that enables people studying proteins to do their research,” Young says, “and that it helps us understand how humans develop into the complex organisms that they are, how mutations disrupt those natural processes, and how to generate therapeutic hypotheses and design drugs to treat dysfunction in a cell.”

The researchers also validated many of the model’s predictions with experimental tests in cells.

“It really excited me to be able to go from computational design all the way to trying these things in the lab,” Barzilay says. “There are a lot of exciting papers in this area of AI, but 99.9 percent of those never get tested in real systems. Thanks to our collaboration with the Young lab, we were able to test, and really learn how well our algorithm is doing.”

Developing the model

The researchers trained and tested ProtGPS on two batches of proteins with known localizations. They found that it could correctly predict where proteins end up with high accuracy. The researchers also tested how well ProtGPS could predict changes in protein localization based on disease-associated mutations within a protein. Many mutations — changes to the sequence for a gene and its corresponding protein — have been found to contribute to or cause disease based on association studies, but the ways in which the mutations lead to disease symptoms remain unknown.

Figuring out the mechanism for how a mutation contributes to disease is important because then researchers can develop therapies to fix that mechanism, preventing or treating the disease. Young and colleagues suspected that many disease-associated mutations might contribute to disease by changing protein localization. For example, a mutation could make a protein unable to join a compartment containing essential partners.

They tested this hypothesis by feeding ProtGOS more than 200,000 proteins with disease-associated mutations, and then asking it to both predict where those mutated proteins would localize and measure how much its prediction changed for a given protein from the normal to the mutated version. A large shift in the prediction indicates a likely change in localization.

The researchers found many cases in which a disease-associated mutation appeared to change a protein’s localization. They tested 20 examples in cells, using fluorescence to compare where in the cell a normal protein and the mutated version of it ended up. The experiments confirmed ProtGPS’s predictions. Altogether, the findings support the researchers’ suspicion that mis-localization may be an underappreciated mechanism of disease, and demonstrate the value of ProtGPS as a tool for understanding disease and identifying new therapeutic avenues.

“The cell is such a complicated system, with so many components and complex networks of interactions,” Mitnikov says. “It’s super interesting to think that with this approach, we can perturb the system, see the outcome of that, and so drive discovery of mechanisms in the cell, or even develop therapeutics based on that.”

The researchers hope that others begin using ProtGPS in the same way that they use predictive structural models like AlphaFold, advancing various projects on protein function, dysfunction, and disease.

Moving beyond prediction to novel generation

The researchers were excited about the possible uses of their prediction model, but they also wanted their model to go beyond predicting localizations of existing proteins, and allow them to design completely new proteins. The goal was for the model to make up entirely new amino acid sequences that, when formed in a cell, would localize to a desired location. Generating a novel protein that can actually accomplish a function — in this case, the function of localizing to a specific cellular compartment — is incredibly difficult. In order to improve their model’s chances of success, the researchers constrained their algorithm to only design proteins like those found in nature. This is an approach commonly used in drug design, for logical reasons; nature has had billions of years to figure out which protein sequences work well and which do not.

Because of the collaboration with the Young lab, the machine learning team was able to test whether their protein generator worked. The model had good results. In one round, it generated 10 proteins intended to localize to the nucleolus. When the researchers tested these proteins in the cell, they found that four of them strongly localized to the nucleolus, and others may have had slight biases toward that location as well.

“The collaboration between our labs has been so generative for all of us,” Mikhael says. “We’ve learned how to speak each other’s languages, in our case learned a lot about how cells work, and by having the chance to experimentally test our model, we’ve been able to figure out what we need to do to actually make the model work, and then make it work better.”

Being able to generate functional proteins in this way could improve researchers’ ability to develop therapies. For example, if a drug must interact with a target that localizes within a certain compartment, then researchers could use this model to design a drug to also localize there. This should make the drug more effective and decrease side effects, since the drug will spend more time engaging with its target and less time interacting with other molecules, causing off-target effects.

The machine learning team members are enthused about the prospect of using what they have learned from this collaboration to design novel proteins with other functions beyond localization, which would expand the possibilities for therapeutic design and other applications.

“A lot of papers show they can design a protein that can be expressed in a cell, but not that the protein has a particular function,” Chinn says. “We actually had functional protein design, and a relatively huge success rate compared to other generative models. That’s really exciting to us, and something we would like to build on.”

All of the researchers involved see ProtGPS as an exciting beginning. They anticipate that their tool will be used to learn more about the roles of localization in protein function and mis-localization in disease. In addition, they are interested in expanding the model’s localization predictions to include more types of compartments, testing more therapeutic hypotheses, and designing increasingly functional proteins for therapies or other applications.

“Now that we know that this protein code for localization exists, and that machine learning models can make sense of that code and even create functional proteins using its logic, that opens up the door for so many potential studies and applications,” Kilgore says.

Cellular traffic congestion in chronic diseases suggests new therapeutic targets

Many chronic diseases have a common denominator that could be driving their dysfunction: reduced protein mobility, which in turn reduces protein function. A new paper from the Young Lab describes this pervasive mobility defect.

Greta Friar | Whitehead Institute
November 26, 2024

Chronic diseases like type 2 diabetes and inflammatory disorders have a huge impact on humanity. They are a leading cause of disease burden and deaths around the globe, are physically and economically taxing, and the number of people with such diseases is growing.

Treating chronic disease has proven difficult because there is not one simple cause, like a single gene mutation, that a treatment could target. At least, that’s how it has appeared to scientists. However, research from Whitehead Institute Member Richard Young and colleagues, published in the journal Cell on November 27, reveals that many chronic diseases have a common denominator that could be driving their dysfunction: reduced protein mobility. What this means is that around half of all proteins active in cells slow their movement when cells are in a chronic disease state, reducing the proteins’ functions. The researchers’ findings suggest that protein mobility may be a linchpin for decreased cellular function in chronic disease, making it a promising therapeutic target.

In this paper, Young and colleagues in his lab, including postdoc Alessandra Dall’Agnese, graduate students Shannon Moreno and Ming Zheng, and research scientist Tong Ihn Lee, describe their discovery of this common mobility defect, which they call proteolethargy; explain what causes the defect and how it leads to dysfunction in cells; and propose a new therapeutic hypothesis for treating chronic diseases.

“I’m excited about what this work could mean for patients,” says Dall’Agnese. “My hope is that this will lead to a new class of drugs that restore protein mobility, which could help people with many different diseases that all have this mechanism as a common denominator.”

“This work was a collaborative, interdisciplinary effort that brought together biologists, physicists, chemists, computer scientists and physician-scientists,” Lee says. “Combining that expertise is a strength of the Young lab. Studying the problem from different viewpoints really helped us think about how this mechanism might work and how it could change our understanding of the pathology of chronic disease.”

Commuter delays cause work stoppages in the cell

How do proteins moving more slowly through a cell lead to widespread and significant cellular dysfunction? Dall’Agnese explains that every cell is like a tiny city, with proteins as the workers who keep everything running. Proteins have to commute in dense traffic in the cell, traveling from where they are created to where they work. The faster their commute, the more work they get done. Now, imagine a city that starts experiencing traffic jams along all the roads. Stores don’t open on time, groceries are stuck in transit, meetings are postponed. Essentially all operations in the city are slowed.

The slow down of operations in cells experiencing reduced protein mobility follows a similar progression. Normally, most proteins zip around the cell bumping into other molecules until they locate the molecule they work with or act on. The slower a protein moves, the fewer other molecules it will reach, and so the less likely it will be able to do its job. Young and colleagues found that such protein slow-downs lead to measurable reductions in the functional output of the proteins. When many proteins fail to get their jobs done in time, cells begin to experience a variety of problems—as they are known to do in chronic diseases.

Discovering the protein mobility problem

Young and colleagues first suspected that cells affected in chronic disease might have a protein mobility problem after observing changes in the behavior of the insulin receptor, a signaling protein that reacts to the presence of insulin and causes cells to take in sugar from blood. In people with diabetes, cells become less responsive to insulin — a state called insulin resistance — causing too much sugar to remain in the blood. In research published on insulin receptors in Nature Communications in 2022, Young and colleagues reported that insulin receptor mobility might be relevant to diabetes.

Knowing that many cellular functions are altered in diabetes, the researchers considered the possibility that altered protein mobility might somehow affect many proteins in cells. To test this hypothesis, they studied proteins involved in a broad range of cellular functions, including MED1, a protein involved in gene expression; HP1α, a protein involved in gene silencing; FIB1, a protein involved in production of ribosomes; and SRSF2, a protein involved in splicing of messenger RNA. They used single-molecule tracking and other methods to measure how each of those proteins moves in healthy cells and in cells in disease states. All but one of the proteins showed reduced mobility (about 20-35%) in the disease cells.

“I’m excited that we were able to transfer physics-based insight and methodology, which are commonly used to understand the single-molecule processes like gene transcription in normal cells, to a disease context and show that they can be used to uncover unexpected mechanisms of disease,” Zheng says. “This work shows how the random walk of proteins in cells is linked to disease pathology.”

Moreno concurs: “In school, we’re taught to consider changes in protein structure or DNA sequences when looking for causes of disease, but we’ve demonstrated that those are not the only contributing factors. If you only consider a static picture of a protein or a cell, you miss out on discovering these changes that only appear when molecules are in motion.”

 Can’t commute across the cell, I’m all tied up right now

Next, the researchers needed to determine what was causing the proteins to slow down. They suspected that the defect had to do with an increase in cells of the level of reactive oxygen species (ROS), molecules that are highly prone to interfering with other molecules and their chemical reactions. Many types of chronic-disease-associated triggers, such as higher sugar or fat levels, certain toxins, and inflammatory signals, lead to an increase in ROS, also known as an increase in oxidative stress. The researchers measured the mobility of the proteins again, in cells that had high levels of ROS and were not otherwise in a disease state, and saw comparable mobility defects, suggesting that oxidative stress was to blame for the protein mobility defect.

The final part of the puzzle was why some, but not all, proteins slow down in the presence of ROS. SRSF2 was the only one of the proteins that was unaffected in the experiments, and it had one clear difference from the others: its surface did not contain any cysteines, an amino acid building block of many proteins. Cysteines are especially susceptible to interference from ROS because it will cause them to bond to other cysteines. When this bonding occurs between two protein molecules, it slows them down because the two proteins cannot move through the cell as quickly as either protein alone.

About half of the proteins in our cells contain surface cysteines, so this single protein mobility defect can impact many different cellular pathways. This makes sense when one considers the diversity of dysfunctions that appear in cells of people with chronic diseases: dysfunctions in cell signaling, metabolic processes, gene expression and gene silencing, and more. All of these processes rely on the efficient functioning of proteins—including the diverse proteins studied by the researchers. Young and colleagues performed several experiments to confirm that decreased protein mobility does in fact decrease a protein’s function. For example, they found that when an insulin receptor experiences decreased mobility, it acts less efficiently on IRS1, a molecule to which it usually adds a phosphate group.

From understanding a mechanism to treating a disease

Discovering that decreased protein mobility in the presence of oxidative stress could be driving many of the symptoms of chronic disease provides opportunities to develop therapies to rescue protein mobility. In the course of their experiments, the researchers treated cells with an antioxidant drug—something that reduces ROS—called N-acetyl cysteine and saw that this partially restored protein mobility.

The researchers are pursuing a variety of follow ups to this work, including the search for drugs that safely and efficiently reduce ROS and restore protein mobility. They developed an assay that can be used to screen drugs to see if they restore protein mobility by comparing each drug’s effect on a simple biomarker with surface cysteines to one without. They are also looking into other diseases that may involve protein mobility, and are exploring the role of reduced protein mobility in aging.

“The complex biology of chronic diseases has made it challenging to come up with effective therapeutic hypotheses,” says Young, who is also a professor of biology at the Massachusetts Institute of Technology. “The discovery that diverse disease-associated stimuli all induce a common feature, proteolethargy, and that this feature could contribute to much of the dysregulation that we see in chronic disease, is something that I hope will be a real game changer for developing drugs that work across the spectrum of chronic diseases.”

A blueprint for better cancer immunotherapies

By examining antigen architectures, MIT researchers built a therapeutic cancer vaccine that may improve tumor response to immune checkpoint blockade treatments.

Bendta Schroeder | Koch Institute
November 25, 2024

Immune checkpoint blockade (ICB) therapies can be very effective against some cancers by helping the immune system recognize cancer cells that are masquerading as healthy cells.

T cells are built to recognize specific pathogens or cancer cells, which they identify from the short fragments of proteins presented on their surface. These fragments are often referred to as antigens. Healthy cells will will not have the same short fragments or antigens on their surface, and thus will be spared from attack.

Even with cancer-associated antigens studding their surfaces, tumor cells can still escape attack by presenting a checkpoint protein, which is built to turn off the T cell. Immune checkpoint blockade therapies bind to these “off-switch” proteins and allow the T cell to attack.

Researchers have established that how cancer-associated antigens are distributed throughout a tumor determines how it will respond to checkpoint therapies. Tumors with the same antigen signal across most of its cells respond well, but heterogeneous tumors with subpopulations of cells that each have different antigens, do not. The overwhelming majority of tumors fall into the latter category and are characterized by heterogenous antigen expression. Because the mechanisms behind antigen distribution and tumor response are poorly understood, efforts to improve ICB therapy response in heterogenous tumors have been hindered.

In a new study, MIT researchers analyzed antigen expression patterns and associated T cell responses to better understand why patients with heterogenous tumors respond poorly to ICB therapies. In addition to identifying specific antigen architectures that determine how immune systems respond to tumors, the team developed an RNA-based vaccine that, when combined with ICB therapies, was effective at controlling tumors in mouse models of lung cancer.

Stefani Spranger, associate professor of biology and member of MIT’s Koch Institute for Integrative Cancer Research, is the senior author of the study, appearing recently in the Journal for Immunotherapy of Cancer. Other contributors include Koch Institute colleague Forest White, the Ned C. (1949) and Janet Bemis Rice Professor and professor of biological engineering at MIT, and Darrell Irvine, professor of immunology and microbiology at Scripps Research Institute and a former member of the Koch Institute.

While RNA vaccines are being evaluated in clinical trials, current practice of antigen selection is based on the predicted stability of antigens on the surface of tumor cells.

“It’s not so black-and-white,” says Spranger. “Even antigens that don’t make the numerical cut-off could be really valuable targets. Instead of just focusing on the numbers, we need to look inside the complex interplays between antigen hierarchies to uncover new and important therapeutic strategies.”

Spranger and her team created mouse models of lung cancer with a number of different and well-defined expression patterns of cancer-associated antigens in order to analyze how each antigen impacts T cell response. They created both “clonal” tumors, with the same antigen expression pattern across cells, and “subclonal” tumors that represent a heterogenous mix of tumor cell subpopulations expressing different antigens. In each type of tumor, they tested different combinations of antigens with strong or weak binding affinity to MHC.

The researchers found that the keys to immune response were how widespread an antigen is expressed across a tumor, what other antigens are expressed at the same time, and the relative binding strength and other characteristics of antigens expressed by multiple cell populations in the tumor

As expected, mouse models with clonal tumors were able to mount an immune response sufficient to control tumor growth when treated with ICB therapy, no matter which combinations of weak or strong antigens were present. However, the team discovered that the relative strength of antigens present resulted in dynamics of competition and synergy between T cell populations, mediated by immune recognition specialists called cross-presenting dendritic cells in tumor-draining lymph nodes. In pairings of two weak or two strong antigens, one resulting T cell population would be reduced through competition. In pairings of weak and strong antigens, overall T cell response was enhanced.

In subclonal tumors, with different cell populations emitting different antigen signals, competition rather than synergy was the rule, regardless of antigen combination. Tumors with a subclonal cell population expressing a strong antigen would be well-controlled under ICB treatment at first, but eventually parts of the tumor lacking the strong antigen began to grow and developed the ability evade immune attack and resist ICB therapy.

Incorporating these insights, the researchers then designed an RNA-based vaccine to be delivered in combination with ICB treatment with the goal of strengthening immune responses suppressed by antigen-driven dynamics. Strikingly, they found that no matter the binding affinity or other characteristics of the antigen targeted, the vaccine-ICB therapy combination was able to control tumors in mouse models. The widespread availability of an antigen across tumor cells determined the vaccine’s success, even if that antigen was associated with weak immune response.

Analysis of clinical data across tumor types showed that the vaccine-ICB therapy combination may be an effective strategy for treating patients with tumors with high heterogeneity. Patterns of antigen architectures in patient tumors correlated with T cell synergy or competition in mice models and determined responsiveness to ICB in cancer patients. In future work with the Irvine laboratory at the Scripps Research Institute, the Spranger laboratory will further optimize the vaccine with the aim of testing the therapy strategy in the clinic.