Closest look yet at killer T-cell activity could yield new approach to tackling antibiotic resistance An in-depth look at the work of T-cells, the body's bacteria killers, could provide a roadmap to effective drug treatments.

In a study that could provide a roadmap for combatting the rising threat of drug-resistant pathogens, researchers have discovered the specific mechanism the body’s T-Cells use to kill bacteria.

University of Michigan researchers, in collaboration with colleagues at Harvard University, have discovered a key difference between the way immune cells attack bacteria and the way antibiotics do. Where drugs typically attack a single process within bacteria, T-Cells attack a host of processes at the same time.

On Thursday, the journal Cell published findings from a team headed by U-M’s Sriram Chandrasekaran and Harvard’s Judy Lieberman. It’s a study with potential implications for drug-resistant pathogens—a problem projected to kill as many as 10 million people annually across the globe by the year 2050.

“We have a huge crisis of antibiotic resistance right now in that most drugs that treat diseases like tuberculosis or listeria, or pathogens like E.coli, are not effective,” said Chandrasekaran, an assistant professor of biomedical engineering. “So there is a huge need for figuring out how the immune system does its work. We hope to design a drug that goes after bacteria in a similar way.”

We’ve reached a point where we take what antibiotics can do for granted, and we can’t do that anymore.Sriram Chandrasekaran

Killer T-Cells, formally known as cytotoxic lymphocytes, attack infected cells by producing the enzyme granzyme B. How this enzyme triggers death in bacteria has not been well understood, Chandrasekaran said.

Proteomics – a technique that measures protein levels in a cell—and computer modeling, allowed researchers to see granzyme B’s multi-pronged attack targeting multiple processes.

Chandrasekaran and his team monitored how T-Cells deal with three different threats: E. coli, listeria and tuberculosis.

“When exposed to granzyme B, the bacteria were unable to develop resistance to the multi-pronged attack, even after exposure over multiple generations,” Chandrasekaran said. “This enzyme breaks down multiple proteins that are essential for the bacteria to survive.

“It’s essentially killing several birds with one stone.”

The possible applications of the new findings on T-Cells run the gamut from the creation of new medications to the re-purposing of previously-approved drugs in combination to fight infections by mimicking granzyme B.

Chandrasekaran’s team is now looking at how bacteria hide to avoid T-Cell attacks.

And the need for a new approach in some form is dire. World Health Organization officials describe antibiotic resistance as “one of the biggest threats to global health, food security and development today.”

Sriram Chandrasekaran, Assistant Professor of Biomedical Engineering, shows a computer model of a pathway for a potential disease or infection. Photo: Joseph Xu

Each year, an estimated 700,000 deaths are linked to antibiotic-resistant bacteria, according to the World Health Organization. Projections show that number skyrocketing to 10 million by 2050.

England’s top health official, Sally Davies, recently said the lost effectiveness of antibiotics would mean “the end of modern medicine.”

“We really are facing—if we don’t take action now—a dreadful post-antibiotic apocalypse,” she was quoted saying earlier this month. “I don’t want to say to my children that I didn’t do my best to protect them and their children.”

Of particular concern is the fact that there are few new antibiotics in the pipeline. The heyday of new antibiotics occurred the 1940s through the 1960s, with releases eventually grinding almost to a halt by the end of the twentieth century.

“We’ve reached a point where we take what antibiotics can do for granted, and we can’t do that anymore,” Chandrasekaran said. “We’re taking inspiration from the human immune system, which has been fighting infections for thousands of years.”

The paper is titled, “Granzyme B disrupts central metabolism and protein synthesis in bacteria to promote an immune cell death program.” The research is funded by the National Institutes of Health, Harvard University and the University of Michigan.

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Bionic heart tissue: U-Michigan part of $20M center Scar tissue left over from heart attacks creates dead zones that don’t beat. Bioengineered patches could fix that.

The University of Michigan is partnering on an ambitious $20 million project to grow new heart tissue for cardiac patients. The new research center has been awarded to Boston University (BU), with strong partnership from U-M and Florida International University (FIU).

“A heart attack creates scar tissue, and the heart never returns to full function. But for every person, we could create a living patch that a surgeon could stitch in,” said Stephen Forrest, who leads the nanotechnology aspect of the project and is U-M’s Peter A. Franken Distinguished University Professor of Engineering. “It’s very audacious.”

The project is a National Science Foundation Engineering Research Center. These 5-year grants are typically renewed for another 5 years, so the researchers are looking at a 10-year timeline to go from the current state of tissue engineering to working, implantable heart tissue.

A heart attack creates scar tissue, but we could create a living patch that a surgeon could stitch in.Steve Forrest

“Heart disease is one of the biggest problems we face,” said David Bishop, director of the new center and a BU professor of electrical and computer engineering and physics. “This grant gives us the opportunity to define a societal problem, and then create the industry to solve it.”

The living patches the researchers are developing would consist of heart muscle cells, blood vessels to carry nutrients in and waste out, and optical circuitry to make the heart muscle cells beat in synchrony. Already, researchers in the lab have been developing ways to structure cells in scaffolds that mimic particular organs and grow blood vessels into artificial tissues. But typically, working implants have been static, biodegradable materials such as artificial windpipes that the body gradually replaces with tissue. Working tissue, like heart muscle, would need to be responsive as soon as it was implanted.

Engineering Research Center grants are extremely competitive, with only four of more than 200 applicants receiving an award in 2017. These centers are designed to work directly with industry to translate breakthroughs along the way out of the lab and into healthcare. Just producing a more true-to-life “heart on a chip” could aid the pharmaceutical industry in developing better treatments for problems such as arrhythmia.

Ramcharan and her colleagues in Lahann’s lab will help design and produce a polymer-protein construct that mimics the 3D matrix connecting the cells in human heart muscle. Heart muscle cells moving into this environment will then be able to link up into a single tissue. Photo: Joseph Xu, Michigan Engineering Communications & Marketing.

In order to produce the heart tissue, the team intends to start with an artificial scaffold that mimics the 3D structure of heart tissue. Joerg Lahann, a U-M professor of chemical engineering, will work with the team building the flexible polymer scaffold, as well as on the attachment and monitoring of cells within that framework.

“Michigan is pleased to lend expertise to the development of implantable heart tissue, which could improve and extend so many lives,” said Alec D. Gallimore, the Robert J. Vlasic Dean of Engineering. “Our faculty members are leaders in nanotechnology and in developing materials that support and interact with living cells and tissues, two areas that are critical to the project’s success.”

The 3D scaffold will initially be peppered with nanometer-sized gold patches that act as attachment points for protein fragments, called peptides, which will then serve as anchors for the cells. They will be printed onto the gold patches using a technique developed by Forrest and Max Shtein, a U-M associate professor of materials science and engineering. This method, called organic vapor jet printing, was initially invented for mass-producing electronic devices.

“The adaptation of this technology to biological systems represents a radically new step,” said Forrest. U-M will receive $2.8 million for these contributions.

Christopher Chen, the center’s director of cellular engineering and a BU professor of biomedical engineering, will lead the effort to grow heart muscle cells on the scaffold and infuse the tissue with blood vessels. Meanwhile, Alice White, director of nanomechanics and chair of the BU mechanical engineering department will work closely with Arvind Agarwal, an FIU professor of mechanical and materials engineering, to produce an artificial nervous system that uses light to synchronize the heartbeat in the tissue.

Stacy Ramcharan, a doctoral student in chemical engineering, uses a computerized system to layer polymer fibers, forming a scaffold for growing cells into artificial tissues. Photo: Joseph Xu, Michigan Engineering Communications & Marketing.

“It’s humbling to have the opportunity to work on something that could really be a game changer,” says Bishop. “If we succeed, we’ll save a lot of lives and add meaningful years for many people.”

In addition to the technical thrusts led by Forrest, Chen and White, Thomas Bifano, a professor of mechanical engineering and director of BU’s Photonics Center, will direct imaging.

Along with the core partners, Harvard Medical School, Columbia University, the Wyss Institute at Harvard, Argonne National Laboratory, the École Polytechnique Fédérale de Lausanne in Switzerland, and the Centro Atómico in Argentina will offer expertise in bioengineering, nanotechnology, and other areas.

Forrest is also the Paul G. Goebel Professor of Engineering, and a professor of electrical engineering and computer science, material science and engineering, and physics. Lahann is also a professor of material science and engineering, biomedical engineering, and macromolecular science and engineering. Shtein is also an associate professor of chemical engineering, macromolecular science and engineering, and art and design. Gallimore is also the Richard F. and Eleanor A. Towner Professor, an Arthur F. Thurnau Professor, and a professor of aerospace engineering.

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Reading cancer’s chemical clues A nanoparticle-assisted optical imaging technique could one day read the chemical makeup of a tumor.

 

A tumor’s chemical makeup holds valuable clues about how to fight it. But today, it’s difficult or impossible to examine the chemistry inside a tumor. A nanoparticle-assisted optical imaging technique could one day enable doctors to read those clues in real time, providing a non-invasive precision medicine approach that could match treatment to individual tumors.

“Tumors vary widely from one patient to the next, so the more we know about them, the more effective our treatments become. This is especially important with chemotherapy because of its high cost and severe side effects,” said Xueding Wang, a University of Michigan professor of biomedical engineering who helped develop the technique. “This could form the basis of precision medicine treatments that offer better outcomes, fewer side effects and lower costs.”

Most of us are working in the dark with regard to tumor imaging. -Xueding Wang

Doctors already know, for example, that some treatments don’t work on acidic tumors while others are ineffective against tumors that have low oxygen levels. If they know the chemical makeup of a given tumor, they can start the right treatment immediately, then keep close tabs on its effectiveness over time.

In a recent paper, U-M researchers successfully used the process to get a three-dimensional view of the pH level inside tumors in mice, and they believe that they will also be able to use it to read a variety of other important chemical markers inside cancers. The new technology is detailed in a paper published September 7 in Nature Communication.

Images of a mouse tumor obtained with the new technique. Row A shows the presence of the nanoparticle itself, in blue. Row B shows the pH of the tumor. Image C shows oxygen saturation and image D shows hemoglobin concentration. Photo courtesy of Janggun Jo, Michigan Engineering

“Most of us are working in the dark with regard to tumor imaging. There are very few cases where we can study the chemistry of a tumor,” said Wang. ”We hope to change that with this technology, which offers a spatially detailed, real-time look at the chemistry inside a tumor, even when it’s deep inside the body.”

The technique uses a two-part system, starting with a purpose-built, injectable nanoparticle that’s absorbed only by cancerous cells. The particles were loaded with a marker dye that changes color in response to the tumor’s pH to measure acidity.

Wang and Raoul Kopelman, the Richard Smalley Distinguished University Professor of Chemistry, Physics and Applied Physics, made the nanoparticles small enough to fit through tiny cracks in the walls of cancer cells called fenestrations—imperfections that form because cancer cells grow so quickly. They then coated the particles with protein fragments, or peptides, that are attracted to cancerous cells.

“The peptides on the particle are like tugboats guiding an ocean liner,” Kopelman explained.

The particles were injected into mice, where they infiltrated the cancerous cells and the pH-sensitive dye did its work. Next, the team read the dye by flashing pulses of laser light into the tumor from outside the mouse’s body and recording the ultrasound signal that’s reflected back.

Chang Lee, Ph.D., examines the pH-sensitive dye used in the new cancer imaging technique. Photo: Akhil Kantipuly, Michigan Engineering

“Inside the body, the laser’s energy turns from light into heat, then from heat into sound, a bit like thunder,” said Wang. “We can use ultrasound to read that sound energy, then digitally convert it back to optical information. That provides a painless, non-invasive way for us to see the color change in the injected dye, even when it’s deep inside the body.”

The researchers caution that an approved treatment is several years off. But they note that the imaging technology is already under clinical trial, as are the individual components of the nanoparticle. In the meantime, they are working on similar approaches that could be used to measure markers other than pH, like potassium and oxygen levels. They envision a treatment that could measure several different aspects of a tumor’s chemistry using a single scan.

The paper is titled “In vivo quantitative imaging of tumor pH by nanosonophore assisted multi-spectral photoacoustic imaging.” The research was supported by funding from the National Institutes of Health through the National Cancer Institute (grant number R01CA186769).

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