Ultrasound Imaging at the Cellular Level

Since 1956 when ultrasound was first used for clinical purposes in Glasgow, Ireland by obstetrician Ian Donald and engineer Tom Brown, a significant amount of research has been done with using ultrasound for both imaging and therapy. Whenever we hear about ultrasound imaging, for most of us, it probably resonates with pregnancy and seeing images of babies developing inside the womb, or ultrasound being used to diagnose and isolate various diseases found in different areas of the body. Physical therapists have also used therapeutic ultrasound since the 1940s to treat a variety of conditions, from tissue relaxation, increasing local blood flow, reduce swelling, and promote bone fracture healing¹. However, ultrasound techniques primarily rely on using sound waves that bounce off the tissues to relay its anatomical shapes and density, thus limiting its ability to penetrate beyond tissue

to the cellular level. In many ways, modern medicine is still dependent upon further evolution of science and technology to bring forth revolutionary changes.

The brilliant minds at the California Institute of Technology (Caltech), like Professor Mikhail Shapiro and his team at the Shapiro lab, are diving more deeply to further the value of this discovery to find answers to help bring new therapies. They have discovered a way to use proteins from photosynthetic microbes found in nature to enhance ultrasound-imaging techniques that go beyond tissues to target the molecular and cellular levels. The foundation of their research is geared toward basic biology and cellular therapy with an overall goal to develop ways to image and control the function of cells inside the body.

During the early stages of research, they found the advantages of using cells as therapeutics, as opposed to molecules, are that cells are more sophisticated and can be programmed to respond and perform very specific tasks. They can also generate more cells and can self-destruct at the appropriate time. Nevertheless, one of the challenges of using cells is that upon injecting them into the targeted area of the body, it is difficult to verify if the cells have reached their destination and if delivery was successful. It is also difficult for cells to receive instructions from the outside world, for example from a physician, which is why a need to create a cellular “antenna” was required to allow for such imaging and communication.

“We’re able to engineer sophisticated cellular agents and administer them into the body, but once they are injected we lack the ability to see them or give them commands to perform functions for which they were originally designed,” adds Professor Shapiro. “Our goal at the Shapiro lab is to give cells that are engineered the ability to communicate with the outside world from inside the body.”

Protein Nanostructured Gas Vesicles

In 2014, Professor Shapiro discovered that he could get ultrasound signals from gas-filled structures found inside water-dwelling organisms, which use these proteins to float on top of the water and get ideal exposure to sunlight. During experimentation, he and his team discovered that these gas vesicles, or protein-shelled nanostructures, would be able to reflect sound waves during ultrasound imaging. This protein contains air on the interior that provides the buoyancy mechanism. Since it contains air, it has a different density from tissue (which is mainly water), allowing the ability to scatter sound waves to use with ultrasound.

Gas vesicles would enhance ultrasound methods to capture images of specific cell types. Professor Shapiro and his team work collaboratively on multiple projects with the goal to use this protein to image gene expression and locating cells within the body. The challenge with this approach is that the structure of gas vesicles are much more complicated than a simple fluorescent protein, which is coded by a single gene. Gas vesicles are encoded by a group of genes – at least eight of them to be precise, that have to work together to form this structure. “Transferring this more complex genetic machinery from one domain of life to another, and from one cell to another, presents a difficult challenge,” expressed Shapiro. Respectively, the Lego-like proteins would also allow interchangeable pieces of other proteins to attach to the surface of gas vesicles so that they can be modified to target specific properties, providing enhanced images of the molecules in various colors for greater definition.

Ultimately, gas vesicle proteins can be engineered to target tissues in the body that display specific cellular targets, for example, certain proteins that are overproduced in tumor cells. Gas vesicles can also be designed with longer and stronger nanostructures to prevent them from collapsing. These added functionalities along with a modular, Lego-like system give Professor Shapiro’s research some traction.

Temperature Gives Bacterial and Human Cells a Way to Communicate

Human cells are not the only cells that can be engineered for specific tasks. A new study conducted by Professor Shapiro’s lab involves using genetically altered bacterial cells to release medicine directly into a specific site of a patient who has a tumor or other disease. By using the heat generated from ultrasound, the medicine would be administered at precisely the right time. Controlling the temperature would grant these genetically modified bacterial and human cells the ability to listen and communicate when properly activated.

Many scientists and engineers have shown interest in experimenting with developing cells into therapy, like with immunotherapy. Immunotherapy uses engineered immune cells administered into the body to recognize tumor antigens and to wipe out cancer. Another example of therapy is using engineered probiotics, which are bacteria that can be swallowed and would produce a beneficial activity in the gastrointestinal tract. Once in the GI tract, they can either displace pathogens; prevent pathogens from taking hold on the GI tract, or release molecules, like anti-inflammatory compounds.

His research also showed that medicine administered into the patient would have the ability to break down and self-destruct when necessary, for example, if the patient had a high fever indicating that the bacterial therapy may not be working effectively and it would be in the patient’s best interest to terminate the therapy. Using the body’s natural temperature as a control mechanism to destroy the cells, it acts as a natural fail-safe kill switch to ensure that the therapy will terminate precisely when required.

The Future of Ultrasound: Noninvasive Therapy to Target Tumors and Cancer Cells

The future of ultrasound tools and imaging could potentially open a new doorway of possibilities for noninvasive procedures for patients with tumors and other diseases. The idea that cells can be engineered to target a specific area of a patient’s body using heat from ultrasound could pave the way to the future of noninvasive surgeries.

“I’m driven by the potential outcome; the ultimate applications and significance of being able to image and communicate with cells inside the body, but I’m equally driven by the challenge of finding and engineering unique proteins, like gas vesicles, that have unusual physical properties. As engineers, we want to understand proteins from both a biological and physical point of view, and engineer them accordingly,” Shapiro adds. His busy team at the Shapiro lab consists of students from many different scientific backgrounds from physics, biology, and chemistry. “I enjoy engaging in challenges that include multiple aspects of these various backgrounds to collectively produce promising results and solutions,” Shapiro says.

Professor Shapiro and his team at Caltech have every reason to be optimistic about their research and the impact it would have on the health outcomes for millions of people. Their exploration and commitment to creating a new field in biomolecular and cellular ultrasound, and its success, would also create a pathway to help inspire and encourage other engineers and scientists around the world to begin using this technology in various applications to benefit many lives.

Mikhail Shapiro, Ph.D.,

Assistant Professor of Chemical Engineering, HMRI Principal Investigator – Biochemistry and Molecular Biophysics, Chemical Engineering

Assistant Professor Mikhail Shapiro completed his B.S. degree at Brown University in 2004 and received his Ph.D. from Massachusetts Institute of Technology (MIT) in 2008. Since joining Caltech in 2013, he and his team have been developing technologies to study biological processes that occur deep inside

living organisms. This research involves molecular and cellular engineering, and uses various forms

of energy: magnetic, mechanical, thermal and chemical. They utilize multiple biophysical methods that include magnetic resonance, ultrasound, infrared and electro-physiology to enable the imaging

and control of biological function.

Professor Shapiro’s work has been featured in several publications highlighting his body of work and

the progress he has made linking magnetic resonance imaging (MRI) and ultrasound signals to gene expressions in cells, including tumor cells and commensal microbes, found in living tissue. This technique could eventually monitor cells and genes non-invasively, which could ultimately lead to non-surgical biopsies or interventions.

Go to www.caltech.edu/news to find out more.