More Effective Cancer Immunotherapy: Stanford’s New Method To Find Antigens That Trigger Specific Immune Cells

The surface of a cell can reveal its secrets. It is embellished with tens of thousands to hundreds of thousands of molecules that aid immune cells in differentiating between friends and enemies. Antigens are some of those obtruding substances that cause the immune system to assault. However, it can be challenging for researchers to locate those antigens in the molecular forest since they frequently differ between individuals.

A group of Stanford researchers has created a novel technique to predict which antigens will trigger a potent immune response more quickly and correctly. Their method might aid in the development of more potent cancer immunotherapies. The research, which was directed by Polly Fordyce, an Institute Scholar at Sarafan ChEM-H, will be published in the journal Nature Methods today, September 5, 2022.

Immune cells known as T cells prowl the body, squeezing past other cells as they go. They employ T cell receptors to read peptides, or brief proteins, that are molecularly encased in larger proteins known as major histocompatibility complexes (pMHCs), which protrude from cell surfaces. Healthy host cells display an assortment of pMHCs that do not stimulate an immune response. To find and eliminate cells with these foreign signatures, T cells must first recognize disease-indicating peptides. It has long been unclear how T cells sensitively distinguish these antigenic peptides from host peptides to prevent unintentionally harming host cells.

According to Fordyce, an assistant professor of bioengineering and genetics, "a T cell can detect a single antigenic peptide among a sea of 10,000 or 100,000 non-antigenic peptides being expressed on cell surfaces."

The T cell crawl holds the secret of selectivity. The connections between receptors and peptides are stressed as a result of T cells moving, and often, this additional stress is enough to break the binding. However, occasionally it has the opposite result. Professor of molecular and cellular physiology and structural biology Chris Garcia, who is also a co-author of the work, has demonstrated before that the most antigenic peptides are those whose connections with T cell receptors become stronger in response to sliding.

Fordyce compared it to a Chinese finger trap. The binding actually lasts longer when the receptor-antigen connection is somewhat pulled.

It is necessary to apply that sliding, or shearing, force between a peptide and a T cell while also monitoring T cell activity to determine the optimal antigen-receptor combinations. To obtain reproducible data for as many potential peptide/T cell receptor pairs as possible, this should ideally be repeated thousands of times. Existing techniques, however, take a lot of time and may only be able to measure one peptide with hundreds of T cells in a single day.

The study's first author, postdoctoral researcher Yinnian Feng, devised a method that enables the team to assess the interactions of 20 different peptides with thousands of T cells in less than five hours.

They created small, spherical beads from a polymer that swells when heated and affixed a few molecules of a particular peptide-studded pMHC to their surfaces to create a simpler system that mimics cells with dangling peptides. They placed a T cell on top of each bead and allowed enough time for the receptors to attach to the peptides before mildly heating the bead. The T cell is stretched to simulate the force it would encounter sliding over body cells as a result of the bead's growth, which widens the space between the tether points. The team then measured the T cells' level of activity after applying that force.

They may track many separate pMHCs by performing hundreds of individual tests concurrently using beads that are each labeled with a distinct color. After each run, they captured two sets of images that were tiled across each slide: one set that revealed the pMHC a specific bead was displaying, and another that revealed the level of activity of each T cell above that bead. They can determine which antigens elicited the highest T cell responses by comparing those photos.

The research team used 21 different peptides to demonstrate their platform and show that their findings supported the existence of known activating and non-activating peptides for one T cell receptor as well as the discovery of a previously unidentified antigen that elicited a potent T cell response. The T cell receptors that generate the highest affinity contacts with antigens in the lab are frequently also triggered by non-antigenic peptides in the body. This presents a difficulty for immunotherapy, which they have already started to address in collaboration with the Garcia lab. This negative effect is hazardous because it causes healthy cells to die.

The research team's technology allowed them to characterize T cell receptors that were specifically designed to recognize tumor antigens without off-target reactivity. They intend to create libraries of more than 1,000 peptides in upcoming studies to find new antigens.

The researchers are hopeful that this method, which is rapid and only needs a small number of cells, or an improved version of it, will one day be used to enhance customized immunotherapies.

In addition to identifying which antigens are capable of potently activating a patient's own T cells to more efficiently target cancer cells, this platform can help improve attempts to create T cells that selectively target cancer cells, according to Fordyce.

Fordyce is a researcher with the Chan Zuckerberg Biohub and a member of Stanford Bio-X, SPARK, and the Wu Tsai Neurosciences Institute. Garcia is a researcher for the Howard Hughes Medical Institute, a member of Stanford Bio-X, the Stanford Cancer Institute, and the Wu Tsai Neurosciences Institute.

The National Institutes of Health and a seed grant from Stanford Bio-X Interdisciplinary Initiatives both contributed to the funding of the project.

By STANFORD UNIVERSITY