Researchers at the USC Viterbi School of Engineering and Keck School of Medicine of USC have engineered a remarkable genetic tool that addresses one of the most persistent obstacles in cancer immunotherapy: the physical softness of tumours.

The work, detailed in the journal Nature Biomedical Engineering, reveals how the soft environment of a tumour enables cancer cells to become “stealthy” and resistant to treatment, providing a powerful new mechanism to tag those evasive cells for destruction.

The paper’s lead author, Jenny Yunjia Qu, a postdoctoral researcher in the lab of Peter Yingxiao Wang, the Dwight C. and Hildagarde E. Baum Department Chair in the Alfred Mann Department of Biomedical Engineering, collaborated with the USC Viterbi and Keck School of Medicine teams to develop a technique that successfully targets hard-to-reach “cancer stem-like cells.” These cell types possess stem cell-like abilities, including self-renewal, which make them key drivers of tumour growth, drug resistance, and cancer recurrence.

CAR T-cell therapy is a revolutionary cancer treatment in which T-cells—a type of white blood cell—are removed from a patient and given the unique chimaeric antigen receptor (CAR).

The CAR binds to cancer cell-associated antigens, directing T-cells to destroy the cancer cells.

CAR T therapy has revolutionised the treatment of blood cancers, but it struggles against solid tumours.

The Wang Lab team has demonstrated the critical role of the physical microenvironment of a tumour in the success of CAR T-cells, particularly when that microenvironment is soft.

“This is a relatively new finding in the field—that soft environments can cause cancer cells to be less killable by CAR-T cell therapy, and other approaches,” said Wang.

“Soft environment cancer cells typically have a greater tendency to become stem cell-like cells, which is also consistent with resistance to being killed by CAR-T therapy.”

Qu said that you could imagine the problem like trying to pierce a hole in something gelatinous, like Jello or tofu—the soft and springy nature of the material making it hard to penetrate.

“CAR T-cells kill tumours by grabbing individual cells, poking holes in their membrane and secreting cytotoxic molecules into the tumour cells. When the CAR T-cell is going in, they can really see, grab and kill the stiffer cells, but they neglect the softer, Jello-like ones,” Qu said.

“The cells are soft, therefore they cannot be grabbed easily, so the CAR T-cells have less efficacy killing them,” Wang said.

“Jenny essentially identified the mechanism in these soft, stem-like tumour cells, and utilised this understanding and molecular insight to rewire them so that they can produce more recognisable markers on their surface. This makes the cells more recognisable by the CAR T-cells, and therefore they can be attacked with a higher probability of being killed.”

To address this immune evasion, the research team first needed a way to identify and track these evasive soft cells.

They engineered a genetic tool called the Mechano-Recorder, which acts like a “black box” inside the cell.

The tool records the level of mechanical softness the cell experiences.

The researchers discovered that softer cancer cells displayed significantly higher intracellular calcium signalling.

The Mechano-Recorder converts this transient calcium signal into a stable, long-lasting fluorescent signal, effectively barcoding these cells with their mechanical experience.

“The recording part is the most exciting part,” said co-author Longwei Liu, assistant professor of ophthalmology and biomedical engineering at USC, “It’s just like you’re taking a picture of the cancer cells at a certain time points, and recording the signal for diagnostic purposes or for developing a therapeutic purpose.”

The team then successfully used this recorder to show that soft environments specifically promote these recorder-positive cells, which exhibit stem-cell-like features, including markers associated with metastasis and hypoxia.

The researchers then leveraged the Mechano-Recorder’s design for therapeutic action,  genetically reprogramming the system to convert the signature of soft, resistant cells into a targetable beacon for immune cell attack.

“What we are trying to do is firstly recognise the difference between the softer and the stiffer cells. And then we rewire the softer cells to express another synthetic antigen,” Qu said.

Qu and her colleagues replaced the fluorescent reporter in the recorder with CD19, a clinically validated antigen commonly used in CAR T therapy for blood cancers.

This process effectively broadcasts a ‘kill me’ signal to CAR T-Cells.

“When the softer cells express this signal, the CD19 CAR T-cells that we engineered will come in and only see those cells with the ‘kill me’ signal, and then they’ll attack,” Qu said.

“Finding the soft spot in solid tumours is a major new direction in making immune medicated therapy a reality,” said W. Martin Kast, a co-author and the Walter A. Richter Cancer Chair and professor of Immunology & Immune Therapeutics at the Keck School of Medicine.

Tested in breast cancer cell lines, patient-derived cells, and mouse models, the rewiring strategy successfully overcame the resistance of soft cancer stem-like cells.

The results showed that these reprogrammed tumours, though softer, exhibited greater T-cell infiltration and enhanced killing efficacy.

Wang added that this Mechano-Recorder and cell barcoding system had broad applications across cancers and many other diseases beyond breast cancer tumours.

“This same approach could be applicable easily or readily extended to other types of solid tumours—essentially all of them,” Wang said.

“In this particular work, we convert a soft-induced stimulus into a “kill me” signal. But we can do the same thing for many other cell signals as well. Any molecular signalling pathway we detect we can convert into a “kill me” signal, or a “help me” signal, depending on what we need, and depending on what kind of disease we are dealing with.”

The team has already successfully tested the approach in models of glioblastoma, pancreatic cancer, and prostate cancer.

Wang said that the team’s latest work establishes a foundational concept: understanding the molecular consequences of a cancer cell’s biophysical cues, like softness, allows researchers to “steal” the molecular information and then wire it to behave in a way that is more beneficial to treatment.

By transforming a tumour’s softness-driven stealth into an obvious target, the work paves an entirely new path for improving immunotherapy outcomes.

Source: USC Viterbi