For years, one of the most powerful weapons against certain blood cancers, called CAR-T cell therapy, has required an elaborate process: Doctors extract a patient's immune cells, ship them to a specialized facility where they're genetically reprogrammed to fight cancer, then ship them back for infusion back into the patient's bloodstream. This has revolutionized cancer treatment, but it takes weeks and can cost hundreds of thousands of dollars, placing it out of reach for many of the patients who need it most.
Now, scientists at UC San Francisco have developed a method to precisely reprogram these cancer-fighting cells directly inside the body, potentially eliminating the manufacturing process, cost, and waiting time that has kept this life-saving therapy out of reach for many patients around the world.
It is the first time that scientists have integrated a large sequence of DNA at a specific site in human T cells that were never removed from the body. Crucially, this targeted approach outperformed the standard method of randomly integrating DNA using viruses, a breakthrough that goes beyond CAR T to advance the fields of cell and gene therapy.
In experiments using mice with humanized immune systems, described March 18 in Nature , the researchers used the method to successfully treat aggressive leukemia, multiple myeloma, and even a solid tumor.
"I think this is just the beginning of a big wave of new therapies that will be truly transformational and save a lot of lives," said Justin Eyquem , PhD, an associate professor of medicine at UCSF and the senior author of the new paper. "I'm incredibly excited to be part of it."
Molecular scissors alter gene
CAR-T cell therapy works by giving T cells — the immune system's disease fighters — new genetic instructions to recognize and destroy cancer cells. These instructions come in the form of chimeric antigen receptors (CARs), molecules that protrude from the surface of T cells like antennae. When a CAR binds to specific proteins on a cancer cell's surface, it triggers the T cell to attack and kill the tumor cell. Seven CAR-T cell therapies are currently approved for use in blood cancers by the U.S. Food and Drug Administration.
But accessing these therapies, which cost $400,000 to $500,000, has proven difficult for many patients. The manufacturing process requires specialized facilities and takes weeks, during which some patients' cancers progress. And before receiving the engineered cells, patients must undergo intensive chemotherapy to clear space in their bone marrow for the new T cells — a punishing process that some patients, especially older or frailer ones, cannot tolerate.
"It's become a global access issue; many patients who would benefit from CAR-T cells either can't afford them or can't get them fast enough," Eyquem said. "There has been a big push in the field to try to move to directly producing these cells in the body."
Re-engineering immune cells in the body, called in vivo manufacturing, could also eliminate the need for preparatory chemotherapy.
To achieve this, Eyquem and his collaborators, including scientists at the Gladstone Institutes, Duke University, and Innovative Genomics Institute, designed a dual-particle system to carry CRISPR-Cas9 gene-editing machinery — the molecular scissors required to alter genes — directly to T cells circulating in the body. One particle was coated with antibodies against CD3, a protein found exclusively on T cell surfaces, ensuring the editing tools reach only their intended targets.
The second particle carried new DNA encoding the cancer-fighting CAR along with instructions to insert it at a specific location in the T cell genome, a site containing a molecular "on switch" only activated in T cells. Only when the gene lands in this exact spot does it coax the immune cells to make the new CARs. The particles were also engineered to evade immediate destruction by the immune system.
"When you manufacture these cells outside the body, you can do a lot of quality control to make sure you only end up with re-engineered T cells," said Eyquem. "Inside the body, we can't do that post-manufacturing quality control, so we really needed to optimize the approach upfront to avoid altering any other cells."
Clear detectable cancer in two weeks
The researchers, led by co-first authors William Nyberg, PhD, and Pierre-Louis Bernard, PhD, both UCSF postdoctoral fellows, tested their approach in mice engrafted with aggressive leukemia. A single injection of the dual-particle system cleared all detectable cancer in nearly all the mice within two weeks. The engineered CAR-T cells made up as much as 40% of immune cells in some organs and successfully eliminated cancer from both the bone marrow and spleen.
The approach also worked against multiple myeloma and, strikingly, against a solid sarcoma tumor. Solid tumors have historically resisted CAR-T therapy, making this result particularly significant.
The T cells engineered inside the body also unexpectedly appeared to outperform those manufactured in the lab.
"What was especially remarkable was that the cells we're generating in vivo actually look better than what we make in the lab," Eyquem said. "We think that when cells are taken out of the body and grown in the lab, they lose some of their 'stemness' and proliferative capacity and that doesn't happen here."
The technology still must be scaled up for use in humans, and clinical trials will be needed to assess safety and efficacy. Eyquem and his collaborators have founded a company called Azalea Therapeutics to advance the dual-particle platform described in this research through clinical development.
"If we can translate this to humans, we could dramatically reduce costs, eliminate waiting times, and potentially allow community hospitals — not just major cancer centers — to offer these life-saving therapies," he said. "That would truly democratize access to CAR-T cell therapy."
