CRISPR instead of viral vectors
T cells for personalized cancer therapy can be edited by CRISPR-Cas
Elena Kleshchenko, PCR.news
The private biopharmaceutical company PACT Pharma has developed a new personalized T-cell therapy, and T-cells with receptors specific to tumor neoantigens are created using CRISPR, not viral vectors. (It is quite difficult to make vectors individually for each patient.) The results of the first clinical trial are published in the journal Nature (Foy et al., Non-viral precision T cell receptor replacement for personalized cell therapy).
T cells can inhibit the development of cancer by recognizing altered proteins on the surface of tumor cells. Unfortunately, there are not enough such cells in the body. Isolation of cancer-detecting T-cell receptors (TCR) and the creation of therapeutic T—cells with such receptors is a promising new method of therapy.
In phase 1 of the clinical trial, 16 patients with metastatic solid tumors resistant to standard treatments (mainly colorectal cancer, as well as breast cancer and lung cancer) received genetically engineered T cells that express receptors targeting their individual cancer antigens.
Blood was taken from patients, T-cells were isolated with the help of special reagents. More than 5,000 mutations targeting various subtypes of the immune system HLA (receptors presenting cellular antigens, including tumor antigens) have been identified in the genomes of these cells. Sequencing of TCR genes made it possible to identify 175 unique tumor-specific receptors. Personalized libraries containing hundreds of tumor antigens were obtained by comparing DNA and RNA samples of peripheral blood and a patient's tumor biopsy.
To create a personalized therapy, one or more receptors were selected for each patient. T-cells were edited using CRISPR-Cas9 — in one step, their own TCR (TCRa and TCRß) and nokin were knocked out into the TRAC locus of the genes of two tumor-specific TCR chains. Homologous recombination with a fragment from a plasmid was used for nokin. Then the T cells were grown in culture for 11 days to get them in large quantities, and injected into the patient.
Some patients underwent a biopsy after the infusion and found that the edited T-cells were mainly directed to the tumor. Treatment led to stabilization of the disease in 5 out of 16 patients, while in the remaining 11 the disease progressed. Only two patients had temporary adverse reactions associated with T-cell therapy (one had fever and chills, the other had confusion), and all patients had expected side effects associated with concomitant chemotherapy.
"This study is a significant step forward in the development of personalized cancer treatment using isolated immune receptors that can specifically recognize unique mutations in a patient's own cancer," said Anthony Ribas of the University of California, Los Angeles, one of the lead authors of the paper. James Heath, president of the Institute of Systems Biology in Seattle, and Nobel laureate David Baltimore, Professor Emeritus at Caltech and a member of the UCLA Comprehensive Johnson Cancer Center, also participated in the development of the technology. Nine academic centers participated in the study.
The authors emphasize that using CRISPR instead of vectors makes T-cell therapy more accessible. "We are working to make personalized TCR T-cell therapy a reality for all cancer patients," said Stephanie Mandl, director of science at PACT Pharma. — A key element for achieving this goal of wide application is our own catalog of 64 human leukocyte antigens (HLA), which covers patients of all nationalities. This is a key difference from most modern T-cell therapies, which are limited to only one HLA molecule, common in people of European descent."
Among the limitations of the proposed approach, the authors of the article mention the long time required to characterize potential antigens, isolate, clone and test T-cell receptors, as well as the different affinity of patient-specific T-cell receptors to tumor antigens. They noted that they see ways to increase the effectiveness of therapy.
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