"Enhanced" CAR-T lymphocytes
The combination of immunopreparation and CAR-T-lymphocytes saved the lives of mice with cancer
Oleg Lischuk, N+1
American researchers have managed to repeatedly increase the effectiveness of therapy of oncohematological diseases with lymphocytes with chimeric antigen receptors (CAR-T-lymphocytes) in preclinical trials. To do this, they used an experimental drug — a modified human cytokine that stimulates lymphocytic growth. In in vitro experiments and on animal models of mouse and human tumors, such therapy led to the persistent destruction of cancer cells, while CAR-T-lymphocytes themselves gave only a temporary effect. The report on the work was published in the journal Nature Communications (Kim et al., A long-acting interleukin-7, rhIL-7-hyFc, enhances CAR T cell expansion, persistence, and anti-tumor activity).
In the most general terms, the principle of CAR-T therapy is that T-lymphocytes are taken from patients or donors and their main receptor (TCR), designed to recognize foreign cells, is replaced with an artificially created (CAR) specific to a given tumor. After that, the genetically modified cells are injected into the patient.
In preclinical trials, this technique has made it possible to achieve long-term remission and even complete resorption of tumors in patients with certain oncological (mainly hematological) diseases. In 2017, the US Food and Drug Administration (FDA) approved its clinical use for the first time, as of June 2022, it has registered four CAR-T-lymphocyte preparations: axicabtagen ciloleucel, tisagenleucel, brexucabtagen autoleucel and lysocabtagen maraleucel. All of them are intended for the treatment of B-cell lymphoma. This is due to the presence of CD19 antigen inherent only in B-lymphocytes, which serves as a convenient target for therapy, ensuring its effectiveness.
Experiments on CAR-T therapy of other oncological diseases often demonstrate an insufficiently persistent effect, which is due to the gradual depletion of CAR-T lymphocytes, in which they lose effector functions and express inhibitory receptors, and the death of these cells.
To increase the functionality and survival of antitumor lymphocytes, the staff of Washington University in St. Louis, led by John DiPersio, used a modified recombinant human interleukin-7 (responsible for the maturation and survival of lymphocytes), hybridized with a stable biological platform in the form of a chimeric Fc fragment of immunoglobulin (prolongs the effect and prevents an undesirable immune reaction). A similar drug, known as rhIL-7-hyFc, NT-I7 or efineptakin alpha, is currently undergoing the first phase of clinical trials in combination with pembrolizumab for tumor immunotherapy.
At the first stage of the work, the researchers cultured CD19+ tumor cells with anti-CD19 CAR-T lymphocytes (UCART19) and different concentrations of rhIL-7-hyFc (10, 100 and 1000 nanograms per milliliter plus a control group) in a nutrient medium. Without the drug, immunotherapeutic cells stopped multiplying by the end of the second week of the experiment, adding it to the culture prevented the death of UCART19, stimulated their proliferation and maintained polyfunctionality, especially at concentrations above 10 nanograms per milliliter.
To evaluate the effectiveness in vivo, NSG immunodeficient mice with vaccinated human tumors (B-cell lymphoma and CD33+ acute myeloid leukemia) and ordinary animals with mouse CD19+ lymphoma were used. All of them were treated with "profile" CAR-T lymphocytes (UCART19, UCART33 and mCART19, respectively). On the 1st, 15th and 29th days of therapy, they were injected with 10 nanograms of rhIL-7-hyFc per kilogram of body weight and compared with the control group.
In all cases, the modified interleukin-7 ensured stable reproduction of CAR-T lymphocytes throughout the observations, as well as increased their efficiency (cytotoxicity) and reduced depletion.
In the presence of the drug, all animals with human oncological diseases remained alive and, in most cases, free of tumors at the end of the experiment (day 175, which is about one fifth of the average life expectancy of a laboratory mouse), while without it (only with CAR-T) they lived up to a maximum of day 40, and without treatment, they died within two weeks.
In the experiment with mouse lymphoma, 4 out of 4 animals who received mCART19 with rhIL-7-hyFc survived until its end (more than 100 days), 3 out of 4 received only mCART19, and 1 out of 4 in two control groups.
"Enhanced" CAR-T lymphocytes (green) and macrophages (red) attack mouse cancer cells (blue). Michael Rettig / Bursky Center for Human Immunology & Immunotherapy Programs.
Additional experiments have shown that rhIL-7-hyFc also reduces the minimum effective dose of CAR-T lymphocytes, while increasing their quantity and quality in vivo.
If the results obtained can be reproduced in humans, the technique can be applied in the foreseeable future, since, as mentioned above, rhIL-7-hyFc is already undergoing clinical trials, albeit for a different indication.
Previously, different laboratories managed to maintain the functionality and prolong the life of CAR-T lymphocytes, giving them a kind of "rest", turning off the synthesis of the regulatory protein Cbl-b and activating the non-canonical signaling pathway of the transcription factor NF-kB.
The experimental use of CAR-T cells includes not only the treatment of a wide range of oncological diseases, but also the therapy of viral infections, fibrosis and autoimmune diseases, as well as countering cellular aging.
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