Projects / Programmes
Regulation of the therapeutic effect of CAR-T cells for increased safety and efficiency
| Code |
Science |
Field |
Subfield |
| 3.04.00 |
Medical sciences |
Oncology |
|
| Code |
Science |
Field |
| B000 |
Biomedical sciences |
|
| Code |
Science |
Field |
| 3.02 |
Medical and Health Sciences |
Clinical medicine |
CD19-CAR T cells, NFAT2 based arteficial transcription factors, proliferation regulation, B cell lymphoma, heterodimeriaztion systems
Researchers (1)
| no. |
Code |
Name and surname |
Research area |
Role |
Period |
No. of publicationsNo. of publications |
| 1. |
34069 |
PhD Duško Lainšček |
Biotechnology |
Head |
2018 - 2020 |
148 |
Organisations (1)
| no. |
Code |
Research organisation |
City |
Registration number |
No. of publicationsNo. of publications |
| 1. |
0104 |
National Institute of Chemistry |
Ljubljana |
5051592000 |
21,007 |
Abstract
CAR-T cells present a revolutionary therapeutic option for various malignant diseases based on their ability to specifically recognize the selected tumour surface markers in MHC independent manner, triggering cell activation and cytokine production that results in killing cancerous cell expressing surface markers recognized by the chimeric antigen receptor (CAR). CAR T-cells are highly potent serial killers that may however cause serious side effects so their activity needs to be carefully controlled.
CARs are typically composed of an extracellular antigen-recognition domain (most often single-chain variable fragment (scFV) derived from a monoclonal antibody or antigen-binding fragment (Fab)), a transmembrane domain, which is connected to an intracellular signaling domain, containing CD3ζ domain linked with one or two costimulatory domains, required for the second generation T cell activation.
The main therapeutic goal is a specific T cell activation with sufficient destruction of tumorous cells in a safe manner. In order to increase T cell activation, 4-1BB to prolong T cell survival and CD28 to increase the potency of T cell response, was introduced into the next generation CAR-T cells . In some cases the higher number of infused CARs resulted in greater cytokine production leading towards the cytokine release syndrome (CRS) and other toxicities, which can be treated with some specific drugs, like IL-6 antibodies or with high dosage of corticosteroids, but this can result in CAR T cell depletion. In order to avoid CRS, sometimes lower numbers of CARs are introduced into patients or several infusions are made, which again can end either in no therapeutic effect or in severe CRS. Several attempts were made to influence CAR T cell number but still no such switch is developed that can effectively regulate infused CAR T cell number.
In this project we propose to design a mechanism to control proliferation and activation state of the infused CARs by using artificial transcription factor that could be controlled by the physician treating the patient. The goal of the proposed project is to contribute to the development of more efficient and safer CAR T cell-based cancer immunotherapy by the design and experimental verification of a novel strategy for the control of proliferation and activation state of CAR-T cells based on the engineered key endogenous NFAT (Nuclear factor of activated T cells) transcription factor, which can be regulated by small chemical regulators.
We plan to produce stably expressing CD19-CAR T cell line by utilizing different chemically induced heterodimerization domains. We will virally transduce T cells that will be isolated from PBMC which will be obtained within the collaboration with the Blood Transfusion Centre of Slovenia. We will design tNFAT21-593 (NFAT2 without its own transactivation domain) artificial transcription factors which will upon inducing heterodimerization bind to its protein counterparts, fused either with the activator or repressor domains. This will result in regulated gene transcription, leading to higher/lower IL2 production and afterwards also to higher/lower CD19-CAR T cell proliferation. Based on this chemically regulated proliferation we will influence CD19 CAR T cell proliferation and action. The effect of this novel regulation of CAR T cells will be tested in vitro on different B cell cancerous cells (Raji, BCWM cells). Therapeutic use and also increased safety towards lowering the risk of CRS development will be tested in vivo on animal cancer model as well.
Significance for science
Current cancer therapy based on CAR-T cells that is already in the clinical use relies basically on unregulated action of T cells within the human body after infusion of the cells. Although several small-molecule growth and action regulators have been proposed, there is still no switch available that could regulate both activated and resting state of therapeutic cells, which means that there is no negative regulator of T cell growth available. Additionally, in contrast to the regulation of CAR assembly we plan to engineer an endogenous signalling pathway of T cells, which is likely to have stronger effect. The proposed projects proposes development of a new CD-19 CAR-T cell line whose growth, proliferation and activation can be regulated by small molecules so the number of CAR-T cells can be also negatively regulated if required. This new approach of regulating the number of circulating CAR T cells addresses an aspect towards increasing the safety of this therapy-decreasing the risk of CRS.
CAR T cell therapy is probably the hottest therapeutic method, with many advantages but also many dangerous aspects, such as CRS. Therefore any improvement of the safety and efficiency of this therapeutic approach may offer significant regards in terms of a rapid translation into therapeutic application. If CAR T cell therapy becomes safer and more reproducible it will likely be implemented for increasing number of patients at earlier stage and also for additional indications. Based on engineered NFAT-driven regulation of CAR T cell proliferation and activation, as proposed in this projects, physicians could regulate the number of T cells and adjust the response for each patient by a simple small drug infusion (without killing therapeutic cells as in some published safety switches), so CRS could be avoided, leading to reduced number of fatal complications in cancer CAR-T cell based therapy.
Significance for the country
Current cancer therapy based on CAR-T cells that is already in the clinical use relies basically on unregulated action of T cells within the human body after infusion of the cells. Although several small-molecule growth and action regulators have been proposed, there is still no switch available that could regulate both activated and resting state of therapeutic cells, which means that there is no negative regulator of T cell growth available. Additionally, in contrast to the regulation of CAR assembly we plan to engineer an endogenous signalling pathway of T cells, which is likely to have stronger effect. The proposed projects proposes development of a new CD-19 CAR-T cell line whose growth, proliferation and activation can be regulated by small molecules so the number of CAR-T cells can be also negatively regulated if required. This new approach of regulating the number of circulating CAR T cells addresses an aspect towards increasing the safety of this therapy-decreasing the risk of CRS.
CAR T cell therapy is probably the hottest therapeutic method, with many advantages but also many dangerous aspects, such as CRS. Therefore any improvement of the safety and efficiency of this therapeutic approach may offer significant regards in terms of a rapid translation into therapeutic application. If CAR T cell therapy becomes safer and more reproducible it will likely be implemented for increasing number of patients at earlier stage and also for additional indications. Based on engineered NFAT-driven regulation of CAR T cell proliferation and activation, as proposed in this projects, physicians could regulate the number of T cells and adjust the response for each patient by a simple small drug infusion (without killing therapeutic cells as in some published safety switches), so CRS could be avoided, leading to reduced number of fatal complications in cancer CAR-T cell based therapy.
Most important scientific results
Final report
Most important socioeconomically and culturally relevant results
Final report
