CAR T cell

In biology, chimeric antigen receptors —also known as chimeric immunoreceptors, chimeric T cell receptors or artificial T cell receptors—are receptor proteins that have been engineered to give T cells the new ability to target a specific antigen. The receptors are chimeric in that they combine both antigen-binding and T cell activating functions into a single receptor.
CAR T cell therapy uses T cells engineered with CARs to treat cancer. T cells are modified to recognize cancer cells and destroy them. The standard approach is to harvest T cells from patients, genetically alter them, then infuse the resulting CAR T cells into patients to attack their tumors.
CAR T cells can be derived either autologously from T cells in a patient's own blood or allogeneically from those of a donor. Once isolated, these T cells are genetically engineered to express a specific CAR, using a vector derived from an engineered lentivirus such as HIV. The CAR programs the T cells to target an antigen present on the tumor cell surface. For safety, CAR T cells are engineered to be specific to an antigen that is expressed on a tumor cell but not on healthy cells.
After the modified T cells are infused into a patient, they act as a "living drug" against cancer cells. When they come in contact with their targeted antigen on a cell's surface, T cells bind to it and become activated, then proceed to proliferate and become cytotoxic. CAR T cells destroy cells through several mechanisms, including extensive stimulated cell proliferation, increasing the degree to which they are toxic to other living cells, and by causing the increased secretion of factors that can affect other cells such as cytokines, interleukins and growth factors.
The surface of CAR T cells can bear either of two types of co-receptors, CD4 and CD8. These two cell types, called CD4+ and CD8+, respectively, have different and interacting cytotoxic effects. Therapies employing a 1-to-1 ratio of the cell types apparently provide synergistic antitumor effects.

History

The first chimeric receptors containing portions of an antibody and the T cell receptor were described in 1987 by Yoshihisa Kuwana et al., at Fujita Health University and Kyowa Hakko Kogyo, Co. Ltd., in Japan, and independently in 1989 by Gideon Gross and Zelig Eshhar at the Weizmann Institute in Israel. Originally termed "T-bodies", these early approaches combined an antibody's ability to specifically bind to diverse targets with the constant domains of the TCR-α or TCR-β proteins.
In 1991, chimeric receptors containing the intracellular signaling domain of CD3ζ were shown to activate T cell signaling by Arthur Weiss at the University of California, San Francisco. This work prompted CD3ζ intracellular domains to be added to chimeric receptors with antibody-like extracellular domains, commonly single-chain fraction variable domains, as well as proteins such as CD4, subsequently termed first generation CARs.
A first generation CAR containing a CD4 extracellular domain and a CD3ζ intracellular domain was used in the first clinical trial of chimeric antigen receptor T cells by the biotechnology company Cell Genesys in the mid 1990s, allowing adoptively transferred T cells to target HIV infected cells, although it failed to show any clinical improvement. Similar early clinical trials of CAR T cells in solid tumors in the 1990s using first generation CARs targeting a solid tumor antigens such as MUC1 did not show long-term persistence of the transferred T cells or result in significant remissions.
In the early 2000s, co-stimulatory domains such as CD28 or 4-1BB were added to first generation CAR's CD3ζ intracellular domain. Termed second generation CARs, these constructs showed greater persistence and improved tumor clearance in pre-clinical models. Clinical trials in the early 2010s using second generation CARs targeting CD19, a protein expressed by normal B cells as well as B-cell leukemias and lymphomas, by investigators at the NCI, University of Pennsylvania, and Memorial Sloan Kettering Cancer Center demonstrated the clinical efficacy of CAR T cell therapies and resulted in complete remissions in many heavily pre-treated patients. These trials ultimately led in the US to the FDA's first two approvals of CAR T cells in 2017, those for tisagenlecleucel, marketed by Novartis originally for B-cell precursor acute lymphoblastic leukemia, and axicabtagene ciloleucel, marketed by Kite Pharma originally for diffuse large B-cell lymphoma. There are now six FDA-approved CAR T therapies.

Production

The first step in the production of CAR T-cells is the isolation of T cells from human blood. CAR T-cells may be manufactured either from the patient's own blood, known as an autologous treatment, or from the blood of a healthy donor, known as an allogeneic treatment. The manufacturing process is the same in both cases; only the choice of initial blood donor is different.
First, leukocytes are isolated using a blood cell separator in a process known as leukocyte apheresis. Peripheral blood mononuclear cells are then separated and collected. The products of leukocyte apheresis are then transferred to a cell-processing center. In the cell processing center, specific T cells are stimulated so that they will actively proliferate and expand to large numbers. To drive their expansion, T cells are typically treated with the cytokine interleukin 2 and anti-CD3 antibodies. Anti-CD3/CD28 antibodies are also used in some protocols.
The expanded T cells are purified and then transduced with a gene encoding the engineered CAR via a retroviral vector, typically either an integrating gammaretrovirus or a lentiviral vector. These vectors are very safe in modern times due to a partial deletion of the U3 region. The new gene editing tool CRISPR/Cas9 has recently been used instead of retroviral vectors to integrate the CAR gene into specific sites in the genome.
The patient undergoes lymphodepletion chemotherapy prior to the introduction of the engineered CAR T-cells. The depletion of the number of circulating leukocytes in the patient upregulates the number of cytokines that are produced and reduces competition for resources, which helps to promote the expansion of the engineered CAR T-cells.

Delivery

Clinical applications

As of March 2019, there were around 364 ongoing clinical trials happening globally involving CAR T cells. The majority of those trials target blood cancers: CAR T therapies account for more than half of all trials for hematological malignancies. CD19 continues to be the most popular antigen target, followed by BCMA. In 2016, studies began to explore the viability of other antigens, such as CD20. Trials for solid tumors are less dominated by CAR T, with about half of cell therapy-based trials involving other platforms such as NK cells.

Cancer

T cells are genetically engineered to express chimeric antigen receptors specifically directed toward antigens on a patient's tumor cells, then infused into the patient where they attack and kill the cancer cells. Adoptive transfer of T cells expressing CARs is a promising anti-cancer therapeutic, because CAR-modified T cells can be engineered to target potentially any tumor associated antigen.
Early CAR T cell research has focused on blood cancers. The first approved treatments use CARs that target the antigen CD19, present in B-cell-derived cancers such as acute lymphoblastic leukemia and diffuse large B-cell lymphoma. There are also efforts underway to engineer CARs targeting many other blood cancer antigens, including CD30 in refractory Hodgkin's lymphoma; CD33, CD123, and FLT3 in acute myeloid leukemia ; and BCMA in multiple myeloma. Aside from CD19, CARs targeting the multiple myeloma antigen B-cell maturation antigen have achieved the most clinical success so far. CARs targeting BCMA were initially reported by Robert Carpenter and James Kochenderfer et al. Anti-BCMA CAR T cells have now been tested in many clinical trials, and anti-BCMA CAR T-cell products have been approved by the U.S. Food and Drug Administration.
CAR T cells have also been found to be effective in treating glioblastoma. A single infusion is enough to show rapid tumor regression in a matter of days.
Solid tumors have presented a more difficult target. Identification of good antigens has been challenging: such antigens must be highly expressed on the majority of cancer cells, but largely absent on normal tissues. CAR T cells are also not trafficked efficiently into the center of solid tumor masses, and the hostile tumor microenvironment suppresses T cell activity.

Autoimmune disease

While most CAR T cell studies focus on creating a CAR T cell that can eradicate a certain cell population, other potential uses for this technology have emerged. T cells can mediate tolerance to antigens. A regulatory T cell outfitted with a CAR could confer tolerance to a specific antigen, which could be utilized in organ transplantation or rheumatologic diseases like lupus. It has been investigated as a treatment for ulcerative colitis and rheumatoid arthritis. Clinical trials have launched for systemic sclerosis, myositis and rheumatoid arthritis.

Approved therapies

CAR T cell	Company	Approval Agency: Date	Target	Antigen recognition domain	Intracellular signaling domain	Indication	Agency Product Number, Drug Label
tisagenlecleucel	Novartis	FDA: 08/30/2017 EMA: 08/22/2018 MHLW: 05/15/2019	CD19	scFV	41BB - CD3ζ	B-cell precursor ALL Diffuse large B-cell lymphoma Follicular lymphoma	FDA:125646, EMA:004090,
axicabtagene ciloleucel	Kite Pharma / Gilead	FDA: 10/18/2017 EMA: 08/27/2018 NMPA: 06/23/2021 MHLW: 12/22/2022	CD19	scFV	CD28 - CD3ζ	Diffuse large B-cell lymphoma Follicular lymphoma Primary mediastinal large B-cell lymphoma	FDA:125643, EMA:004480,
brexucabtagene autoleucel	Kite Pharma / Gilead	FDA: 07/24/2020 EMA: 12/14/2020	CD19	scFV	CD28 - CD3ζ	Mantle cell lymphoma B-cell precursor ALL	FDA:125703, EMA:005102,
lisocabtagene maraleucel	Juno Therapeutics / BMS	FDA: 02/05/2021 EMA: 04/04/2022 MHLW: 12/20/2022	CD19	scFV	41BB - CD3ζ	Diffuse large B-cell lymphoma	FDA: 25714, EMA:004731,
idecabtagene vicleucel	BMS	FDA: 03/26/2021 EMA: 08/18/2021	BCMA	scFV	41BB - CD3ζ	Multiple myeloma,	FDA:125736, EMA:004662,
ciltacabtagene autoleucel	Janssen / J&J	FDA: 02/28/2022 EMA: 05/25/2022	BCMA	VHH	41BB - CD3ζ	Multiple myeloma,	FDA:125746, EMA:005095,
obecabtagene autoleucel	Autolus	FDA: 11/08/2024	CD19	scFV	41BB - CD3ζ	B-cell precursor ALL	FDA: 125813