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嵌合抗原受体T细胞有望成为癌症治疗的“第五支柱”

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CAR T-cells, a Promising “Fifth Pillar” of Cancer Treatment

    Immunotherapy, or the idea of guiding patients’ own immune systems to target malignant cells, has emerged as a promising method of cancer treatment. Following the traditional methods of surgery, chemotherapy, and radiation therapy, as well as the relatively recent method of targeted pharmaceutical therapy, immunotherapy is now seen as a possible “fifth pillar” of cancer treatment (1). The basic premise of immunotherapy is to enhance our naturally existing defence mechanisms, namely the lymphocytes of the immune system, in the hopes of eradicating cancerous cells. One form of immunotherapy that has generated exciting results involves genetically engineered T lymphocytes, or T-cells, that can specifically target and attack cancerous cells (2). These genetically engineered T-cells express chimeric antigen receptors (CARs) that can bind to specific markers, or antigens, on malignant cells. This article hopes to map the path of CAR T-cell therapy from primary research laboratories to potential commercialization, as well as some of the challenges it faces on the way.

    CAR T-cell therapy has its roots extended back to the 1960s when scientists discovered that cells in the immune system can kill tumour cells in mice (3). Since then, the rapid development of genetic engineering techniques has allowed researchers to turn the idea of targeting tumour cell with our own immune system into practice. In 1989, Israeli immunologist Zelig Eshhar and colleges successfully fused parts of T-cell receptors (TCRs), or antigen receptors expressed on the surface of T-cells, with parts of an antibody (4). This design is later known as first-generation CARs. The intracellular signalling domain of first-generation CARs contained only one activation domain. Unfortunately, this design was shown to be ineffective in vivo (5). Second-generation CARs emerged in the early 2000s. They include additional costimulatory molecules to strengthen the T-cells’ capacity to proliferate and to produce cytokines (2,5). Following the successful results of several clinical trials, CAR T-cell therapy was proved by the U.S. Food and Drug Administration (FDA) to be a treatment for certain types of B-cell lymphoma in 2017 (6).

    Without a thorough understanding of the molecular mechanisms behind CAR T-cell actions, any potential applications are only mirages. Following is a brief introduction to the molecular principles underlying CAR T-cell actions. T-cells normally express TCRs that bind to specific antigens in a “lock and key” fashion. A typical TCR consists of two functional domains: an extracellular antigen-binding domain and an intracellular signalling domain. In a genetically engineered CAR, the extracellular antigen binding domain is replaced by an antibody-derived domain, usually a single-chain fragment variable (scFv), that specifically targets tumour cells (5). If we think of tumour antigens, or molecular structures on tumour cells, as a unique lock, we can think of CAR T-cells as being artificially armed with a specific key to that lock. Once the lock and the key form a match, malignant cells expressing the “lock” will be targeted for destruction.

    Some beneficial characteristics of CAR T-cell therapy have caught the attention of researchers, physicians, and patients alike. A normal TCR is only capable of recognizing peptide antigens presented by major histocompatibility complexes (MHCs), a group of cell surface proteins that specialize in processing and displaying antigen-derived proteins (7). In other words, the function of a normal TCR is MHC-dependent. However, CARs recognize their antigens via the antibody-derived domain, which operates in an MHC-independent manner. This MHC-independent characteristic of CAR T-cell activity is significant for cancer treatment because many tumour cells escape immunosurveillance by suppressing MHC expression (2). In other words, CAR T-cells are able to detect the presence of cancerous cells that would otherwise be hidden from the immune system. An additional benefit that CAR T-cell therapy offers is personalized treatments based on specific circumstances of the patients owing to the variability of CARs and the fact that CAR T-cell therapy utilizes the patients’ own immune systems.

    In translating research results into viable cancer therapy, successes have been mixed with tragedies during clinical trials. So far, successes have been seen in targeting CAR T-cells to malignant B lymphocytes, or B-cells, at different developing stages (8–10). B-cells prove to be an easy target for CARs due to the presence of distinctive markers, or “locks”, such as CD19 (2,5). However, CAR T-cells that recognize these markers not only destroy malignant B-cells, but also the healthy ones, resulting in a condition known as B-cell aplasia. Low counts or even complete eradication of B-cells can lead to inadequate immune responses upon infection. This situation can be treated with immunoglobin replacement therapy, yet it is still possible that B-cell aplasia can have long-term complications (5). There is evidence that CAR T-cells can be effective in treating some other forms of lymphoma, however, to achieve complete responses, high levels of conditioning are needed. The conditioning procedure includes treating patients with high doses of chemotherapy, which can be highly toxic to patients (5). The difficulties in treating these types of lymphomas may be due to the fact that they involve solid tumours that are hard to target with CAR T-cells (5). Even more worrying, in one clinical trial conducted by Juno Therapeutics, five patients succumbed to severe brain swelling. There was no clear explanation for the cause of this tragedy. Although researchers had found that CAR T-cell therapy can have adverse effects on neurological functions, brain swelling was not one of the suspected side-effects (11).

    In addition to fatal brain swelling, CAR T-cell therapy has also been shown to have other severe side-effects. One of the most severe side-effects is cytokine release syndrome (CRS), resulting from high levels of cytokines circulating through the body. Cytokines are molecules released by T-cells to assist their immune functions. CRS is associated with serious symptoms such as low blood pressure, capillary leakage, and even cardiac arrest (12). As mentioned above, neurologic toxicities of CAR T-cell therapy have been noticed during clinical trials. The neurotoxic effects of CAR T-cell therapy can range from confusion, language impairments, and sometimes seizures (11). The underlying causes of these adverse neurological effects are still under investigation. Furthermore, CAR T-cells themselves may evoke violent allergic reactions upon infusion into the patients’ bloodstream, a condition known as anaphylaxis. Anaphylaxis can be life-threatening if not treated immediately (12).

    Another challenge comes from the high costs associated with CAR T-cell therapy. One round of CAR T-cell therapy usually follows the steps stated below. First, T-cells are extracted from a patient’s blood. The extracted T-cells are then genetically engineered to express CARs designed for the patient’s condition (12). Genes that encode the designed CARs are often introduced to the T-cells using retrovirus vectors. An alternative way is to use transposons, or genes that can insert themselves into certain positions on the genome, to deliver the CAR genes (2). The genetically engineered cells can now be called CAR T-cells. Next, these CAR T-cells are cultured in laboratories to expand their numbers. The final step is to infuse the CAR T-cells into the patients. If successful, the CAR T-cells will continue to multiply in the patients’ bloodstream and attack their designated targets. Each step in the procedure either involved sophisticated protocols requiring highly-trained personnel and well-equipped laboratories, or extensive care and monitoring of the patients (13). Health Canada approved the first CAR T-cell therapy, named Kymriah, in September 2018, however, no patients outside of clinical trials have received it (14). In the United States, not all insurance programs cover the high costs of CAR T-cell therapy, yet the cost can reach as high as US$475,000 for just one round of treatment (14). We are not sure how much, if any, of the extremely high costs will be covered by the government once CAR T-cell therapy proceeds to full commercialization. If the costs of CAR T-cell therapy remain thus high, it will likely be inaccessible to the majority of patients’ families.

    These challenges may remain problematic for a while, but scientists have already been working on improving the safety and effectiveness of CAR T-cell therapy, as well as lowering the costs to manufacture. For example, one team of scientists aims to enhance the level of control over the place and time of activation of the genetically engineered T-cells (15). Others set out to program the residual CAR T-cells to self-destruct when the appropriate time has come to prevent unintended damage to the patients (16). There are also efforts towards building a type of CAR T-cells that can be universally applied in order to increase the accessibility of the therapy (2).

    In sum, the application of CAR T-cells in cancer treatment is still in its infancy, there are still many concerns and uncertainties that need to be addressed. However, with the successes already seen in treating blood cancers and preliminary research into targeting solid tumours, CAR T-cell therapy has the potential of revolutionizing cancer treatment in that complete eradication of cancerous cells may be achieved. The road of CAR T-cells from primary research to commercialization has not been without obstacles and probably will not be in the future, but if we continue to proceed with caution, CAR T-cell treatment may one day be more than just a last resort and become the true “fifth pillar” of cancer treatment.

References

1. National Cancer Institute. CAR T Cells: Engineering Patients’ Immune Cells to Treat Their Cancers [Internet]. National Cancer Institute. 2017 [cited 2019 Feb 23]. Available from: https://www.cancer.gov/about-cancer/treatment/research/car-t-cells

2. Gomes-Silva D, Ramos CA. Cancer Immunotherapy Using CAR-T Cells: From the Research Bench to the Assembly Line. Biotechnology Journal. 2018 Feb;13(2):1700097. 

3. Memorial Sloan Kettering Cancer Center. CAR T Cells: Timeline of Progress [Internet]. Memorial Sloan Kettering Cancer Center. [cited 2019 Feb 23]. Available from: https://www.mskcc.org/timeline/car-t-timeline-progress

4. Gross G, Waks T, Eshhar Z. Expression of immunoglobulin-T-cell receptor chimeric molecules as functional receptors with antibody-type specificity. Proceedings of the National Academy of Sciences. 1989 Dec 1;86(24):10024–8. 

5. Enblad G, Karlsson H, Loskog ASI. CAR T-Cell Therapy: The Role of Physical Barriers and Immunosuppression in Lymphoma. Human Gene Therapy. 2015 Aug;26(8):498–505. 

6. FDA News Release. FDA approves CAR-T cell therapy to treat adults with certain types of large B-cell lymphoma [Internet]. U.S. Food and Drug Administration. 2017 [cited 2019 Feb 23]. Available from: https://www.fda.gov/newsevents/newsroom/pressannouncements/ucm581216.htm

7. Wieczorek M, Abualrous ET, Sticht J, Álvaro-Benito M, Stolzenberg S, Noé F, et al. Major Histocompatibility Complex (MHC) Class I and MHC Class II Proteins: Conformational Plasticity in Antigen Presentation. Frontiers in Immunology [Internet]. 2017 Mar 17 [cited 2019 Feb 23];8. Available from: https://journal.frontiersin.org/article/10.3389/fimmu.2017.00292/full

8. Porter DL, Levine BL, Kalos M, Bagg A, June CH. Chimeric Antigen Receptor–Modified T Cells in Chronic Lymphoid Leukemia. New England Journal of Medicine. 2011 Aug 25;365(8):725–33. 

9. Davila ML, Riviere I, Wang X, Bartido S, Park J, Curran K, et al. Efficacy and Toxicity Management of 19-28z CAR T Cell Therapy in B Cell Acute Lymphoblastic Leukemia. Science Translational Medicine. 2014 Feb 19;6(224):224ra25-224ra25. 

10. Kochenderfer JN, Dudley ME, Kassim SH, Somerville RPT, Carpenter RO, Stetler-Stevenson M, et al. Chemotherapy-Refractory Diffuse Large B-Cell Lymphoma and Indolent B-Cell Malignancies Can Be Effectively Treated with Autologous T Cells Expressing an Anti-CD19 Chimeric Antigen Receptor. Journal of Clinical Oncology. 2015 Feb 20;33(6):540–9. 

11. Couzin-Frankel J. Worries, confusion after cancer trial deaths. Science. 2016 Dec 9;354(6317):1211–1211. 

12. Leukemia & Lymphoma Society. Chimeric Antigen Receptor (CAR) T-cell Therapy [Internet]. Leukemia & Lymphoma Society. 2018 [cited 2019 Feb 23]. Available from: https://www.lls.org/treatment/types-of-treatment/immunotherapy/chimeric-antigen-receptor-car-t-cell-therapy

13. Kunert A, Straetemans T, Govers C, Lamers C, Mathijssen R, Sleijfer S, et al. TCR-Engineered T Cells Meet New Challenges to Treat Solid Tumors: Choice of Antigen, T Cell Fitness, and Sensitization of Tumor Milieu. Frontiers in Immunology [Internet]. 2013 [cited 2019 Mar 2];4. Available from: https://journal.frontiersin.org/article/10.3389/fimmu.2013.00363/abstract

14. Grant K. Uncertain, costly, but filled with hope: Gene therapy about to go mainstream in Canada [Internet]. The Globe and Mail. 2018 [cited 2019 Feb 23]. Available from: https://www.theglobeandmail.com/canada/article-uncertain-costly-but-filled-with-hope-gene-therapy-about-to-go/

15. Roybal KT, Williams JZ, Morsut L, Rupp LJ, Kolinko I, Choe JH, et al. Engineering T Cells with Customized Therapeutic Response Programs Using Synthetic Notch Receptors. Cell. 2016 Oct;167(2):419-432.e16. 

16. Hamers L. How to make CAR-T cell therapies for cancer safer and more effective [Internet]. ScienceNews. 2018 [cited 2019 Feb 23]. Available from: https://www.sciencenews.org/article/how-make-car-t-cell-therapies-cancer-safer-and-more-effective