CAR T-Cell Therapy: An Emerging Frontier in The Fast Lane

CAR T-Cells: Chimeric Antigen Receptor (CAR) T-cells are a new frontier of cancer therapy and have shown a tremendous clinical performance in pediatric Acute Lymphoblastic Leukemia (ALL) and non-Hodgkin Lymphoma patients. The therapy has shown potential to expand to other forms of cancer, including solid tumors. CAR T-cells are T cells that have been collected from a patient and engineered using viral transduction methods to express CARs. CAR consists of a cytoplasmic signaling domain, transmembrane domain and an extracellular antigen recognition domain2 (see Figure 1). CAR T-cells are expanded ex vivo and reinfused into the patient1.

Figure 1:    CAR T-cells contain a molecular construct (CAR) that makes them specific for ALL cells.

Figure 1: CAR T-cells contain a molecular construct (CAR) that makes them specific for ALL cells.

Advances in cellular engineering allow a continuous production of T-cells following infusion into the patient (expansion) and improve the time that CAR T-cells survive in the circulation (persistence)1. Early attempts took several weeks to generate a batch of CAR T-cells, this time has been reduced by several labs to 7 days1.

The CAR complex presented on a T-cell is designed to identify and bind to specific targets on a tumor, such as glycolipids, carbohydrates and proteins (i.e. CD19 on B-cells). Upon binding, CAR T-cells become activated, causing cytokine secretion and T-cell proliferation. A tumor recognized by CAR T-cells is therefore targeted for elimination by the immune system2 (see Figure 2).

Figure 2:   CAR T-cells are made of T-cells separated from a patient’s blood. They are engineered to express CAR, and reinfused into the patient to target ALL cells.

Figure 2: CAR T-cells are made of T-cells separated from a patient’s blood. They are engineered to express CAR, and reinfused into the patient to target ALL cells.

 Relevant Clinical Trials: Early experimental success motivated further studies, leading to two approved single-use therapies - tisagenlecleucel (KymriahTM) and axicabtagene ciloleucel (YescartaTM). Kymriah is a CAR T-cell therapy targeting CD19 in children and adolescents suffering from ALL, which is the most common form of cancer in children and in over 80% of cases the disease arises in B cells1. First-line treatment is intensive chemotherapy. However, upon relapse following a stem cell transplant or chemotherapy, there are very few options of further treatment, according to Stephan Grupp (M.D., PhD) – an expert from the Children’s Hospital of Philadeplphia (CHOP)1. As a result, ALL relapse is the leading cause of death in childhood cancer. Kymriah was initially approved by the FDA in August 2017 due to very promising clinical trial results4. Updated analysis of 75 Kymriah-infused patients (Pivotal Phase 2 trial ELIANA by Novartis) with ≥3 months follow-up has shown an 81% overall remission rate. 6 months relapse-free survival was 80%, whereas event-free survival at 6 months was 73%, and 50% at 12 months. Overall survival in the cohort of 75 patients was 90% at 6 months, and 76% at 12 months3.

Yescarta was approved in October 2018. Initially, a small trial led by National Cancer Institute (NCI), showed that Yescarta generated complete responses in half of the advanced diffuse large B-cell lymphoma patients. James Kochenderfer (M.D., NCI), the trial’s lead investigator, stated that the data gave a first real glimpse of the potential the approach has in treating patients with aggressive lymphomas1. Importantly, up until now such cases were ‘’virtually untreatable’’1. Kite Pharmaceuticals (who hold a research agreement with NCI for development of ACT-based therapies) funded a further larger trial called ZUMA-1. By employing 101 aggressive non-Hodgkin lymphoma patients, ZUMA-1 confirmed the earlier findings, giving a 72% response rate after a single infusion and 51% complete remission at 7.9 months median follow-up, serving as a basis for the FDA approval5.

Side Effects: Side effects include B-cell aplasia (therapy killing off healthy B cells), cerebral edema (swelling in the brain: very rare yet encountered), and foremost cytokine release syndrome (CRS)1. CRS is a rapid mass cytokine release into the bloodstream, which could lead to very high fever as well as a precipitous drop in blood pressure. It is considered an on-target side effect of the CAR T-cell therapy, demonstrating that active T-cells are functioning in the body. The research team at CHOP observed that IL-6 cytokine levels were increased in patients suffering from CRS. Tocilizumab (Actemra®), a drug used to treat inflammatory conditions through IL-6 blockage, is the standard therapy in severe CRS management1.

Commercial Performance: Kymriah quarterly sales, as of April 2018, were rather disappointing, with the treatment amassing $12 million against the predicted $159 million. Problematically, thus far the drug was limited to a relatively small population, with 3,100 new patients each year. Kymriah targets only about 30% of these, which are patients responsive to the therapy within one month, with the treatment cost of $475,000 per patient6. Yescarta, on the other hand, costs $373,000 and Gilead, the owners of Kite Pharma, have secured support from Medicare, where Medicare will cover a part of the costs of the treatment7. However, Kymriah is currently debated to decrease its price per treatment down to $160,000 and it has recently gained further approval, expanding its clinical use to relapsed large B-cell lymphoma.

Future Prospects: The recent success and potential therapeutic use has become evident. The numbers of clinical trials focusing on CAR T-cells grew from 5 to 1801. The majority of trials focus on CD19-targeted CAR T-cells. However, a portion of patients are unresponsive to this treatment, and in many cases ALL cells eventually stop expressing CD19, leading to relapse. With this, CD22 is an increasingly more popular target for CAR T-cell therapy, together with therapies that target several CD proteins (i.e. CD19, CD22, CD123)1. Such approaches are hoped to prolong the effectiveness and improve the efficacy of CAR T-cell therapy by targeting several key antigens simultaneously.

Endeavors are being made to adapt CAR T-cell therapy to solid tumors1. However, this is proving difficult due to a lack of unique antigens. For example, clinical trials are being conducted using CAR T-cells targeting mesothelin, a protein overexpressed in cases of lung and pancreatic cancers. EGFRvIII found in the majority glioblastoma cells is another such case. These, however, have not shown the success displayed with ALL trials. Solid tumors are surrounded by a microenvironment that blunts immune responses1. It seems that to successfully target solid tumors, a next-generation CAR T-cell therapy is needed, targeting specific antigens of the tumor microenvironment.

Laboratory Resources: CAR T-cell preparation is a multi-step process requiring a variety of aperture, all of which is commercially available from different manufacturers. Initially, the leukapheresis process allows the removal of leukocytes from patient’s blood. Lymphocyte-enriched apheresis is obtained through cell wash and counterflow centrifugal elutriation9. The technology is also useful at later stages of T-cell formulation. Here, machines such as Cell Saver 5+, Terumo Elutra, COBE2991 or Fresenius Kabi LOVO can be useful9,10:

Haemonetics Corporation (Cell Saver): https://www.haemonetics.com/en/

TerumoBCT (Teruma Elutro, COBE2991): https://www.terumobct.com/

Fresenius (Fresenius Kabi LOVO): http://lovo.fresenius-kabi.us/

Next, T-cell purification, activation and ex vivo expansion is needed to obtain a T-cell yield9. Activation and growth is achieved using magnetic anti-CD3/anti-CD28 monoclonal antibodies or cell-based artificial antigen-presenting cells (aAPCs). The function of bioreactor is to provide a favorable environment for cell growth and vector transduction. Beads and bioreactors are available from9,10:

Thermofisher (Dynabeads): https://www.thermofisher.com/uk/en/home/life-science/cell-analysis/cell-isolation-and-expansion/cell-expansion/human-t-cell-expansion.html

Miltenyi Biotec (Miltenyi MACS GMP ExpAct Treg Beads): https://www.miltenyibiotec.com/GB-en/products/cell-manufacturing-platform/macs-gmp-portfolio/activation-and-expansion-tools/macs-gmp-expact-treg-kit.html

GE Healthcare Life Sciences (WAVE Bioreactor): https://www.gelifesciences.com/en/gb/shop/cell-culture-and-fermentation/rocking-bioreactors

Wilsonwolf (G-Rex Bioreactor): http://www.wilsonwolf.com/

Creating CAR T-cells requires the usage of viral vectors that transduce the cells and incorporate CAR into the leukocyte DNA9. Oxford Biomedica and Creative Biolabs manufacture viral vectors suitable for CAR T-cell production.

Oxford Biomedica: http://www.oxfordbiomedica.co.uk/

Creative Biolabs: https://www.creative-biolabs.com/car-t/

Bibliography:

1.     National Cancer Institute. (2017). CAR T Cells: Engineering Immune Cells to Treat Cancer. [online]

2.     Pang, Y., Hou, X., Yang, C., Liu, Y. and Jiang, G. (2018). Advances on chimeric antigen receptor-modified T-cell therapy for oncotherapy. Molecular Cancer, [online] 17(1), pp.1-10.

3.     ASCO post. (2018). Updated Analysis of ELIANA Trial Shows Longer-Term Durable Remissions With Tisagenlecleucel in Children, Young Adults With Relapsed/Refractory ALL. [online]

4.     Caffrey, M. (2018). With Approval of CAR T-Cell Therapy Comes the Next Challenge: Payer Coverage. [online] ajmc.com.

5.     Gilead.com. (2018). Kite’s Yescarta™ (Axicabtagene Ciloleucel) Becomes First CAR T Therapy Approved by the FDA for the Treatment of Adult Patients With Relapsed or Refractory Large B-Cell Lymphoma After Two or More Lines of Systemic Therapy | Gilead. [online]

6.     Labiotech.eu. (2018). Novartis' CAR-T Therapy Kymriah Is Not Living Up to Expectations. [online]

7.     Shock Exchange (2018). Gilead: Will Medicare Boost Yescarta Sales?. [online]

8.     Sagonowsky, E. (2018). Novartis could cut its Kymriah price to $160,000 and keep its profit margins: study | FiercePharma. [online]

9.     Levine, B., Miskin, J., Wonnacott, K. and Keir, C. (2017). Global Manufacturing of CAR T Cell Therapy. Molecular Therapy - Methods & Clinical Development, [online] 4, pp.92-101.

10.  Wang, X. and Rivière, I. (2016). Clinical manufacturing of CAR T cells: foundation of a promising therapy. Molecular Therapy - Oncolytics, [online] 3(16015), pp.1-7.

Glioblastoma Multiforme