CAR T-Cell Therapy: A Healthcare Professional's Guide - Manufacturing
CAR T-Cell Therapy: An Overview
Cancer immunotherapy options are expanding at an unprecedented rate. Many forms of immuno-oncology agents are under investigation or have been recently approved, such as the Immune Checkpoint Inhibitors.
Chimeric antigen receptor (CAR) T-cell therapy is a new, investigational form of gene therapy that may offer a promising option for patients and clinicians who are fighting unresponsive or recurrent leukemias or lymphomas that are ineligible for other standard options, such as:
- Bone marrow transplant.
But exactly what is the manufacturing process for this unique type of gene therapy, and how does the patient receive it?
Learn More: CAR T-Cell Mechanism of Action
CAR T-Cell Therapy at the FDA
Axicabtagene ciloleucel (KTE-C19) from Kite Pharma was the first CAR T-cell therapy submitted to the FDA for approval in March 2017. For the current submission, axicabtagene ciloleucel has been studied in relapsed or chemotherapy-refractory B-cell non-Hodgkin lymphoma (B-NHL) patients who are ineligible for autologous stem cell transplant (ASCT). If approved, Kite plans to commercially launch axicabtagene ciloleucel in 2017.
The manufacturing of CAR T-cell therapy is both astounding and complicated at the same time. A bioengineering feat, the CAR T-cells are constructed from the patient's own T lymphocytes to boost their cancer-killing ability, and then re-infused back into the patient with directions to target and kill the cancer cells.
CAR T-cells are also under study for B-cell acute lymphoblastic leukemia (B-ALL), and chronic lymphocytic leukemia (CLL), as well as in numerous solid tumor trials.
Learn More: KTE-C19 Clinical Trials
How Are CAR T-Cells Manufactured?
CAR T-cell therapy is not yet approved by the FDA and is considered investigational; a patient must enter a clinical trial to receive it. Here's the process:
- Leukapheresis: The patient's T-cells are harvested through apheresis and separated from other blood components such as red blood cell, platelets and monocytes. The amounts of specific subsets of T cells, such as CD4 and CD8 may be optimized; however, the optimal subsets of T-cells to improve efficacy and limit toxic effects are still under study.
- T-cell activation: T-cell activation occurs via primary and costimulatory signals. Endogenous cell-based activation, such as with dendritic cells, may be used, but is not reliable and requires further study. In beads-based activation, antibody-coated beads serve as artificial dendritic cells and can activate the isolated T cells. Biotechnology companies have available these established and accessible activators.
- Transduction: In the lab, the activated T-cells are genetically reprogrammed (DNA encoding) using lentiviral or gamma-retroviral recombinant vectors or a transposon system to yield the CAR construct receptor on the surface of the T-cell.
- Expansion of CAR T-cells: There are several CAR T-cell expansion methods that can be used in the lab to ensure a therapeutic dose of genetically modifed cells can be delivered to the patient. Once reproduced, the cells are frozen to ensure stability prior to delivery to the patient for infusion.
On average, the engineering process takes about 2 weeks. This lengthy timeline can be problematic for patients with rapidly progressive disease.
Patient Delivery: The Preinfusion Conditioning Regimen
Before the bioengineered T-cells are reinfused, the patient undergoes a conditioning chemotherapy regimen to reduced the competing T-cell population (lymphodepletion), lower the tumor burden and help reduce serious complications such as cytokine-release syndrome (CRS).
Agents used in conditioning regimens have varied in studies but have frequently included:
- A combination of fludarabine and cyclophosphamide (FC regimen) The FC regimen seems to be commonly used.
- Other agents include pentostatin and bendamustine.
Researchers note that the best conditioning regimen is still under study, but studies have shown an improved 6-month progression-free survival as well as a longer duration of CAR T-cell survival for patients who receive a preinfusion conditioning regimen. Zhang and colleagues reported that the 6-month progression free survival (PFS) was 94.6% for patients given the lymphodepletion regimen prior to cell infusion, significantly higher than 54.5% in patients without lymphodepletion.
Learn More: Adverse Events Linked With CAR T-Cell Therapy
In the manufacturing facility the CAR structure is synthesized using the patient's own T-cells. The CAR is a fusion protein that contains 3 distinct functional domains. For example, with axicabtagene ciloleucel (KTE-C19):
- The extracellular domain is an antibody fragment that elicits signals to activate the engineered T cells to recognize and bind to target antigens (i.e., CD19) on the surface of the cancer cell. The single chain antibody for KTE-C19 is known as FMC-63.
- The intracelluar components called the essential activation domain (CD3) - signal 1 - and the costimulatory domain - signal 2 - provide signals that allow the CAR T-cells to activate, proliferate and survive. Signal 1 and signal 2 have a synergistic effect, leading to tumor cell apoptosis.
- The CAR-T cells then circulate throughout the body to kill cancer cells, including those that have metastasized.
Many different tumor antigen targets are under research; however, the most common antigen that has been targeted in lymphoma and leukemia trials is known as "cluster of differentiation 19" (CD19).
CD19 is expressed on the surface of most B cells, both healthy and cancerous. However, CD19 is not present on other healthy cells and the CAR T construct does not target these cells. However, healthy B-cells are targeted by the CAR T therapy and leads to B-cell aplasia which is managed by regular immunoglobulin replacement.
Other tumor targets under study include CD20 and CD22.
Quality and Education: Necessary Steps
The manufacturing process for CAR-T cells is similar, even though the design and structure of the tumor specific scFv (single chain fragment variable) may be proprietary. The methods used to develop the fusion protein that makes a CAR T-cell are unique to pharmaceutical companies. These proprietary methods may be one variable that leads to various efficacy and toxicity outcomes from study to study.
The CAR-T cell-manufacturing programs used currently are labor intensive and patient-specific. For wide adoption and commercialization of CAR T-cell therapy in clinical practice, current good manufacturing procedure (cGMP) and quality control are crucial steps and must be implemented. In addition, this complex manufacturing technique requires a highly skilled workforce to maintain the GMP facility.
In the future, the development of off-the-shelf or universal CAR T-cells from allogenic donors could streamline the manufacturing process, shorten the time to therapy for patients, and allow one lot of drug to treat many patients.
Education will be a large component of CAR T-cell therapy if deemed approval by the FDA. In this fragile patient population, serious adverse events such as cytokine release syndrome, neurotoxicity, cerebral edema, and B cell aplasia require immediate medical care and ongoing education to the patient, family and health care providers.
Finished: CAR T-Cell Therapy: A Healthcare Professional's Guide - Manufacturing
- Zhang T, Cao L, Xie J, et al. Efficiency of CD19 chimeric antigen receptor-modified T cells for treatment of B cell malignancies in phase I clinical trials: a meta-analysis. Oncotarget. 2015 Oct 20; 6(32):33961-71. Accessed May 21, 2017 at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4741817/
- Wang X. Riviere I. Clinical manufacturing of CAR T cells: foundation of a promising therapy. Molecular Therapy Oncolytics. 2016;3:16015; doi:10.1038/mto.2016.15; published online 15 June 2016. Accessed May 25, 2017 at http://www.sciencedirect.com/science/article/pii/S2372770516300390
- Maus M, Levine B. Chimeric antigen receptor T-cell therapy for the community oncologist. The Oncologist 2016;21:1-10. Accessed May 23, 2017 at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4861363/
- Wei G, Ding L, Wang J, et al. Advances of CD19-directed chimeric antigen receptor-modified T cells in refractory/relapsed acute lymphoblastic leukemia. Exp Hematol Oncol. 2017; 6: 10. Published online 2017 Apr 14. doi: 10.1186/s40164-017-0070-9 Accessed May 25, 2017 at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5391552/
- Feinberg F, Fillman J, Simoncini J, et al. CAR-T Cells: The next era in immuno-oncology. AJMC.com. Feb. 2017 Accessed May 22, 2017 at http://www.ajmc.com/journals/evidence-based-oncology/2017/february-2017/car-t-cells-the-next-era-in-immuno-oncology-/P-4
- Kite Pharma. CAR Video. Accessed May 25, 2017 at http://kitepharma.com/car-video/
- Neelapu S. An Interim Analysis of the ZUMA-1 Study of KTE-C19 in Refractory, Aggressive Non-Hodgkin Lymphoma. Clinical Advances in Hematology & Oncology. Vol. Volume 15, Issue 2, February 2017. Accessed April 22, 2017 at http://www.hematologyandoncology.net/index.php/archives/february-2017/an-interim-analysis-of-the-zuma-1-study-of-kte-c19-in-refractory-aggressive-non-hodgkin-lymphoma/