You may have recently heard about cases of acute lymphoid leukemia (ALL) being cured in a few adults and children using a new type of cell therapy [1]. In these studies, scientists took a patient’s own immune cells, altered their genetic material so that these cells would attack the leukemia, then put the modified immune cells back into the patient. The immune cells used in this therapy are T cells that are engineered to have a chimeric antigen receptor (CAR). The final product, called a CAR T cell, is a revolutionary alternative to traditional cancer therapies like radiation and chemotherapy. This technology allows for specific targeting and elimination of cancer cells without harming healthy cells. The premise behind CARs is that the human body already has the potential to fight infection and disease– CAR T cells simply harness this ability and give the system a boost.
Harnessing the Power of T Cells
The human immune system is incredibly robust in its capacity to recognize and eliminate disease, both rapidly and with high specificity in its ability to distinguish between different types of invaders. T cells are a powerful type of white blood cell that contribute to immune defense by sensing and targeting a particular type of danger. Instead of recognizing an entire infected or diseased cell, T cells recognize a small piece of their target, called an antigen. The surface of the T cell has sensors called T cell receptors (TCRs) to recognize the antigen via a lock and key mechanism. The antigen is thought of as the key that comes into contact with the TCR, the lock, flipping a switch to relay signals within the T cell that instruct it to kill the cell that has the antigen.
But seeing an antigen through its TCR is not enough to activate a T cell. One of the most important features of T cells is their ability to discriminate between real danger and a false alarm by requiring two signals for activation. These signals come from an antigen-presenting cell (APC) that engulfs diseased cells and presents their antigens to other immune cells. The first of these signals is the interaction of the T cell’s TCR with its antigen. The second signal is received through the T cell’s CD28 surface protein that recognizes other proteins made by APCs only when they engulf diseased or infected cells, and not when they engulf healthy cells. This mechanism double checks to ensure T cells will not kill healthy cells. Once both signals are engaged, the T cell is activated and ready to kill the cells in the body that have the target antigen on their surface. Given the specificity and function of T cells, they are great candidates for engineering to fight cancer.
Figure 1. (1) The APC shows the target antigen to the T cell on the surface and simultaneously gives a co-stimulatory signal based on the sensing of proteins by CD28. The relay proteins inside the T cell connected to the TCR and CD28 then give signals to activate the T cell. (2) When the T cell sees a cancer cell with the antigen, the TCR binds the antigen and (3) kills the cancer cell.
Genetic Engineering of CARs
For a T cell to be functional, two signals are absolutely necessary: 1) antigen recognition and 2) co-stimulation. In normal T cells, each of these signals requires a separate sensor outside the cell and proteins inside the cell to relay the signal. In engineered T cells, a protein called a single-chain variable fragment (scFv) is engineered to take the place of the TCR. In designing a CAR against cancer, the biggest challenge is to find an antigen that can distinguish a healthy cell from a cancerous cell. Once the appropriate antigen is identified, an scFv can be designed to recognize the antigen. In the case of ALL, researchers engineered an scFv to recognize an antigen called CD19 that is found on most ALL cancer cells [2].
With the scFv designed as the cell-surface antigen sensor, the second task is to address the co-stimulatory signal. When a T cell comes into contact with the cancer cell that has the antigen, this one interaction alone does not allow for killing of the cancer cell. The cancer cell cannot provide a co-stimulatory signal to CD28 from the surface and the technology of CARs looks to bypass the need for an antigen-presenting cell. So to facilitate this process in CAR T cells, the scFv is engineered to transmit both the first (TCR) and second (co-stimulatory) signal whenever it binds to the antigen. This way, both signals are relayed through one carefully engineered sensor instead of through the two distinct surface sensors required of natural T cells. Thus, the CAR T cell can be activated to kill the cancer cell. This new sensor is called a Chimeric Antigen Receptor (CAR), because the components of two receptors are combined into one chimeric receptor.
Figure 2. The CAR T cell has the scFv on the surface as the sensor and is connected to the relay proteins from CD28 and the TCR inside the cell so that the CAR T cell can kill the cancer cell.
Once the CAR is designed, how does it become a part of the T cell? A harmless virus is used to infect the T cell, and it inserts the CAR gene (the segment of DNA that instructs the cell to make the CAR protein) into the T cell’s genetic material. Since the cells constantly make protein using their DNA as a blueprint, incorporating the CAR gene into the cell allows the T cell to have a constantly renewable source of CAR proteins—a more stable solution than providing the protein itself, which could be broken down over time within the cell.
CARs on the road to fighting cancer
To use this technology in the clinic, physicians first draw the patients’ blood and separate out the T cells. Then they use a virus, as described above, to insert the CAR gene into the T cells. After making sure the freshly-made CAR T cells respond to the correct antigen and are safe, doctors inject the cells back into the patient’s bloodstream. Once inside the body, the engineered cells find the cancerous cells and destroy them. For ALL, this is an ideal method because the cancerous cells are white blood cells that are also in the bloodstream [3].
There are quite a few challenges to bringing CARs to the forefront of cancer therapy for other types of cancers. Among the issues is the difficulty in finding antigens that will distinguish the cancer cell from a healthy cell. Cancer cells can evade the immune system by posing as healthy cells, so this makes the task that much more daunting. If a CAR recognized an antigen that was on both healthy and cancer cells, the CAR T cells could kill perfectly healthy cells along with the cancer. In addition, once the CAR T cells are in the body, they need to be able to encounter their target for killing. In ALL, the cancer is made up of blood cells, so delivering the CAR T cells in the bloodstream allows them to find their cancer targets quickly. However, this is not so easy for other types of cancers. The brain, for example, is a tough place for T cells to enter, so CAR T cells against brain cancers may have to be delivered directly to the brain or include an additional protein to lead them there—advances in the CAR technique that may take years to perfect. The initial success of CAR T cells, however, gives a promising outlook for progress in cancer therapy in the years to come.
Vinidhra Mani is a graduate student in the Immunology Program at Harvard Medical School.
References
1. Denise Grady. “In Girl’s Last Hope, Altered Immune Cells Beat Leukemia.” NY Times. December 9, 2012. <http://www.nytimes.com/2012/12/10/health/a-breakthrough-against-leukemia-using-altered-t-cells.html?pagewanted=all&_r=3&.>
2. Davilla, ML, et. al. “How do CARs work?”. Oncoimmunology. December 1, 2012.
3. Mayo Clinic Diseases and Conditions: “Acute Lymphocytic Leukemia.” <http://www.mayoclinic.com/health/acute-lymphocytic-leukemia/DS00558>.
Further Reading
Alberts, B., et. al. “Helper T Cells and Lymphocyte Activation.” U.S. National Library of Medicine. <http://www.ncbi.nlm.nih.gov/books/NBK26827/>.
CAR Clinical Trial: Grupp, S.A., et. al. “Chimeric Antigen Receptor–Modified T Cells for Acute Lymphoid Leukemia”. New England Journal of Medicine. March 25, 2013.
Maher, J. “Immunotherapy of Malignant Disease Using Chimeric Antigen Receptor Engrafted T Cells”. ISRN Oncology. November 14, 2012. <http://www.hindawi.com/isrn/oncology/2012/278093/>.