Helen Lyons
Abstract
A team of researchers at UCSF has developed a rapid way to design patient-specific immune cells that may be used to fight cancer, harmful autoimmune diseases, and infectious illnesses.
A New Role for CRISPR
Scientists working in the lab of Alex Marson developed a new way to use CRISPR/Cas9 to insert long stretches of desired DNA into a specific location in the DNA of T cells. CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats), is a class of DNA sequences that was originally discovered in bacteria, which works together with CAS (CRISPR-associated) proteins to recognize, chop out and remove specific segments of DNA. T cells are a set of specific immune cells that customize the body’s response to specific pathogens.
The new method these scientists designed was published in the July 11, 2018 issue of Nature. The technique they outlined is faster and more precise than previously described methods, allowing introduction of designer DNA into specific locations within the DNA of T cells to create new cell lines in a little over a week. Previously, this process took months to achieve, and often resulted in the insertion of new DNA into random areas of the T cell genome by using viral vectors, potentially harming the recipient T cells. Additionally, it was deemed difficult if not impossible to insert large sequences of DNA into the cells. The new method outlined in the paper allows insertion of longer (greater than 1 kb) DNA sequences into T-cells than was previously thought possible.
The team of researchers led by Theodore Roth, MD/PhD student at UCSF, used a pulse of electricity to make cell membranes briefly permeable, allowing introduction of RNA/protein complexes of CRISPR-Cas9 along with “designer” double stranded DNA into T cells. Roth used previous experience in the lab to guide thousands of trials testing different ratios of DNA, CRISPR-Cas9 complexes, electrical fields, and cell culture techniques until he finally identified a recipe that allowed for reliable successful introduction of DNA into the T cells.
Applications in Cancer, Autoimmune Diseases and Infections
The resulting T cell lines are considered a powerful new tool in the fight against cancer. The DNA inserted into T cells codes for a new receptor, allowing the T cell to more easily attach to cancer cells and then attack them. These researchers demonstrated that they could use their method to create T-cells that were effective in killing melanoma cells both in vitro and in a mouse model.
Scientists also believe this technique can be used to treat autoimmune diseases. The group worked with T cells from 3 children in a family with an autoimmune disorder. An important receptor on the children’s T cells is defective, and without that receptor, a class of immune cells called regulatory T cells (T-regs) cannot develop. Because these siblings’ T-regs cannot function normally, they cannot regulate the other immune cells they would otherwise keep in check. As a result, in this family, the immune system is overactive. The researchers were able to show that inserting normal working copies of the receptor into these children’s T cells in vitro resulted in an increase in normal cellular functionality.
Finally, scientists hope that the modification of T cell receptors will also help fight infection. HIV, for example, relies on the presence of particular T cell receptors to gain entry into the T-cells they infect. In fact, certain individuals born with mutations in those receptors do not become infected with HIV even when exposed to the virus. By editing the receptors on T cells in a patient’s body, scientists hope that HIV from an infected T cell will not be able to spread to uninfected T cells, limiting the progression from HIV infection to AIDS.
Although Marson and Roth are receiving the majority of acclaim for their roles in this groundbreaking work, there are actually 45 authors of this paper, highlighting the importance of teamwork in scientific discovery. This breakthrough has the potential to revolutionize the way so many diseases are treated, and is a testament to the power of collaboration and perseverance in devising a solution to an important problem that scientists had struggled with for years.
References
[1] Roth, Theodore, et. al. (11/07/2018). Reprogramming human T cell function and specificity with non-viral genome targeting. Nature. 405–409. Retrieved: 03/08/18.
[2] Farley, Pete. (11/07/2018). T Cell Engineering Breakthrough Sidesteps Need for Viruses in CRISPR Gene-Editing. Science Daily. https://www.sciencedaily.com/releases/2018/07/180711131204.htm. Retrieved: 03/08/18.
[3] Dean, Michael et. al. (27/09/1996). Genetic Restriction Of HIV-1 Infection and Progression to AIDS by a Deletion Allele of the CKR5 Structural Gene. Science. 1856-1862. Retrieved: 16/03/18
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