The amazing results from CAR T-cell therapies for leukaemia in 2013 (Brentjens et al., 2013; Dudley et al., 2013; Grupp et al., 2013) sparked the explosion of interest in cell therapy that we see today. While the great potential of cell therapy is clear, there are many technological challenges to overcome in manufacture. One of the biggest challenges relates to the need to be able to select large numbers of certain cell types from vast populations. This challenge recently caught the attention of the cytometry industry, establishing therapeutic cell sorting as an R&D priority.
A large community of scientists are now trying to bring T-cell-based immunotherapy to the market. Leading contenders Juno and Kite are racing to develop CD19 CART treatments for acute lymphoblastic leukaemia (ALL) and related disorders. Many more researchers are trying to apply T-cell therapies to other more common types of cancer. Notwithstanding these significant advances, the recent deaths sadly caused by CD19 CART clinical trials remind us again of how potent these cellular immunotherapies are, and of the critical need for better control of the therapeutic cell population.
We think that ongoing safety concerns may soon change the way CD19 CART therapies are commonly produced. So far it has been assumed by many that bulk transduction of PBMCs (peripheral blood mononuclear cells) with the CAR vector, followed by expansion in vitro, was good enough to make a safe and effective therapy. However, the high variability in T-cell subsets within the PBMC fraction, and the highly variable expression of CAR groups, means that more effective sorting and separation of cells is very likely to be necessary.
Beyond the concern for safety, researchers developing cell therapies for common cancers need even greater control of cells to produce viable therapies, because cells of common types of cancer, such as breast cancer, skin cancer and prostate cancer, are much more difficult to target with immunotherapy. These cells share surface markers with normal cells, evolve a microenvironment to evade the immune system, and cannot be killed indiscriminately without serious health consequences: the so-called “on-target off-tumour” effects.
Looking at the efforts of scientists at the cutting edge of cell therapy highlights several themes that all require much greater control of cells, either better control of T-cell subsets or better control of genetic modification, both of which may require a much better therapeutic sorter (see: http://stemcellassays.com/2016/05/crude-versus-defined-car-t-cell-therapy-product/).
One exciting example, recently published in Science, identifies T-cells reactive to clonal markers of a cancer that differ from normal tissue, combining these with genetic modification to switch off the T-cells’ inhibitions in the cancer environment (McGranahan et al., 2016 and https://www.ucl.ac.uk/news/news-articles/0216/040316-tumours-seed-destruction). The result is that the potential therapy is based on a small fraction of the PBMCs which must be separated with high yield and high purity.
Another very different example is a T-cell therapy that is not related to cancer. Regulatory T-cells (Tregs) show great promise to induce tolerance to a transplanted organ (Canavan et al., 2016; Safinia et al., 2015). However, the relevant Tregs are a small fraction of the total PBMCs (1 – 5%) and must be separated with multiple molecular markers (CD4+CD25+CD45RA+). Currently, there is no way to sort with anything near the required throughput, purity and yield.
In future posts, we will share what we have learned about the requirements of therapeutic cell sorting, the approaches that leading researchers are taking to address this problem, and the competing technologies. We will also report on our own experience and progress developing an ultra-high-throughput therapeutic cell sorter to fulfil this urgent clinical need.
Brentjens, R.J., Davila, M.L., Riviere, I., Park, J., Wang, X., Cowell, L.G., Bartido, S., Stefanski, J., Taylor, C., Olszewska, M., et al. (2013). CD19-targeted T cells rapidly induce molecular remissions in adults with chemotherapy-refractory acute lymphoblastic leukemia. Sci. Transl. Med. 5, 177ra38.
Canavan, J.B., Scottà, C., Vossenkämper, A., Goldberg, R., Elder, M.J., Shoval, I., Marks, E., Stolarczyk, E., Lo, J.W., Powell, N., et al. (2016). Developing in vitro expanded CD45RA+ regulatory T cells as an adoptive cell therapy for Crohn’s disease. Gut 65, 584–594.
Dudley, M.E., Kassim, S.H., Carpenter, R.O., Yang, J.C., Phan, G.Q., Hughes, M.S., Sherry, R.M., Feldman, S., Spaner, D., Nathan, D.-A.N., et al. (2013). Effective Treatment Of Chemotherapy-Refractory Diffuse Large B-Cell Lymphoma With Autologous T Cells Genetically-Engineered To Express An Anti-CD19 Chimeric Antigen Receptor. Blood 122, 168–168.
Grupp, S.A., Kalos, M., Barrett, D., Aplenc, R., Porter, D.L., Rheingold, S.R., Teachey, D.T., Chew, A., Hauck, B., Wright, J.F., et al. (2013). Chimeric Antigen Receptor–Modified T Cells for Acute Lymphoid Leukemia. N. Engl. J. Med. 368, 1509–1518.
McGranahan, N., Furness, A.J.S., Rosenthal, R., Ramskov, S., Lyngaa, R., Saini, S.K., Jamal-Hanjani, M., Wilson, G.A., Birkbak, N.J., Hiley, C.T., et al. (2016). Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade. Science 351, 1463–1469.
Safinia, N., Scotta, C., Vaikunthanathan, T., Lechler, R.I., and Lombardi, G. (2015). Regulatory T cells: serious contenders in the promise for immunological tolerance in transplantation. Immunol. Toler. 438.
Samson Rogers leads Cellular Highways, TTP’s initiative in therapeutic cell sorting. He has previously worked on a variety of life science technologies at TTP, co-founded two startups, and published research papers on techniques of measuring biomolecules and cells. Samson has a PhD from Cambridge and an MBA from the ESMT.