Universal Stem Cells: Making the Unsafe Safe

In their recent letter, González et al. discuss the safety of using universal stem cells in regenerative medicine (González et al., 2020González B.J. Creusot R.J. Sykes M. Egli D. How Safe Are Universal Pluripotent Stem Cells?.Cell Stem Cell. 2020; 26: 307-308Abstract Full Text Full Text PDF PubMed Scopus (4) Google Scholar). The term “universal” conveys that the cells have been genetically modified to escape immune rejection when transplanted into allogeneic recipients. As a group that has worked to develop universal cells (Harding et al., 2019Harding J. Vintersten-Nagy K. Shutova M. Yang H. Tang J.K. Massumi M. Izaidfar M. Izadifar Z. Zhang P. Li C. et al.Induction of long-term allogeneic cell acceptance and formation of immune privileged tissue in immunocompetent hosts.bioRxiv. 2019; https://doi.org/10.1101/716571Crossref Scopus (0) Google Scholar), we have from the beginning held cell therapy safety as paramount. We are grateful that the recent commentary brought these issues to the forefront and want to add to their important points. One starting point for discussing the potential risks of universal cells is to consider the alternatives that could be used to prevent rejection in upcoming cell-based therapies. For instance, while immune suppressive drugs are invaluable tools, they do not always work and increase the risk of life-threatening infections and cancer. Alternatively, the induction of immune tolerance through mixed chimerism may be efficacious for solid organ transplants (Zuber and Sykes, 2017Zuber J. Sykes M. Mechanisms of Mixed Chimerism-Based Transplant Tolerance.Trends Immunol. 2017; 38: 829-843Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar), but hematopoietic cell transplantation can also be a life-threatening procedure. This makes it unrealistic for many cell therapy candidates, especially chronic or non-life-threatening conditions such as blindness, diabetes, arthritis, burns, and spinal cord injury, among many others. A primary goal of a universal cell is to escape immune rejection with a risk profile that is lower than the status quo. A benefit of this vision is the potential for “off-the-shelf” cell products that could treat patients with immediate needs. This will not be possible with autologous induced Pluripotent Stem Cells (iPSCs) even if the costs are dramatically lowered, since they take time to generate and will each have to be independently tested and equipped with safety systems (Merkle et al., 2017Merkle F.T. Ghosh S. Kamitaki N. Mitchell J. Avior Y. Mello C. Kashin S. Mekhoubad S. Ilic D. Charlton M. et al.Human pluripotent stem cells recurrently acquire and expand dominant negative P53 mutations.Nature. 2017; 545: 229-233Crossref PubMed Scopus (214) Google Scholar). They may also be immunogenic (Zhao et al., 2015Zhao T. Zhang Z.N. Westenskow P.D. Todorova D. Hu Z. Lin T. Rong Z. Kim J. He J. Wang M. et al.Humanized Mice Reveal Differential Immunogenicity of Cells Derived from Autologous Induced Pluripotent Stem Cells.Cell Stem Cell. 2015; 17: 353-359Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar). These same safety requirements will apply to large banks of HLA-diverse iPSCs, which themselves cannot provide complete coverage to diverse populations and do not solve the hazards of minor antigen mismatch. New solutions bring new challenges. The immune system is vital for clearing potentially cancerous and infected cells, and universal cells could be intrinsically dangerous. We should eliminate the possibility of making a tumorigenic or transmissible cell line, such as that found in Tasmanian Devils and capable of spreading through biting (Hamede et al., 2013Hamede R.K. McCallum H. Jones M. Biting injuries and transmission of Tasmanian devil facial tumour disease.J. Anim. Ecol. 2013; 82: 182-190Crossref PubMed Scopus (86) Google Scholar). As we articulated in our recent review (Lanza et al., 2019Lanza R. Russell D.W. Nagy A. Engineering universal cells that evade immune detection.Nat. Rev. Immunol. 2019; 19: 723-733Crossref PubMed Scopus (37) Google Scholar) and primary work (Harding et al., 2019Harding J. Vintersten-Nagy K. Shutova M. Yang H. Tang J.K. Massumi M. Izaidfar M. Izadifar Z. Zhang P. Li C. et al.Induction of long-term allogeneic cell acceptance and formation of immune privileged tissue in immunocompetent hosts.bioRxiv. 2019; https://doi.org/10.1101/716571Crossref Scopus (0) Google Scholar), it is essential that any universal cell also contain rationally designed kill-switch systems (Liang et al., 2018Liang Q. Monetti C. Shutova M.V. Neely E.J. Hacibekiroglu S. Yang H. Kim C. Zhang P. Li C. Nagy K. et al.Linking a cell-division gene and a suicide gene to define and improve cell therapy safety.Nature. 2018; 563: 701-704Crossref PubMed Scopus (44) Google Scholar) that provide extremely tight control over cell survival and/or proliferation. It is true that genetically integrated kill-switches can undergo loss-of-function mutations and silencing (Iwasawa et al., 2019Iwasawa C. Tamura R. Sugiura Y. Suzuki S. Kuzumaki N. Narita M. Suematsu M. Nakamura M. Yoshida K. Toda M. et al.Increased Cytotoxicity of Herpes Simplex Virus Thymidine Kinase Expression in Human Induced Pluripotent Stem Cells.Int. J. Mol. Sci. 2019; 20: 810Crossref Scopus (7) Google Scholar), but these kinds of problems have solutions. Our recent publication (Liang et al., 2018Liang Q. Monetti C. Shutova M.V. Neely E.J. Hacibekiroglu S. Yang H. Kim C. Zhang P. Li C. Nagy K. et al.Linking a cell-division gene and a suicide gene to define and improve cell therapy safety.Nature. 2018; 563: 701-704Crossref PubMed Scopus (44) Google Scholar) describes one of them and provides an acceptable cell safety level for most therapeutic uses. This publication was, unfortunately, incorrectly cited by González et al. as addressing the problem of immune escape. In fact, this work addresses several key challenges raised in their commentary and provides a new paradigm for cell safety. Our solution involves the transcriptional linkage of an inducible kill-switch to a cell division essential gene (CDEL), which dramatically lowers the probability of mutational escape or silencing. Our work also shows that the safety level jumps by orders of magnitude as the number of CDEL kill-switch integrations is increased (see Figure 2 of Liang et al., 2018Liang Q. Monetti C. Shutova M.V. Neely E.J. Hacibekiroglu S. Yang H. Kim C. Zhang P. Li C. Nagy K. et al.Linking a cell-division gene and a suicide gene to define and improve cell therapy safety.Nature. 2018; 563: 701-704Crossref PubMed Scopus (44) Google Scholar). In principle, multiple CDELs as well as other classes of essential genes, such as those needed for protein synthesis, can be targeted. This could be done in the same cell line and with multiple kill-switches that work by different mechanisms, such as the combination of Thymidine Kinase and iCasp9. We even demonstrated the feasibility of these kinds of targeted genomic edits by generating transgenic mice where every cell had homozygous insertion of a kill-switch into a prototypical CDEL. Furthermore, our work provides a mathematical framework for calculating the probability that a kill-switch “escapee” could arise, which is based on several key parameters including the number of cells needed and whether the kill-switch is integrated at both CDEL alleles. Importantly, this framework provides a tangible way to quantify risk so that patients and clinicians can evaluate the risk versus reward of a specific cell therapy intervention. A generalizable point from our work is that, like any challenge, designing safe, universal cells is not an intractable problem, but one that will have rational and hard-won solutions. They will come with the understanding that all interventions have non-zero risk, and like any novel therapy, kill-switch-containing universal cells will require incremental safety and efficacy testing before widespread use. Ultimately, this way forward could allow us to leverage the benefits of off-the-shelf therapeutic cells while understanding and controlling their risk profile. J.H. and K.N. are partially supported by a grant from Medicine by Design (University of Toronto). The cited work from A.N.’s laboratory was funded by a CIHR Foundation Grant, Canadian Research Chair and grants from Fighting Blindness Canada . J.H. is employed part time, and K.V.N. full time, by panCELLa Inc. A.N. is the co-founder of the company, which has an exclusive license for the Safe Cell (Liang et al., 2018Liang Q. Monetti C. Shutova M.V. Neely E.J. Hacibekiroglu S. Yang H. Kim C. Zhang P. Li C. Nagy K. et al.Linking a cell-division gene and a suicide gene to define and improve cell therapy safety.Nature. 2018; 563: 701-704Crossref PubMed Scopus (44) Google Scholar) and induced allogeneic cell acceptance (Harding et al., 2019Harding J. Vintersten-Nagy K. Shutova M. Yang H. Tang J.K. Massumi M. Izaidfar M. Izadifar Z. Zhang P. Li C. et al.Induction of long-term allogeneic cell acceptance and formation of immune privileged tissue in immunocompetent hosts.bioRxiv. 2019; https://doi.org/10.1101/716571Crossref Scopus (0) Google Scholar). How Safe Are Universal Pluripotent Stem Cells?González et al.Cell Stem CellMarch 05, 2020In BriefMajor breakthroughs in stem cell differentiation to therapeutically relevant cell types have led us to the doorstep of novel cell therapies that can replace disease management with a cure. Degenerative diseases including diabetes, heart diseases, eye diseases, and neurological disorders could benefit from a renewable source of tissues made from stem cells. Indeed, the first stem cell-based clinical trials involving a small number of patients are now underway. A key obstacle to the wide adoption of cell-based therapies is the immune system, which forms a transplantation barrier and rejects the cells from another person. Full-Text PDF Open Archive