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Published on: July 22, 2019
Mitigation of chromosome loss in clinical CRISPR-Cas9-engineered T cells
Connor A Tsuchida1,2,3, Nadav Brandes4,3, Raymund Bueno4,3,5
1University of California, Berkeley - University of California, San Francisco Graduate Program in Bioengineering, University of California, Berkeley, Berkeley, CA, USA.
Abstract:
CRISPR-Cas9 genome editing has enabled advanced T cell therapies, but occasional loss of the targeted chromosome remains a safety concern. To investigate whether Cas9-induced chromosome loss is a universal phenomenon and evaluate its clinical significance, we conducted a systematic analysis in primary human T cells. Arrayed and pooled CRISPR screens revealed that chromosome loss was generalizable across the genome and resulted in partial and entire loss of the chromosome, including in pre-clinical chimeric antigen receptor T cells. T cells with chromosome loss persisted for weeks in culture, implying the potential to interfere with clinical use. A modified cell manufacturing process, employed in our first-in-human clinical trial of Cas9-engineered T cells, 1 dramatically reduced chromosome loss while largely preserving genome editing efficacy. Expression of p53 correlated with protection from chromosome loss observed in this protocol, suggesting both a mechanism and strategy for T cell engineering that mitigates this genotoxicity in the clinic.
Insights
CRISPR-Cas9 gene editing can cause chromosome loss in T cells, a safety concern for therapies. A new manufacturing process significantly reduced this loss, enhancing T cell therapy safety.
Area of Science:
- Genomics
- Immunotherapy
- Cellular Engineering
Background:
- CRISPR-Cas9 genome editing advances T cell therapies but carries a risk of chromosome loss, a potential safety issue.
- The clinical significance and universality of Cas9-induced chromosome loss in primary human T cells require thorough investigation.
Approach:
- Conducted systematic analysis using arrayed and pooled CRISPR screens in primary human T cells.
- Evaluated chromosome loss in pre-clinical chimeric antigen receptor T cells.
- Assessed a modified cell manufacturing process in a first-in-human clinical trial for Cas9-engineered T cells.
Key Points:
- Chromosome loss was a generalizable phenomenon across the genome, leading to partial or entire chromosome loss.
- T cells with chromosome loss demonstrated persistence in culture for weeks, posing a clinical concern.
- A modified manufacturing process significantly reduced chromosome loss while maintaining genome editing efficacy.
Conclusions:
- Cas9-induced chromosome loss is a significant genotoxicity concern in T cell engineering.
- The p53 pathway plays a role in mitigating Cas9-induced chromosome loss.
- The modified cell manufacturing process and understanding p53's role offer strategies to improve the safety of CRISPR-Cas9-based T cell therapies.
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