Workflow Schematic

Gene editing is being used to improve cell-based therapies, including disruption of checkpoint protein genes to create engineered cells refractory to tumor immunosuppression or knockdown of endogenous TCRs to reduce mispairing of endogenous and engineered TCRs and/or to create allogeneic donor cells. The work published in Mol Ther., 24(9), 1570-1580, 2016 and summarized above augmented their previous virus- based CRISPR-mediated delivery method with the MaxCyte GTx®. Activated T cells were first electroporated with multiple mRNAs encoding Cas9 and AAV helper proteins. Electroporated cells were then transduced with AAV for delivery of CRISPR gRNA targeting Tim3 alone or Tim3 & CD3. The dual gRNA targeting both Tim3 and CD3 lead to >60% knockdown of both proteins as assessed via FACS. See publication for detailed methods.
Summary
- Non-viral engineering enables rapid development of next-generation therapies — such as endogenous T cell receptor knockout or disruption of checkpoint inhibitors — with the benefit of simplified, more cost-effective manufacturing.
- MaxCyte Flow Electroporation® Technology (co)delivers a diversity of payloads including mRNA, sgRNA, RNPs, and plasmid & minicircle DNA providing flexibility for sophisticated, non-viral engineering including:
– transient mRNA expression
– nuclease-mediated gene editing (CRISPR, TALEN, ZFN)
– transposon insertion (Sleeping Beauty, piggyBac) - MaxCyte clinical scalability and regulatory compliance provide for streamlined clinical translation of new therapies.
- The high efficiency and low toxicity of MaxCyte Flow Electroporation provides for strong expression of exogeneous genes such as CARs and/or high gene disruption frequencies that bolster therapeutic efficacy.
- Development and optimization of MaxCyte Flow Electroporation for new cell types and payloads is a rapid, straightforward process.