MaxCyte® Flow Electroporation Technology: A Safe, Reliable and Effective Method for Engineering CAR T Cells
Recombinant T cell expression of Chimeric Antigen Receptors (CARs) has shown extraordinary efficacy in numerous clinical trials as an adoptive cell therapy to treat hematological malignancies. Still, CAR T therapy faces significant challenges, ranging from long lead times and expensive manufacturing to complicated vector-engineering. Also, CAR T cell production routinely employs random integration of viruses or transposons, which carries an inherent risk of genotoxicity and costly, long-term patient follow-up. CAR engineering by transient mRNA transfection could be safer but more cost-prohibitive, requiring several doses per patient. DNA is a promising alternative but can cause sensitive T cells to lose functional capacity or induce apoptosis.
Here we describe Nano-S/MARt (nS/MARt), a novel DNA vector platform for stable CAR expression with minimal disruption of T cell activity. This antibiotic-free, nanovector technology uses scaffold/matrix attachment regions (S/MARs) for DNA vector maintenance and replication, and transfects primary human T cells efficiently and without toxicity. When combined with GMP-compliant MaxCyte Flow Electroporation® and CliniMACS Prodigy® automated cell processing, nS/MARt enabled the production of recombinant T cells with stable CAR expression and enhanced anti-tumor activity in only five days. The result was a shortened manufacturing protocol, producing safer cell therapeutics for thousands of patients from a single batch.
Optimized Nanovectors Provide Prolonged Transgene Expression in Primary CD3+ Cells
nS/MARt Delivery by Electroporation Has Minimal Impact on Human T Cells and Provides Superior Functionality
nS/MARt T Cells Mediate Efficient Tumor Killing in vivo
Development of GMP-Compatible, Large-Scale CAR T Cell Manufacturing with nS/MARt Vectors
- In preclinical experiments, T cells engineered using the MaxCyte ExPERT GTx instrument to deliver nS/MARt vectors were compared to LV-transduced cells. MaxCyte electroporation enabled successful modification of human T cells, with more than 80% of viable cells expressing the transgene.
- Additionally, the CD4:CD8 ratio of nS/MARt electroporated cells was not altered and the T cell proliferation capacity remained intact.
- More interestingly, not only did CAR T cells generated with these technologies retain healthy T cell characteristics but also killed target cells more effectively than LV-transduced T cells.
- This data proves that MaxCyte electroporation is a safe and effective method to genetically modify cells.