WEBINAR –The Next Generation of CAR-T Cells: New Techniques in Gene Editing and Rapid Assessment with Real Time Live Cell Analysis

Chimeric antigen receptor (CAR) T cell therapy has yielded promising results in recent clinical trials. In this webinar, experts in CAR-T cell development discuss new techniques to further improve functionality.

Topics covered in this webinar:

  • Developing allogeneic anti-CD19 CAR T cells by utilizing an engineered homing endonuclease (ARCUS nuclease) specific for the T-cell receptor alpha constant (TRAC) locus
  • Reducing off-target events and enhancing gene-editing efficiency and production of CAR T cells.
  • Rapid assessment of CAR constructs targeting cleaved MUC1
  • Analysis of CAR-T cells at different time points to simulate treating early stage versus late stage, aggressive cancers
  • Generating in vitro data on the xCELLigence Real Time Cell Analysis Platform to predict the effect of the CAR-T cells in vivo.

Watch Webinar on Demand

Featured Speakers:
Mark C. Johnson, Ph.D.
Team Leader, Cell Therapy Discovery
Precision Biosciences Optimization of Nucleases for Gene-editing in Chimeric Antigen Receptor (CAR) T Cell Manufacturing
Cynthia Bamdad, Ph.D
Chief Executive Officer
Minerva BiotechnologiesRapid Assessment of CAR-T Cells Targeting Cleaved MUC1

Presentation 1:

Optimization of Nucleases for Gene-editing in Chimeric Antigen Receptor (CAR) T Cell Manufacturing
Chimeric antigen receptor (CAR) T cell therapy has yielded promising results in recent clinical trials, particularly for treating CD19+ hematological malignancies. To further improve CAR T functionality, next generation cells will be precisely gene-edited utilizing nucleases that produce targeted double-strand breaks in the host genome. For example, nucleases may be used to enable the insertion of a CAR gene into a targeted site by the process of homology directed repair, or knockout genes such as PD-1 to improve efficacy. Gene-editing specificity is paramount, as unintended DNA damage can lead to deficiencies in CAR T function and reduce the efficiency of cell production. To develop allogeneic anti-CD19 CAR T cells, we utilize an engineered homing endonuclease (ARCUS nuclease) specific for the T-cell receptor alpha constant (TRAC) locus. Although initial nucleases promoted efficient gene-editing, a series of optimization studies increased nuclease specificity for the TRAC locus while reducing off-target events. Of the nucleases screened, five candidates were selected based on analysis of off-targeting events for comparative studies of in vitro functionality and CAR T cell manufacturing efficiency. Using sensitive, real-time measurement of cytolysis with the ACEA Biosystems xCELLigence instrument, we observed increased cytolysis of target cells using CAR T cells made with two of our optimized nucleases compared to the parental nuclease. Furthermore, in vitro functionality correlated with increased TRAC-editing and CAR gene insertion during cell manufacturing. Collectively, these data show that our ARCUS nucleases can be optimized to reduce off-targeting events, thereby enhancing gene-editing efficiency and production of CAR T cells.

Presentation 2:

Rapid Assessment of CAR-T Cells Targeting Cleaved MUC1
Minerva Biotechnologies is developing a pipeline of CAR-T therapeutics for solid tumor cancers. Using the xCELLigence RTCA-MP instrument that enables simultaneous testing of 576 different conditions, we were able to rapidly assess 60 different CAR constructs and choose a clinical lead. Importantly, we paused experiments to add CAR-T cells at different time points to simulate treating early stage versus late stage, aggressive cancers. We also paused experiments to add cleavage enzymes that are overexpressed in cancers to more accurately mimic the tumor microenvironment and to see in real-time their effect on tumor growth and susceptibility to CAR-T mediated cell killing. Various CAR-T cell “training” protocols were tested, which enabled the development of a novel stimulation step that increases specific cell killing but does not increase non-specific cell killing. xCELLigence was faster and more accurate than FACS. xCELLigence in vitro data perfectly predicted the effect of the CAR-T cells in vivo.

Watch Webinar on Demand

Meet the Speakers:

Mark C. Johnson, Ph.D., is a team leader in Cell Therapy Discovery at Precision BioSciences, a biotech located in Durham, North Carolina that solves problems in cancer, genetic disease, and agriculture through gene editing. Mark’s team develops allogeneic CAR-T cells using Precision’s proprietary ARCUS technology, and their current goal is to improve the efficacy, persistence, and specificity of CAR T cell therapies by applying CAR T cells carrying novel genetic edits to a variety of in vitro and in vivo tumor models. Prior to joining Precision BioSciences, Mark was a postdoctoral fellow in the Immunology Department at the University of Washington. He completed his graduate studies in microbiology and immunology at the University of North Carolina, Chapel Hill.Dr. Cynthia Bamdad is CEO of Minerva Biotechnologies. Dr. Bamdad holds a BS in Physics and a Ph.D. in Biophysics from Harvard University. While a PhD student, Bamdad invented the first electronic DNA chip, which formed the cornerstone of a startup (now GenMark) that sold within two years to Motorola for $300MM. Bamdad’s invention is now an FDA-approved medical device. Dr. Bamdad was on the Board of Directors of Pharmacyclics and was critical to the turnaround strategy that brought it from $15MM valuation to $21B sellout to AbbVie. At Minerva, Dr. Bamdad discovered that early stem cells grow by the same mechanism as 75% of all cancers; she discovered how stem cells limit self-replication and how cancers override this natural shut-off switch. Minerva has developed a CAR T for solid tumors that is scheduled for human trials in Q4 2018. Bamdad is inventor on over 150 patents and pending applications.