CARAMBA: a first-in-human clinical trial with SLAMF7 CAR-T cells prepared by virus-free Sleeping Beauty gene transfer to treat multiple myeloma

Authors:
Sabrina Prommersberger , Michael Reiser , Julia Beckmann , Sophia Danhof , Maximilian Amberger , Patricia Quade-Lyssy , Hermann Einsele , Michael Hudecek , Halvard Bonig , Zoltán Ivics 
In:
Source: Gene Ther
Publication Date: (2021)
Issue: 28: 560-571
Research Area:
Immunotherapy / Hematology
Gene Expression
Regenerative medicine
Cells used in publication:
T cell, human stim.
Species: human
Tissue Origin: blood
Platform:
4D-Nucleofector™ X-Unit
Experiment

taken from Miskey et al: CD8+ and CD4+ T cells were purified from PBMC and activated by stimulation with anti-CD3/anti-CD28 beads (Thermofisher) in separate cultures. Transfection of transposon and transposase vectors was performed on day 2 with 10 µg transposon MC and 5 µg SB100X transposase MC per 1×106 T cells on a 4DNucleofector according to the manufacturer’s instructions (Lonza). Transfected CD8+ and CD4+ T cells were expanded using the G-Rex culture system (Wilson-Wolf). On day 14, SLAMF7 CAR-expressing T cells were quantified using the EGFRt marker, and the SLAMF7 CAR-T product formulated to comprise CD8+CAR+ and CD4+CAR+ T cells at a 1:1 ratio.

Abstract

Clinical development of chimeric antigen receptor (CAR)-T-cell therapy has been enabled by advances in synthetic biology, genetic engineering, clinical-grade manufacturing, and complex logistics to distribute the drug product to treatment sites. A key ambition of the CARAMBA project is to provide clinical proof-of-concept for virus-free CAR gene transfer using advanced Sleeping Beauty (SB) transposon technology. SB transposition in CAR-T engineering is attractive due to the high rate of stable CAR gene transfer enabled by optimized hyperactive SB100X transposase and transposon combinations, encoded by mRNA and minicircle DNA, respectively, as preferred vector embodiments. This approach bears the potential to facilitate and expedite vector procurement, CAR-T manufacturing and distribution, and the promise to provide a safe, effective, and economically sustainable treatment. As an exemplary and novel target for SB-based CAR-T cells, the CARAMBA consortium has selected the SLAMF7 antigen in multiple myeloma. SLAMF7 CAR-T cells confer potent and consistent anti-myeloma activity in preclinical assays in vitro and in vivo. The CARAMBA clinical trial (Phase-I/IIA; EudraCT: 2019-001264-30) investigates the feasibility, safety, and anti-myeloma efficacy of autologous SLAMF7 CAR-T cells. CARAMBA is the first clinical trial with virus-free CAR-T cells in Europe, and the first clinical trial that uses advanced SB technology worldwide.