Page 31 - SaxoCell Annual Report22/23

CAR- CAReNK-AID
NK

The goal of the CAReNK-AID project is to develop innovative cell-based therapies for the
treatment of autoimmune diseases, such as Systemic Lupus Erythematosus (SLE), type 1 diabetes
and pemphigus vulgaris, for which there are currently no suitable treatments. In this project,
natural killer (NK) cells are genetically engineered with chimeric antigen receptors (CAR) to target
and eliminate autoreactive B cells, which are the primary cause of certain autoimmune diseases,
via their surface molecule CD19.

Project lead: Torsten Tonn

Partner: TUD Dresden University of Technology, University Hospital Dresden, University of
Leipzig

A CAR targeting CD19 (CD19.CAR) was constructed and stably transduced into the NK cell line NK-
92. After confirming the activity of CD19.CAR against CD19+ B cells in vitro, primary NK cells were
transduced with CD19.CAR and the transduction efficiency and cytotoxicity were compared. A
GMP procedure for the production of activated, unmodified primary NK cells on a clinical scale has
been established, validated and forms the basis for the implementation of genetically modified
primary NK cells in patients with severe autoimmune diseases. A study protocol is currently being
developed to evaluate safety and efficacy in patients with various autoimmune diseases in which
depletion of CD19-based B cells may hold promise for therapeutic benefit.

In addition, process optimizations focusing on NK cell expansion as well as non-viral gene transfer
are being carried out to enable standardized and automated manufacturing for later phases of
clinical development. Here, a CD19-expressing feeder cell line was successfully established to
enable selective expansion of CD19.CAR NK cells. Various mutations in the CD19.CAR binding site
of the CD19 molecule were introduced to improve expansion properties. For non-viral CAR
delivery, DNA-based constructs for mRNA production have been developed, including
mutagenesis to exploit the natural cap-1 structure with reduced immunogenicity. Self-amplifying
mRNAs (sa-mRNAs) were added to the range of mRNA candidates, and optimal nanoparticle
candidates for mRNA transfection were identified based on studies in standard cell lines.
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