Building a next-generation platform for DNA-based antibody therapeutics

Antibody gene transfer (AGT) aims to deliver the monoclonal antibody (mAb) genes rather than the protein to patients, enabling prolonged in vivo expression at low cost. Using a conventional plasmid DNA and ubiquitous CAG promoter (pCAG), our group previously achieved pre-clinical proof of concept (PoC) for intramuscular AGT in mice and sheep by means of electroporation. Our current research focuses on improving various aspects of the platform. First, we aim to compare product properties of in vitro and in vivo produced mAbs, an uncharted area. Second, we aim to generate a more potent and safer platform through plasmid backbone, promoter and sequence engineering. Finally, we seek to expand platform application beyond mAbs and therefore focus on nanobodies, whose small size and modular nature are a perfect fit for gene transfer.

To address the characterization, we produced mAbs in vitro and in vivo, using lipofection and intramuscular AGT, respectively, with the pCAG. The preliminary results show an N-glycan heterogeneity specific to the expression source. Ongoing research is focused on the significance of these findings. To address the safety and potency of the platform, we expressed mAbs in vivo using a minimal plasmid, containing few bacterial sequences and therefore exhibiting an improved safety profile. Resulting peak plasma mAb levels were similar to those obtained with the pCAG (6.5 µg/ml). A 50% improvement in mAb expression came from the use of a muscle-specific promoter, driving peak concentrations above 11 µg/mL in mice. Engineering the mAb sequence is part of an ongoing investigation. To expand the platform, we used pCAG to design a multivalent antitumoral nanobody construct. Upon intramuscular gene transfer in an athymic nude tumor model, this construct led to peak levels of 270 ng/mL, which remained above 200 ng/mL during the 10 weeks of follow-up. Furthermore, the DNA-based nanobody demonstrated potent anti-tumor response, which improved efficacy over intravenous dosing of the protein counterpart.
In conclusion, we demonstrate progress in building a next-generation platform for DNA-based antibody therapeutics. First, in vitro and in vivo produced mAbs demonstrate distinct glycosylation features. Second, we show that plasmid backbone and cassette engineering can improve both safety and potency of the platform. Finally, we demonstrate PoC for DNA-based nanobodies in oncology and highlight the benefit over conventional protein-based delivery.

Authors

Giles Vermeire(1), Elien De Smidt(2), Nick Geukens(2), Paul Declerck(1), Kevin Hollevoet(1)

Organisations

"Laboratory for Therapeutic and Diagnostic Antibodies, KU Leuven - University of Leuven, 3000 Leuven, Belgium (1), PharmAbs, the KU Leuven Antibody Center - University of Leuven, 3000 Leuven, Belgium (2) "

Presenting author

Giles Vermeire, PhD Researcher, KU Leuven
giles.vermeire@kuleuven.be
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