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Kissling, L.

Publications and source records attributed to Kissling, L..

2 recordsLinked to original sources

BindCraft: one-shot design of functional protein binders

Protein-protein interactions (PPIs) are at the core of all key biological processes. However, the complexity of the structural features that determine PPIs makes their design challenging. We present BindCraft, an open-source and automated pipeline for de novo protein binder design with experimental success rates of 10-100%. BindCraft leverages the weights of AlphaFold21 to generate binders with nanomolar affinity without the need for high-throughput screening or experimental optimization, even in the absence of known binding sites. We successfully designed binders against a diverse set of challenging targets, including cell-surface receptors, common allergens, de novo designed proteins, and multi-domain nucleases, such as CRISPR-Cas9. We showcase the functional and therapeutic potential of designed binders by reducing IgE binding to birch allergen in patient-derived samples, modulating Cas9 gene editing activity, and reducing the cytotoxicity of a foodborne bacterial enterotoxin. Lastly, we utilize cell surface receptor-specific binders to redirect AAV capsids for targeted gene delivery. This work represents a significant advancement towards a "one design-one binder" approach in computational design, with immense potential in therapeutics, diagnostics, and biotechnology.

bioinformatics↗

Safe delivery of AAV vectors to the liver of small weaned pigs by ultrasound guided percutaneous transhepatic portal vein injection

One challenge for liver-directed gene therapy is sufficient vector delivery to the target tissue while minimizing loss of the applied vector dose to other tissues. Infusion via peripheral veins is the least invasive approach; however, it results in systemic diffusion and substantial vector dilution. Here, we describe a safe and minimally invasive method to deliver adeno-associated virus (AAV) vectors to the liver of small weaned pigs by ultrasound-guided percutaneous trans-hepatic portal vein injection. 4-week-old piglets were infused with [~]2.5x1014 vector genomes comprising a dual-rAAV2/9 vector system with a split adenine base editor for in vivo inactivation of PCSK9 to reduce LDL-cholesterol levels. Animals had no signs of discomfort and tolerated the procedure well. However, despite 45% editing of the target site with the applied adenine base editor system in cultivated porcine cells, we only found low amounts of AAV vector genomes and neither detectable transgene-expression nor successful editing in the treated pig livers. We hypothesize that rapid proliferation of pig hepatocytes caused AAV vector dilution, leading to a loss of the vectors from the nucleus, and hence insufficient base editor protein expression for achieving detectable editing rates. Nonetheless, ultrasound-guided percutaneous transhepatic injection to the portal vein is well-tolerated in piglets and has potential for human (neonate) application.

molecular biology↗