Felix Goerdeler, Phd Student at Max Planck Institute of Colloids & Interfaces
Find out more at mpikg.mpg.de.
Protozoan parasites such as Plasmodium, Leishmania or Toxoplasma are responsible for some of the most severe health problems worldwide.
The intracellular forms of these parasites carry a dense cover of carbohydrate chains attached to a lipid moiety called glycosylphosphatidylinositols (GPIs).
Whereas mammalian GPIs usually anchor proteins to the cell surface, the majority of parasitic GPIs is protein-free. Previous work revealed that these GPIs are highly immunogenic and can affect the severity of symptoms in the host.
A well-characterized example are Plasmodium GPIs which act as inflammatory toxins in malaria.
Nanobodies are the smallest antigen-binding fragments from heavy-chain-only antibodies exclusively found in camelids (camel, llama, alpaca).
Their single-domain nature allows straightforward expression in bacterial systems and their small size (approx. 15 kDa) enables them to reach less accessible antigens, e.g. intracellular targets, while maintaining high affinity and stability.
Furthermore, nanobodies can be easily functionalized to give them additional properties, such as multivalency or multispecificity, or to couple them to effector molecules, such as drugs or fluorescence labels. Due to the high structural complexity of glycans, hardly any glycan-targeting nanobodies were described so far.
Here, we show the successful development of glycan-targeting nanobodies against parasitic GPIs. Affinity measurements with synthetic GPIs confirmed that nanobody binding is glycan-dependent. Furthermore, we found that our nanobody recognizes native glycans on the surface of Plasmodium falciparum, Leishmania major/mexicana and Theileria annulata parasites but not Babesia, Besnoitia or Neospora which renders it a useful diagnostic tool.
Next, we sought to functionalize the nanobody for therapeutic applications. Foreign and immunogenic glycan epitopes such as rhamnose or aGal, are present on the food that we consume. They initiate the development of glycan-specific naturally-circulating antibodies. We aim to exploit the presence of such antibodies to amplify an anti-parasitic response using synthetic glycan-coupled nanobodies. The functionalized nanobodies will recruit naturally-circulating antibodies and subsequent anti-parasitic complement response.
Nicolas Bery, Postdoctoral Scientist at Cancer Research Centre of Toulouse (CRCT)
Find out more at crct-inserm.fr.
Tumour-associated KRAS mutations are the most prevalent in the three RAS-family isoforms and involve many different amino-acids.
Therefore, molecules able to interfere with mutant KRAS protein are potentially important for wide-ranging tumour therapy.
We describe here (1) the selection and characterisation of KRAS-specific antibody mimetics (Designed Ankyrin Repeat Proteins, DARPins) and (2) the engineering of these DARPins into RAS degraders based on protein macromolecules fused to specific E3 ligases.
The KRAS-specific DARPin K19 fused to the VHL E3 ligase is compared to a pan-RAS intracellular single domain antibody (iDAb) fused to the UBOX domain of the CHIP E3 ligase.
We demonstrate that while the KRAS-specific DARPin degrader induces specific proteolysis of both mutant and wild type KRAS, it only inhibits proliferation of cancer cells expressing mutant KRAS in vitro and in vivo.
Pan-RAS protein degradation, however, affects proliferation irrespective of the RAS mutation.
These data show that specific KRAS degradation is an important therapeutic strategy to affect tumours expressing any of the range of KRAS mutations.
