Researchers from the University of Zurich have engineered a protein coat shielding for the most commonly used vector in clinical gene therapy, human adenovirus type 5, for improved cancer drug delivery.
Viruses have their own genetic material and can infect human cells in a very specific manner. They will then reproduce as directed by their own genes but using the resources of the host cell. These properties make them interesting “gene shuttles” to fight hereditary diseases or cancer.
There are innumerable different viruses, but the human adenovirus 5, which normally causes the symptoms of a typical cold, has substantial advantages: Its genome can be replaced completely by an artificial one which contains only “useful” genes.
Without any of the viral genes left, the virus can no longer replicate and trigger diseases. In addition, the genome of the adenovirus is very large and does not integrate into human chromosomes.
Until now the use of adenoviruses in tumor therapy has been very limited. They lack the ability to infect cancer cells and therefore cannot inject the genetic blueprints for the therapeutic molecules to fight the disease.
Moreover, adenoviruses are efficiently neutralized by the immune system and very rapidly eliminated by the liver. Researchers led by Andreas Plueckthun, professor at the Department of Biochemistry at the University of Zurich, have now succeeded in rebuilding the viruses so that they effectively recognize and infect tumor cells.
“For this purpose we have created molecules which act as an adapter between the virus and the tumor cell,"
explained Markus Schmid, first author of the study.
[caption id=“attachment_94329” align=“aligncenter” width=“680”] Overview of the knob-adapter complexes.
Knob-binding DARPins are trimerized through SHP and bind the knob in a quasi-covalent manner.
The retargeting module (orange), a target-specific DARPin, allows binding of tumor biomarkers like HER2 or EGFR. Non-targeting (control) DARPins are shown in blue.
Credit: Markus Schmid, et al. CC-BY[/caption]
The adapters, which cling very tightly to the coat of the virus, can - depending on their version - bind to different surface molecules on the tumor cell. The scientists tested adapters for several receptors such as HER2 and EGFR, which are present on various types of cancer cells.
Only viruses which were equipped with these adapters were able to infect the tumor cells.
In a further step, researchers hid the virus under a novel protein coat, which serves as camouflage for the virus and which protects it from the immune system. As a basis for this shield the researchers used an existing antibody that they redesigned.
In an interdisciplinary collaboration between the different research teams, the exact architecture of the complete protective coat was described almost down to the level of atoms.
The shield does not only protect the redesigned virus from the immune cells but also prevents the virus from being eliminated by the liver, which normally quickly removes unmodified adenoviruses from the bloodstream, often making therapeutic applications impossible.
[caption id=“attachment_94328” align=“aligncenter” width=“680”] EM structure of shielded virus and crystal structure of hexon–scFv complex.
(a, b) Comparison of EM structure of naked and shielded HAdV5. Color reflects distance to the core (white: <32 nm, red: 32–38 nm, yellow 41 nm, green 43 nm, cyan 46 nm, blue 48 nm).
Trivalent shield proteins (green–blue) bind all over the capsid, resulting in a dense cover of viral capsid proteins (red–light green).
(c) High-resolution crystal structure of scFv–hexon complex elucidates the atomic interactions. Both heavy (magenta) and light (cyan) chain of the scFv bind to the tower of a hexon monomer (three different shades of blue, one scFv displayed as surface, others as cartoon), formed mainly by HVR2 and HVR7.
The structure also shows few interactions with HVR5. Importantly, all three epitopes in the trimeric hexon were occupied with three scFvs
Credit: Markus Schmid, et al. CC-BY[/caption]
The virus, redesigned using sophisticated protein engineering techniques, works. With its shield and its adapter, these viral gene shuttles efficiently infected tumor cells in laboratory animals.
Using these stealth gene shuttles, the UZH scientists want to develop novel therapies for different types of cancer. The numerous advantages of adenoviruses will likely help to tackle one of the greatest problems of cancer medicine - the development of resistances against drugs.
“With this gene shuttle, we have opened up many avenues to treat aggressive cancers in the future, since we can make the body itself produce a whole cocktail of therapeutics directly in the tumor,"
said biochemist Andreas Plueckthun.
The work was funded by the Schweizerische Nationalfonds Sinergia program, predoctoral fellowships from the Forschungskredit of the University of Zurich, and the Maxi Foundation.
Markus Schmid, Patrick Ernst, Annemarie Honegger, Maarit Suomalainen, Martina Zimmermann, Lukas Braun, Sarah Stauffer, Cristian Thom, Birgit Dreier, Matthias Eibauer, Anja Kipar, Viola Vogel, Urs F. Greber, Ohad Medalia & Andreas Plückthun Adenoviral vector with shield and adapter increases tumor specificity and escapes liver and immune control Nature Communications volume 9, Article number: 450 (2018) doi:10.1038/s41467-017-02707-6
Top Image: the adenovirus (left) camouflages itself from the immune system thanks to its protective coat (right). Credit: University of Zurich