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Nanoparticles with switchable shells for virus sensing and inhibition

Periodic Reporting for period 1 - rSAMs-NANO (Nanoparticles with switchable shells for virus sensing and inhibition)

Reporting period: 2018-09-03 to 2020-09-02

The development of antiviral drugs and virus sensors remains a challenge in the field of medical therapies and diagnosis. The current approach for the design of new antiviral drug candidates relies on the synthesis of monovalent drugs that inhibit virus replication at the late stage of infection. The existing antiviral treatment however suffers from low effectiveness due to the emerging drug resistance mutation. In this context, the development of synthetic drugs with multivalent ligand representation is considered an advantageous alternative for the treatment of severe viral infections such as Influenza. Influenza is an acute respiratory disease and can spread rapidly and widely in the winter season. About 20% of children and 5% of adults worldwide develop symptomatic influenza each year. It causes a broad range of illnesses, from symptomless infection to primary viral and secondary bacterial pneumonia. Rapid diagnosis of influenza viruses during an outbreak is critical for disease control. There are currently three types of diagnostic tests for influenza viruses: virus isolation, antigen capture immunoassays, and molecular diagnostic tests. Although effective and sensitive, these methods require trained personnel and a long testing time. Hence, the development of probes and sensors capable of rapid typing and subtyping of the influenza virus is highly desirable. The current design of the new efficient drugs and diagnostics of Influenza includes the synthesis of complex multivalent compounds as dendrimers, fullerenes, polymers, etc. and requires time-consuming multistep organic synthesis. In this project, we applied the rSAM technique for the engineering of multivalent virus inhibitors and the development of the new effective sensors that can provide flexible recognition of the influenza viruses. The rSAMs are pH-switchable versions of thiol-SAMs monolayers that mimic the complex multivalent carbohydrate arrays present on the cellular surfaces. The rSAMs can be constructed using easily accessible small ligands that allow avoiding the above-mentioned synthetic limitations. The main objectives of the rSAM-NANO project were 1) to investigate the use of rSAMs as dynamic nanoparticle shells for multivalent inhibition of pathogen infection and colonization, and 2) to assess such systems as new antibody-free ultrasensitive and robust sensors for rapid in situ virus diagnostics and viral load of the patient.
Multivalency is essential for the adherence of bacteria or virus particles onto the host cell surfaces. Inhibition of these interactions can restrain the spreading of an infection by blocking the receptor at the early stages of infection. Current drug design and diagnostics are exploring the multivalency concept, i.e. binding of biological targets based on multiple weak interactions. In contrast to classical drug design relying on high-affinity inhibitors, the multivalency concept relies on commonly dendritic architectures with peripheries featuring a high density of ligands, for example, saccharides, capable of simltaneously interacting with biointerfacial receptors. In this context, the surface adaptability where ligands can diffuse laterally to optimize receptor binding is important for generating strong multivalent interactions. The topic of this project concerns Reversible Self-assembled Monolayers (rSAMs) and their applications as dynamic nanoparticle shells for multivalent interactions at biointerfaces. rSAMs are pH-switchable versions of thiol-SAMs that allow a reversible and ordered introduction of affinity reagents on sensor surfaces. Contrary to traditional SAMs of alkanethiols on gold, the rSAMs are tunable to the nature of the head group and layer order and stability, while featuring pH responsiveness and the dynamic nature of non-covalently build assemblies.

The objectives of this project were:
1) to investigate the use of rSAMs as dynamic nanoparticle shells for multivalent inhibition of pathogen infection, and
2) to assess such systems as nanoplasmonic sensors for antibody-free ultrasensitive and robust sensors for rapid in situ detection of viruses.

The project’s objectives have been largely fulfilled and advanced the state of the art in the following ways:
1) We prepared and studied a series of rSAMs layers comprising two different bioactive ligands and studied their affinity to influenza virus proteins such as neuraminidase and hemagglutinin
2) We demonstrated that mixed rSAMs featuring ligand-terminated groups form adaptable surfaces that are optimal for multivalent receptor binding
3) The selected rSAMs were transferred on the 3D surfaces such as gold nanoparticles, nanorods, and nanosquares
4) We have demonstrated that the resulted 3D multivalent hybrid materials bind the viral proteins and can be used as for antibody-free ultrasensitive and robust sensors for rapid in situ detection of viruses and virus inhibition.

Overall, the project has produced commercially exploitable results. We have engaged in a proof-of-concept study with leading sensor companies and the ER is currently acting as principal investigator in an industry-academia collaborative project. Finally, the technology developed by the ER has been patented and will be disseminated in a number of publications in preparation.
Scientific impacts. The research in rSAM-Nano has demonstrated a highly flexible and adaptable new technology with a potential impact on the domains of glycobiology, antiviral research, and clinical diagnostics. A generic approach to design virus receptors featuring unprecedented affinities has been key in this endeavor. We have shown that the approach can be used to prepare 3D core-dynamic shell nanoparticles for multivalent virus inhibition as well as 2D dynamic rSAM chips for optical or gravimetric sensing of intact virus or key viral proteins. Given the ease of optimizing these receptors, we expect the technology to particularly interesting in rapid response actions targeting new pandemic threats. Moreover, we expect that our approach will allow the development of rapid, sensitive, and accurate detection of viral particles. These features are highly important for the development of new protocols for the early-stage virus diagnostics and viral load of the patient. Moreover, our approach represents a valuable rational strategy that can be applied to address the challenges in the domain of anti-viral drug design and improve the potential treatment of emerging viruses. We are convinced that the results of this project will pave the way for future innovations benefitting the European biomedical industry.

Economic societal impacts. The innovative technologies improving the virus surveillance programs are of fundamental importance to public health. The global anti-viral market demonstrated rapid growth in recent years and the Covid-19 pandemic will strongly accelerate this trend. The key market challenges remain the high cost of drug development and the need for the techniques providing an accurate evaluation of the antiviral treatment efficiency. In this context, the proposal rSAM-Nano meets well the market demands but success can only be achieved provided that the results are effectively commercialized. This will benefit society as a whole by creating jobs as well as novel medical solutions including tools for diagnosis and anti-viral treatment.