Towards new therapeutic strategies: Zika-driven oncolysis of brain tumor cells

Brain tumours are amongst the most complicated and expensive types of cancer to treat. Brain tumour cells are typically highly aggressive and invasive, and the available treatment options may not be successful and often have side-effects which leave surviving patients with chronic long-term illnesses or disabilities. The occurrence of CNS tumours are rising in many countries, and Latin American countries such as Brazil now have some of the highest rates of occurrence of these types of cancer. In addition, in many Low and Middle Income Countries (LMICs) such as Brazil, there is very high income inequality which is often associated with disparities in access to proper healthcare; brain cancers require sophisticated and long term treatments and access to appropriate, let alone leading-edge treatments can be highly variable. In this project, our goal is to develop a new therapy against brain tumours called oncolytic viral therapy. This type of anti-tumour therapy makes use of viruses which are able to very specifically infect and destroy cancer cells. Although this type of therapy has been successfully developed for other types of cancer, oncolytic viral therapy has not yet been applied to brain cancers. This project builds on a recent finding made at the University of Sao Paulo in Brazil whereby the Zika virus is able to infect and destroy specific types of brain cancer cell. Zika virus is best known as an infectious mosquito-borne virus that can cause developmental defects in the brains of babies born to women with Zika infection. Brazil has been at the centre of this public health crisis, and much has been learned about how the Zika virus does this over the last few years. The recent discovery of the potential of Zika as a therapeutic agent against brain cancers now raises important questions that we will address in this project. In common with other pathogenic viruses, Zika invades human host cells and then uses the host cell machinery to replicate and produce more virus particles. How this happens in brain tumour cells is entirely unknown. This project aims to understand how proteins produced by the Zika virus interact and control proteins in the host human cells. Preliminary studies have indicated that certain types of brain cancer cell are susceptible to the Zika virus, and others less so. We aim to discover why this is so, and what the specific signatures of susceptible brain tumour cells are. In terms of a future oncolytic viral therapy, answering these questions will allow us to (1) design therapeutics that mimic Zika and (2) understand which brain tumours are best suited to the new therapy, thereby targeting the therapy to those patients who can benefit most. If successful, our project will contribute towards the development of a new therapeutic option that can be used to treat patients with one of the most aggressive forms of cancer. By design, our project will be a collaboration between the University of Sao Paulo, where this discovery was initially made and the University of Southampton. Specific "proteomic" techniques which allow the highly sensitive measurement of thousands of proteins in human cells are used at the University of Southampton, and through this collaboration this technique will be applied to develop Zika as an oncolytic therapy. In addition, cross-training of early career researchers between the partner laboratories will allow the exchange of expertise, and in particular the training of Brazilian researchers in proteomic techniques, an important and very powerful approach with the potential to impact many other biomedical research areas.

Recent work has shown that Zika virus (ZIKV) is capable of specifically infecting and destroying stem-like cancer cells from aggressive CNS embryonal tumors. Tumor cells closely resembling neural stem cells at the molecular level, with activated Wnt signaling, were more susceptible to the oncolytic effects of ZIKV and activated Wnt signaling increased ZIKV-induced tumor cell death. How Wnt signaling promotes susceptibility of CNS tumour cells to ZIKV-induced oncolysis is unknown as are the ZIKV-host cell protein-protein interactions that mediate infection. This project will use unique ATRT and medulloblastoma cell-lines that are susceptible to oncolysis by ZIKV and analyse them using a multi-omics approach. Interaction and expression proteomic techniques as well as RNA-Seq will be used to identify ZIKV-host cell protein-protein interactions and to identify protein networks implicated in ZIKV susceptibility, and possible mechanisms by which activated Wnt signaling enhances this process. This pump priming project will focus on identifying protein-protein interactions for two of the non-structural ZIKV proteins (the NS-3 helicase and NS-5 polymerase) and will also provide proof of principle towards a future larger and comprehensive study of the ZIKV protein interactome. The function of selected protein-protein interactions will be analyzed using siRNA and co-immunolocalization to identify host proteins and mechanisms required for oncolysis. This study will provide key information that will advance the goal of using ZIKV as oncolytic therapy. First, a detailed molecular profile of ZIKV infected CNS tumor cells will identify pathways and processes that promote ZIKV infection, providing a means to assess which types of CNS tumour cells could be targeted using ZIKV. Second, the identification of specific protein-protein interactions may identify molecular mechanisms that could be targeted in therapeutic development.

An important area of scientific impact will be in the area of fundamental CNS tumour cell biology. We will use cutting-edge molecular tools to analyse CNS tumour cell lines including a new human medulloblastoma cell line established in Dr Keith Okamoto's laboratory at the University of Sao Paulo. Multi-omic analyses of this unique cell-line (compared to other cancers, there are relatively few medulloblastoma cell-lines available) will provide an important dataset, that is expected to help explain the increased proliferation an stem cell properties of this new cell-line. In addition, researchers working to develop oncolytic viral therapy for CNS tumours or indeed for other cancers will be impacted by our work. For example, this project could discover a new mechanism or pathway that can then be picked up by another research group (academic or commercial) to be developed as a therapy.

By identifying the pathways and processes required for Zika virus infection of CNS tumour cells we will also impact those researchers studying the pathogenic qualities of Zika. Zika infection is an ongoing issue, and development of vaccines or other anti-viral therapies will be enhanced by understanding the process of Zika infection. Researchers studying basic Zika biology, the process of Zika infection and the consequences of infection such as microcephaly will be impacted by this study. In particular, we will contribute to the expertise and momentum of Zika research programmes in Brazil, helping those programmes to grow and providing expertise and access to proteomic approaches.

Currently available treatment options for CNS tumours typically include surgery followed by radiation and chemotherapy. But even if successfully treated, these types of tumours often leave the patients with cognitive and motor deficits for the rest of their lives. New treatment options that do not have such side effects or that are more effective are needed. This project, to acquire an initial understanding of Zika virus and how it might be harnessed in the future as an oncolytic viral therapy will provide a potential new therapy enabling better outcomes for patients with CNS tumours.

Proteomics has a high entry barrier through the many and varied technical choices and the high initial and maintenance costs of cutting-edge mass-spectrometry instrumentation. In addition, conducting proteomics experiments invariably requires optimization and is reliant on a range of specialist expertise not often available in typical molecular/biological research environments. An important impact of this project will be to provide access to the cutting-edge instrumentation available through the Centre for Proteomic Research at the University of Southampton and the associated expertise through the Centre's personnel to the team at the University of Sao Paulo.

The project personnel will gain in several ways from participation in this project. The postdoctoral fellow (PDRA) will work on a hugely exciting project with potential to make profound and lasting scientific contributions. The PDRA will work both at the University of Southampton and the University of Sao Paulo, and will benefit from the highly complementary research groups at each institution. In addition, the PDRA will benefit from experiencing scientific research in two very different research environments, helping him or her in their future career. This project will also benefit the PhD students who will participate in technical training exchanges. We know from exchange visits that have already taken place between our laboratories that these can have important and lasting impacts on the students' PhD projects and their future career goals.