Strategic Projects

In addition to the core research undertaken in the four Overarching Projects and five Signature Projects and the Centre-wide research which is detailed in the previous pages, the CBNS provides funding for activities that meet the strategic objective of the CBNS to leave a legacy of research excellence and innovation, together with a highly skilled and professional workforce, ready to lead bio-nano science research and technology into the future.

Strategic Projects are assessed by all Centre CIs and are supported by a dedicated fund. The strategic funding is managed centrally and all collaborating organisations contribute part of their ARC funding each year. When assessing proposals for strategic funding, the CBNS Executive Committee consider whether the projects: • Involve collaboration between at least two CIs, or members of their research groups; • Involve collaboration between at least two nodes; • Introduce skills, expertise or activities not presently available within the

Centre; • Have a duration no longer than two years (although after two years, successful projects can apply for additional funding to continue the project); • Address at least one non-research

KPI (such as media, public or industry engagement, visitors, training etc.); and • Not exceed an annual cost of $100,000. In addition to small amounts of funding supporting student attendance at workshops, three major projects were funded in 2019. They are detailed in the following section.

Open Data Fit

Leader: Professor Pall Thordarson Co-Leaders: Professor Tom Davis, Professor Ben Boyd, Dr Simon Corrie, Professor Edmund Crampin, Professor Maria Kavallaris AM

The Open Data Fit project1 is creating a portfolio of websites that bring together tools and information of relevance to the CBNS as well as the broader international research community. The project focuses on what we call small-scale data, i.e. the small data sets that are the bread-and-butter of research in bio-nano research, ranging from kinetic measurements (kinetics) and small-angle scattering curves (nanoparticle characterisation) to cell viability measurements (IC50 for drug treatments). The goal of the Open Data Fit is, therefore, to shine a light on smallscale dark data and transform the way scientists share small-scale datasets. To this end, we designed the Open Data Fit project to assist researchers with their everyday work while at the same time capturing the raw data for productive sharing. The opendatafit (http:// opendatafit.org) web portal (tools) we are building will achieve this by allowing the end-user scientist to upload their data and then perform data fit (analysis) on these datasets, while at the same time capturing the raw data, the analysis method and the results simultaneously in an open-access database. With CBNS support, we built and successfully launched our pilot website supramolecular.org in 2015. This website provides tools for end-users to determine binding constants from NMR, UV-Vis or fluorescence titration data. Since it was launched, it has had over 40,000 visits with the average session lasting 3:30 minutes, showing a high level of engagement from end-users. Moreover, the website has been cited 120 times in the literature (Scopus). Significantly, Sir Fraser Stoddart, Nobel Prize Winner in Chemistry in 2016, agreed to become the Patron of this project. In 2019 we saw a step-change in how the project is influencing the conversations about data management both nationally and internationally. In March 2019, the program lead was invited to an NSF-sponsored workshop in Orlando, Florida on FAIR Chemical Data Publishing Guideline Workshop. The exclusive audience included representatives from all the major publishing companies (Springer-Nature, Science, Wiley, RSC, ACS) and as well IUPAC, NSF, NIST and the NIH. The Opendatafit project was one of the five exemplary Chemical Data Reuse Case Studies presented at this meeting. Later in the year Opendatafit effectively formed one of the major platforms within a national consortium named the Australian Characterisation Commons at Scale (ACCS). The ACCS was then successful in obtaining a $1.9-Milliondollar platform grant from the Australian Research Data Commons (ARDC). With co-investment from University partners, this is a $5.3M project running over three years with the Opendatafit component, the “National Tools for Scattering and Beyond” receiving about $360K of this funding package to grow and expand the Opendatafit platform the next three years.

1. Hibbert D. B., & Thordarson P., The death of the Job plot, transparency, open science and online tools, uncertainty estimation methods and other developments in supramolecular chemistry data analysis Chemical Communications 2016, 12792-12805.

JTCC: visualising bio-imaging and data in 3D virtual reality (VR)

Leaders: Associate Professor John McGhee, Professor Tom Davis Co-Leaders: Dr Andrew Lilja, Dr John Bailey, Mr Campbell Strong, Dr Rowan Hughes, Dr Shereen Kadir

2019 progress

Key projects within the JTCC Signature Project made significant progress in 2019 with the appointment of lead Unity developer Nick Gunn, allowing an increased focus on real-time data visualisation and user engagement. Development of the core projects Nanoscapes and Vox-Cells remained high priorities, with both gaining international conference attention. Other ongoing projects, including the VR visualisation of in vivo nanoparticles and multi-user data analysis platforms continued as core areas of research with an increased emphasis on user testing and evaluation as science discovery and education tools.

Sub-projects:

1. 3D Vox-Cell (UNSW, UQ, Monash) This project tackles the standard approach to volume surface extraction and rendering, to provide both investigators and the lay viewer with a new look in on volumetric datasets. We developed a novel approach for efficient construction and lighting of mesh surfaces extracted from 3D volumetric data sets. Several refinements were made to allow for real-time visualisation of complex data sets, making the system ideal for data inspection in VR. The innovative approach and lighting algorithms were provisionally patented and caught the attention of industry partners. Formal user evaluation of the system will be completed in 2020. 2. Nanoscapes (UNSW, UQ, Monash) – Key project for 2020 This ambitious project aims to bring more authentic scales and densities of biological entities to real-time cinematic visualisations of cellular landscapes. Built using the PC gaming platform Unity, Nanoscapes employs a data-first approach to populate a computer-generated cell surface with key molecules and processes in order to observe the action of nanoparticles in tumour environments. The application serves as both an interactive educational tool and a provoking artefact promoting deeper speculation about nanoworlds. 3. Multi-user VR visualisation of highresolution cellular dynamics data (UNSW, CCIA) This project visualises multiple, dynamic cellular imaging data sets (captured by light sheet lattice microscopy) in an immersive and collaborative virtual environment. This project continues to progress steadily with a core focus on the of development of networked multiuser VR that facilitates longdistance collaboration and cooperation. This work was presented in 2019 at the flagship conference for VR in industry applications (VRCAI), highlighting features including remote voice chat, personalised avatars and data manipulation functionality. 4. Design-led 3D VR of in vivo nanoparticle dynamics data (UNSW, UQ) This project aims to visualise CT and PET data in VR to identify the precise number and location of nanoparticles following in vivo administration. The project has made good progress towards formal evaluations involving students and staff at UQ. User testing, scheduled for early 2020, will probe the effectiveness and usability of the application as a tool for data analysis and education.

Publications:

1. BJ Bailey, A Lilja, C Strong, K Moline, M Kavallaris, RT Hughes, J McGhee, Multi-User Immersive Virtual Reality Prototype for Collaborative Visualization of Microscopy Image Data, VRCAI ’19, Nov 2019, Brisbane, Australia. 2. RT Hughes, C Strong, J McGhee, Vox-Cells: Voxel-based visualization of volume data for enhanced understanding and exploration in Virtual Reality (VR), ACM SIGGRAPH 2019 Posters, July 28 – Aug 1, Los Angles. 3. AR Lilja, S Kadir, RT Hughes, NJ Gunn, C Strong, BJ Bailey, RG Parton, J McGhee, Nanoscapes: Authentic Scales and Densities in Real-Time 3D Cinematic Visualizations of Cellular Landscapes, SIGGRAPH Asia 2019 Posters, Nov 2019, Brisbane.

1 User testing and evaluation of the Vox-Cell volumetric data visualisation tool 2 Finalise the Nanoscapes interactive application and undertake formal evaluation of its educational and speculative potential 3 Formal user testing to evaluate the effectiveness of immersive VR as a platform to visualise and interact with nanoscience laboratory data compared to traditional screen-based methods

4 Refine and test embodied interfaces that allow remote multi-user immersive 3D VR data interaction

KEY GOALS FOR 2020

PET-CT imaging data sets viewed within the Vox-Cell visualisation system. Image credit: Rowan Hughes, John McGhee, Kris Thurecht, Zach Houston.

The neuro-nano interface – using nanotechnology to provide a window into pain

Leaders: Dr Nik Veldhuis, Professor Kris Thurecht, Associate Professor John McGhee Co-Leaders: Dr Daniel Poole, Dr Paulina Ramírez García, Dr Mikey Whittaker and Associate Professor Matthew Kearnes

The two-year project aims to bridge the gap between neuroscience and nanotechnology and provide new insights and new outreach/education opportunities through visualisation of pain states and associated discussion about the potential for nanotechnology to change drug targeting and drug properties for pain. Challenges for nanotechnology beyond cancer Encapsulation into nanoparticles (NPs) can improve the therapeutic effectiveness of drugs by enhancing stability, tolerability, delivery and retention in diseased tissues. There is particular interest in using NPs to deliver anti-cancer drugs, perpetuated by the prospect of engineering NPs to target tumour cells via the so-called EPR effect – exploiting the leaky vasculature and poor lymphatic drainage of solid tumours to promote NP accumulation and uptake. In addition to the development of cancer therapies, there are many potential applications for drug delivery in other conditions and where leaky vessels and edema may occur, or where cellular environments change (e.g. shifts in pH or reactive oxygen species). For example, inflammation and infection lead to edema or swelling, and acidification of extracellular microenvironments and this can be exploited by uptake pH-responsive nanoparticles for exclusive drug release in an affected area to increase drug efficacy and avoid side-effects. As a part of the Signature Project A material scientist’s guide to the cell, our collaborative team in the CBNS proposed that using nanoparticles for intracellular drug delivery would be an effective strategy for the delivery of poorly distributed analgesic or antiinflammatory drugs, and may improve analgesic outcomes. However, our understanding of nanotechnology and in particular, bio-nano interactions with central or peripheral nervous systems is very limited. We have a dedicated program for delivery of drugs to endosomal compartments within neurons of the spinal cord, and seek to extend these studies to other neurons and also increase awareness of opportunities for nanotechnology in modulating neuronal processes. Harnessing nanotechnology to provide a window into pain transmission and analgesia Chronic pain is a global problem with few effective treatment options, and serious societal and pharmacological challenges due to over-subscription of opioids, leading to widespread addiction and significant increases in opioidrelated deaths. The CBNS has an opportunity to capitalise on the momentum of our current neuro-focussed nanotechnology and the current highly publicised ‘opioid epidemic’, through imaging and visualisation, to provide more significant insights into pain processes and addressing key questions in neurobiology: – How, where and when does pain occur? – How can we harness nanotechnology to improve treatment options? – Can nano-based diagnostics or biosensors be developed to allow us to “visualise pain”

Sub-projects:

1. Visualising nanoparticle effects in neurons (Monash, UQ) Background – There remains a need for visualisation and models to allow scientists, students and the public to explore/conceptualise the use of nanoparticles for “fine-tuning” signalling processes, rather than releasing cytotoxic payloads for treating cancer. We will generate a series of visualisations to show how nanoparticles improve delivery. Altering intracellular drug distribution – We have recently generated a new suite of fluorescently labelled nanoparticles (pH-responsive micelles and crosslinked stars) to generate 3D images of fluorescent nanoparticle uptake into neurons of the spinal cord. We are currently using confocal microscopy on spinal cord slices and if possible, we will also apply a powerful new lightsheet microscopy technique that can image through deep tissue (whole spine). This will provide new visual insights to demonstrate that nanomaterials and drugs can be directed to pain-inducing targets in spinal neurons. Targets include the Neurokinin 1 Receptor, which moves to intracellular endosomes to drive neuropathic pain (green stain, figure 1). Biosensor and Nanomaterial uptake in painful conditions – The development of nanomaterials that ‘light up in locations where pain occurs, will allow us to visualise pain over time. In collaboration with ProfessorKris Thurecht, we will track fluorescent- or radio-labelled hyperbranched nanomaterials and intend to explore alternative labelling approaches (radiolabelling or infrared dye) and distinct stimulus-responsive systems, to image distribution and signals in different painful scenarios: In the periphery (e.g. arthritic knee joint) – pH or ROS-responsive polymers label

Fig 1. 3D imaging of Neurokinin 1 Receptor in spinal neurons.

with infrared dye to indicate the location of inflammatory, acidified tissue or the presence of immune cells. In the spinal cord – increase presence and activity of microglia in the spine as pain develops. There are multiple benefits for generating these data: • Demonstrate nanomaterials as a measure (biosensing) the onset on pain over time, OR assess ability for analgesics to reverse this signal • Assessing if biosensors can give a score that can be overlaid with behavioural pain scores for a more comprehensive assessment of pain preclinical models 2. Visualising pain transmission (Monash, UNSW) To demonstrate how pain signalling occurs in neurons and how nanoparticle may aid analgesia, animated videos will be generated utilising the technology develop by Associate Professor John McGhee, Professor Rob Parton and Dr Angus Johnston for the JTCC project. These will be visually distinct from previous 3DVAL projects, for visualising neuronal excitability (aka pain) and drug release/distribution. Synthetic neurons will be used in the first instance and new electron microscopy images of cultured neurons, in collaboration with Rob Parton, is also willing to image cultured neurons by electron microscopy, should we need to use real neuronal data to drive the visualisation.

Progress and key goals for 2020:

• pH-responsive nanoparticles (disassembling micelles or stable stars) have been synthesised with a

Cy5-label for tracking biodistribution over time. • Confocal imaging in spinal neurons with these materials has been initiated and conditions for imaging have been confirmed utilising immunostaining of the NK1 Receptor. • Establishing inflammatory models to demonstrate retention/accumulation of nanomaterials in tissue regions where significant edema has occurred, has been determined and ethics approval is close to being finalised. • Generation of 3D neuron with the 3D

VAL group will being in May, 2020. • Discussions will follow with Associate

Professor Matthew Kearnes about developing a narrative for the current concerns when using opioids and the potential uptake of nanotechnology as a new strategy for understanding and combatting pain.

Outreach:

N. Veldhuis, “Signalling from the inside out: A new understanding of receptor signalling in neurons”, Keynote presentation at Victorian Teachers Symposium, La Trobe University, Feb 2020

Publications:

Paulina D. Ramírez-García, Jeffri S. Retamal, Priyank Shenoy, Wendy Imlach and Matthew Sykes et al. ‘A pH-responsive nanoparticle targets the neurokinin 1 receptor in endosomes to prevent chronic pain’, Nature Nanotechnology, (2019) 14(12):115059, doi:10.1038/s41565-019-0568-x