Bioinnovation and drug discovery at ANSTO’s Australian Synchrotron

Introduction

As one of Australia’s major research facilities, the Australian Nuclear Science & Technology Organisation’s (ANSTO) Australian Synchrotron (AS) is a national institute that drives scientific development for both academic and industrial research applications (Fig. 1, 2). It supports a broad range of studies, from physics, chemistry, materials, medicine, biology, biochemistry and more, providing Australia, New Zealand, Singapore and the surrounding region with access to world-class instrumentation and cutting-edge analytical techniques. Synchrotrons have positioned themselves as invaluable tools for drug discovery and biological innovation. By using the high-intensity radiation produced by a synchrotron source, the AS enables the study of the atomic and molecular composition of any substance with high precision. The facility supports a number of techniques or ‘beamlines’, including scattering, crystallography, microscopy and imaging beamlines, able to provide significant data regarding the molecular and structural dynamics of complex biological systems (Fig. 3). The following sections highlight how these powerful tools are accelerating breakthroughs in drug discovery and bioinnovation.

Fig. 1.

Top view of the Australian Synchrotron, showcasing the oval-shaped synchrotron ring, the onsite guest house (top LH corner), and the solar panels on the roof, highlighting the integration of national research infrastructure with sustainable energy solutions.

Fig. 2.

Floorplan of the Australian Synchrotron, showcasing all 18 beamlines, including 15 operational and 3 under construction at the time of writing (ADS1, ADS2, Nanoprobe). This highlights our broad infrastructure supporting advanced research in a wide range of applications including bioinnovation and drug discovery.

Fig. 3.

Schematic representation of the Australian Synchrotron’s integrated capabilities for advancing bioinnovation and drug discovery. It highlights our key beamline capabilities, including crystallography, scattering, imaging and microscopy, as well as user laboratories and data handling systems, supporting cutting-edge research towards therapeutic development. Created in BioRender.

Crystallography: unlocking the 3-D structure of drug targets

X-Ray crystallography has long been one of the most reliable and widely used methods for determining the three-dimensional (3-D) structure of biomolecules, and X-ray crystallography beamlines at the Australian Synchrotron (AS) are playing a central role in structure-based drug discovery and design.¹ The ability to determine the precise atomic arrangement or coordinates of molecules allows scientists to visualise drug targets in unparalleled detail, providing critical information for the design of small molecules that can bind to specific sites on these targets. Crystallography therefore supports a wide range of therapeutic categories, such as infectious,^2–4 autoimmune⁵ and neurodegenerative disease.⁶ For example, synchrotron researchers have elucidated the structures of bacterial enzymes that are involved in antibiotic resistance, allowing the design of new antibiotics that can evade resistance mechanisms.^7,8 In neuroscience research, crystallography has been instrumental in uncovering the structural characteristics of amyloid proteins involved in Alzheimer’s disease, providing essential information for drug design.^9–11 Research into the SARS-CoV-2 virus and others has also used synchrotron crystallography to examine aspects of the virus, from vaccine and possible drug targets to the immune response of infected individuals.^12–14

The crystallographic beamlines at the AS include the Macromolecular (MX1)¹⁵ and Microfocus (MX2)¹⁶ beamlines, and the High Performance Macromolecular Crystallographic (MX3) beamline (Fig. 2, 3). With its microfocus and high flux beam, MX3 is particularly well suited for studying microcrystals, which are often weakly diffracting. These challenging proteins and protein-complexes are often highly flexible representing the native biological state. MX3 as a new state-of-the-art beamline at the facility, will enable significant advancements in techniques such as serial crystallography, time-resolved crystallography and tray screening. These transformative technologies will enable researchers to capture biomolecular interactions and conformational changes in real time, providing a level of detail that was previously unattainable. Such innovations are crucial, allowing the timely unravelling the complexities of drug targets, accelerating the development of targeted therapies and improving the efficacy of new treatments.

The AS is also supporting the development of a Crystallographic Fragment Screening (CFS) pipeline. CFS using synchrotrons is a very effective approach for identifying potential drug candidates.¹⁷ Traditionally, it has been a slow and labour-intensive process, even when using a synchrotron, and primary screening has remained outside of the repertoire of most laboratories. Attempts are now underway in collaboration with key Australian researchers, to take advantage of advances in technologies to establish a standard CFS pipeline for the design of high-affinity molecules for protein interactions. Together with the AS MX suite, this will significantly enhance Australia’s capability for structure-based drug design, enabling the design of new drugs and therapies more specific, effective and efficient.

Scattering methods: studying biological interactions in solution

One of the major techniques at the AS is Small Angle X-ray Scattering (SAXS). SAXS allows researchers to study the shape and structure of macromolecules such as proteins, nucleic acids and lipids and their interactions in solution (Fig. 2, 3). This technique is especially valuable for drug discovery as it provides crucial insights into how potential drug candidates interact with target proteins, enzymes and other biomolecules. SAXS also allows for the real-time tracking of protein structure and conformational changes, aiding in the design of small molecules that target modified or pathogenic forms of disease-associated proteins. The Biological SAXS (BioSAXS) beamline specifically offers high flux radiation to support structural biology research at low concentrations, providing a streamlined and highly automated data collection and processing experience. This is enhanced when used with the ‘Coflow’ sample autoloader, which supports in-line size exclusion chromatography (SEC) coupled with SAXS (SEC-SAXS)¹⁸ and can batch-run up to 96 pre-equilibrated samples. This setup is particularly useful for the high-throughput studies of poorly stable, low concentration protein–protein and protein–ligand interactions in solution. Such studies can identify potential drug binding sites, including any induced conformational changes. These findings are highly relevant for targeting proteins associated with infectious diseases,^19–23 phase separation,^24,25 enzymes functions^26,27 and cancer.²⁸

SAXS also plays a crucial role in optimising lipid nanoparticle (LNP) formulation for enhanced drug delivery. Using the temperature-controlled sample environments, researchers can investigate biomolecular–lipid interactions at different temperatures, providing insights into the dynamics and formulations of LNPs.^29–31 This can be applied, for example, to examine mRNA delivery systems,³² to assess their efficiency or immunogenic response generation. Additionally, a Flowthrough Capillary setup allows for dynamic measurements that involve probing biomolecular or LNP interactions tracked in real time. This is particularly important for measuring LNP delivery and behaviours within biological systems, contributing again to the development of improved systems for vaccine and drug delivery.^33,34 Together these advanced SAXS-focused methodologies are crucial for vaccine and drug development by providing essential information about biomolecular dynamics, molecular interactions and the optimisation of different drug delivery systems.

Microscopy: examining cellular processes in drug development

The Infrared Microspectroscopy (IRM) beamline is another powerful tool for enhancing drug discovery research at the AS (Fig. 2, 3). IRM utilises infrared radiation to examine the molecular components of cells and tissues, allowing researchers to map the chemistry of a system and trace the dynamics of drug candidates on cellular processes in close to real time.³⁵ As infrared radiation is non-ionising, it is particularly useful for determining the effects of pharmaceutical drugs on living cells, with specialised IRM techniques allowing for the sub-cellular chemical mapping of single eukaryotic cells.³⁶ As a non-destructive technique,³⁷ IRM provides a clear picture of a cell’s response to external stimuli, including protein expression modifications, lipid profiles and capturing the chemistry of metabolic processes.³⁸ IRM as a key biomedical tool has therefore been applied to the study of many different disease pathologies, including neurotoxicity,³⁹ Alzheimer’s disease,⁴⁰ diabetes mellitus and various blood diseases including malaria and leishmania.^41,42

IRM has also proven a valuable technique in cancer research for the investigation of drug–tumour cell interactions. By comparing molecular changes in cells prior to and following pharmacological intervention, investigators can derive valuable information regarding drug action and drug resistance mechanisms. This information is important in the identification of compounds that can selectively kill cancer cells without significant causing side effects to healthy tissue.⁴³

Imaging: advanced high-resolution techniques in pharmaceutical development

The Imaging group at the AS, with its Imaging and Medical beamline (IMBL) and the Micro-Computed Tomography (MCT) beamline,⁴⁴ provides essential facilities for the investigation of drug behaviour within biological organs and tissues,⁴⁵ and also supports broader studies in organ⁴⁶ and tissue morphology (Fig. 2, 3).⁴⁷ The Imaging beamlines enable the physiology of disease or the activity of pharmaceutical drug candidates to be captured in situ and at high spatial resolution.⁴⁸ Beamline capabilities include high-resolution whole-organ imaging, small- and large-animal scans such as whole-pig CT, and live-animal lung imaging using phase contrast.^49,50 These capabilities are crucial to the development and validation of realistic preclinical models and to the understanding of complex disease environments, both of which underpin bioinnovation and therapeutic developments.

IMBL also allows researchers to examine the distribution and physiological effect of drug treatment within tissues such as tumours and surrounding organs. Preclinical studies conducted at IMBL have provided valuable insights into the effects of radiation therapy on tumour growth, interaction with adjacent tissues, and overall distribution within the host organism. One particularly promising technique being investigated at IMBL is Microbeam Radiation Therapy (MRT), an experimental cancer treatment using synchrotron-generated, spatially fractionated arrays of micrometre-scale radiation fields referred to as microbeams. MRT has demonstrated advanced tumour control ability while minimising damage to healthy tissues. MRT exhibits a post-treatment window of transient localised increased capillary permeability, which offers promising avenues for drug delivery studies, especially in evaluating how therapeutic agents interact with tumour environments under conditions of high-dose radiation. Such techniques are crucial in assessing treatment efficacy and refining therapeutic approaches at the preclinical stage, as demonstrated in recent studies involving glioma-bearing animal models.^51–53 Imaging is therefore a key aspect of drug discovery; to define the distribution of drugs within an organism, map their transit across biological barriers and also their delivery to the targeted tissues of interest.

The IMBL has also been used for high-resolution visualisation and characterisation of breast cancer tissues, including the detection of microcalcifications, tumour vasculature and tissue boundaries, offering new insights into tumour heterogeneity and aiding in the development of more targeted therapies.^54–57 The MCT beamline can be similarly applied but for higher-resolution imaging of smaller samples and is thus perfectly positioned to analyse the complex architectures of cells and tissues, before, during and after drug treatments. In this way the imaging capabilities at the AS are essential to aid in the design of drug delivery systems for the targeted delivery of drugs with limited potential for unwanted off-target effects.

Conclusion

ANSTO’s AS is a cutting-edge bioinnovation facility, providing crucial infrastructure to underpin pharmaceutical development and discovery in Australia. Its comprehensive array of scattering, crystallography, microscopy and imaging beamlines holds unprecedented potential to track the dynamics of molecular and structural biology and thereby enabling scientists to design better and more targeted therapeutic drugs. Through the capacity to facilitate protein and ligand interaction examination, drug target 3-D structure determination, cellular processes exploration, and observation of interactions between drugs within living organisms, the synchrotron holds potential to determine future directions for medical innovation. Owing to ongoing development in synchrotron technologies, coupled with intense focus in research driven by user demand and collaborations, the AS has the potential to be one of the top providers of national and international collaborations, in the effort to achieve novel treatments of disease. With capabilities that are the latest in innovation and backed by the collaborative model of research, the facility is a utility of the pharmaceutical world and a main collaborator towards achieving improved human health.