SAGC Collaborative Projects

Natural killer (NK) cells offer exciting potential for cancer immunotherapy because of their innate ability to target and destroy tumor cells without the need for prior sensitization. They can recognize and kill cancerous or virally infected cells through direct cytotoxicity and by releasing cytokines that modulate the immune response. NK cells are particularly effective at eliminating cancer cells that evade the adaptive immune system, and their function can be enhanced through techniques like chimeric antigen receptor (CAR) engineering.

NK cells are difficult to obtain from traditional donor sources because peripheral blood provides a limited supply, and the quality of NK cells varies between donors. When expanded in vitro, they can lose their effectiveness (functional exhaustion), and modifying them through gene editing is complex. These limitations make alternative sources, like iPSC-derived NK cells, highly appealing. 

Recent work led by Dr. Gustavo R Rossi (Guimaraes Group - Translational innate immunotherapy, The University of Queensland) detailed a two-step method for generating high-purity Natural Killer cells from an induced pluripotent stem cell (iPSC) derived cell lines (iNK cells). These iNK cells displayed key NK cell markers and enhanced cytotoxicity, showed resilience to cryopreservation, and their cytotoxic potential was further improved by incorporating a chimeric antigen receptor (CAR) construct. 

“A key advantage of the technology was being able to fix our cells from various sources at different time points, and safely ship them to the SAGC for barcoding and sequencing.” says Dr. Fernando Guimaraes.

Having a scalable and consistent source of iNK cells would help address current limitations in traditional NK cell sources, and has the potential to play an exciting role in cancer immunotherapies.

Single Cell RNA Sequencing using Parse Biosciences Evercodeᵀᴹ combinatorial barcoding technology: An important element of this study was comparing transcriptomic profiles of NK cells derived from iPSC with those derived from peripheral blood (PBMC) and cord blood units (CBU). Dr. Fernando Guimaraes and Gustavo became interested in using the Evercodeᵀᴹ technology for their experiment, because of the advantage of being able to fix and store cells from different sources across different time points, and batch together for processing and sequencing. This led them to the SAGC, who with the support of Dr. Jiyoti Verma (Field Application Specialist from Decode Science) conducted the combinatorial barcoding workflow and performed deep sequencing of the libraries on the MGI DNBSEQ G400.

Single Cell Data Processing:  RNA sequencing data was analysed using Trailmaker, Parse Bioscience’s pipeline that aligns reads to a reference genome, generates gene count matrices and performs various QC steps, like filtering out low quality cells and removes doublets etc.

GR Rossi et al. Immunology & Cell Biology 2024; 1–11

Sanfilippo Syndrome is a paediatric lysosomal storage disorder that is commonly known as ‘childhood-onset dementia’, due to the severe cognitive impairment and neuronal loss associated with it. Children affected by this disorder experience symptoms such as hyperactivity, speech and hearing loss, sleep disturbances and motor impairment, all of which are followed by premature death around mid to late adolescence. While gene therapy has shown promising results in children diagnosed under the age of 2, most children are not diagnosed until 4 or 5 years of age. This highlights a critical need to identify new treatment options that could help manage symptoms, slow or stop disease progression and improve quality of life.

To begin to address this gap, the Laboratory for Human Neurophysiology and Genetics, led by Professor Cedric Bardy at SAHMRI and Flinders University, worked in collaboration with the SAGC. The aim was to understand the molecular changes underlying disease phenotypes and identify genetic targets for repurposed therapeutics. Using a human-derived neuronal model of Sanfilippo Syndrome along with state-of-the-art single-cell transcriptomic profiling, electrophysiology and machine learning, the team identified key phenotypes that may cause neurodegeneration. 

As part of this project, the SAGC created mRNA libraries from single-nuclei extractions and sequencing. Together, we have sequenced multiple patient lines and identified dysregulated genes and pathways that can be targeted with repurposed drugs. This has helped Cedric’s team to identify and validate therapeutic targets for Sanfilippo Syndrome, which they hope will fast-track to clinical trials and improve the quality of life for patients.

Sheep experience pain in response to necessary management practices like castration and tail docking, as well as during lambing and due to other natural hazards of life. At the very least that is acute pain but, if sheep are anything like humans and other mammals, then a proportion of sheep will also experience some form of persistent pathological pain. 

To address wellbeing concerns around pain in sheep, objective pain measurement tools are needed to answer questions like: Do you have pain now? How much pain are you in? And have you had pain in the past? This then enables us to develop, and demonstrate the effectiveness of, pain mitigation strategies, as well as to provide objective evidence in support of claims of pain mitigation, and to support ongoing evaluation of best practice and evidence–based recommendations for managing sheep wellbeing.

Led by Professor Mark Hutchinson, researchers at the University of Adelaide’s Neuroimmunopharmacology Lab and Davies Livestock Research Centre are working to identify objective blood-based biomarkers of acute and persistent pain in sheep, with funding support from the Australian Research Council, Meat and Livestock Australia, the University of Adelaide’s Davies Livestock Research Centre, and the Livestock SA Sheep Industry Fund.

As part of this research program, the team have been working with SAGC to identify plasma and blood transcriptomic changes that occur in response to painful husbandry procedures, that could be used to diagnose/quantify acute or persistent pain in sheep. 

Small RNA-seq of nearly 500 sheep plasma samples has been completed to date, alongside ongoing optimisation of protocols for sheep whole blood total RNA sequencing. By combining these blood transcriptomic data with blood cell phenotyping, brain and spinal cord histology and histochemistry, behaviour, and quantitative sensory testing data from their sheep trials, the team aim to not only identify novel objective pain biomarkers, but also build an integrated picture of the short- and long-term molecular, cellular, systems and behavioural consequences of these painful marking procedures, and the mechanisms underlying any transition to and maintenance of chronic pain, in sheep. 

2023 saw the establishment of the South Australian node of the Australian Alliance for Indigenous Genomics (ALIGN), on the back of the successful 2021 Genomics Health Futures Mission grant to advance the benefits of Genomic Medicine for Aboriginal and Torres Strait Islander peoples.

Key priorities of the SA node, which is partially led by SAGC, revolve around extensive multi-omics analyses in the PROPHECY (Predicting Renal, Ophthalmic and Heart Events in the Aboriginal Community) study, Aboriginal-led policy development for Indigenous Genomics projects in SA and capacity development of a SA Indigenous genomics workforce. To help facilitate and guide this work, a South Australian Aboriginal Governance Committee has been established to provide Aboriginal insight and oversight into culturally safe genomics research practices so that we can work towards prioritising community benefit, health and wellbeing. The PROPHECY study is the largest longitudinal study of diabetes in Aboriginal Australians ever conducted. PROPHECY aims to understand why Aboriginal people get type II diabetes at a much younger age, and why they experience associated complications such as kidney disease and blindness at a much younger age than non-Aboriginal Australians.

People who have COVID-19 will excrete fragments of the SARS-CoV-2 virus through faecal waste or other bodily fluids into the wastewater stream through the sewerage system. A wastewater surveillance program therefore provides another mechanism through which community transmission can be monitored to complement clinical surveillance. The point of difference for wastewater surveillance is that it does not rely on clinical presentation by individuals. Wastewater surveillance will detect virus fragments from people who are weakly symptomatic or asymptomatic. The use of wastewater surveillance for detecting and quantifying the presence of SARS-CoV-2 virus fragments was expanded in 2022 to identify the variants of the virus circulating in the community in a manner that is independent of clinical presentation by individuals.

The aim was to provide a complementary source of information to analysis of variants detected from sequencing of clinical isolates. In South Australia, SA Water, SA Health and the SAGC ran a PCR-based screening wastewater surveillance program over the course of a year, whereby samples were collected from the major metropolitan wastewater treatment plants twice a week. These plants service about 65% of the SA population. PCR products from these samples were submitted to SAGC for sequencing. The resulting data produced was used by SA Health to assess changes in prevalence of variants and to monitor the appearance of new variants of concern (results are shown in the figure 1).

Emergence of COVID-19 and the ensuing pandemic has demonstrated the continued threat infectious diseases can pose to our health, economy, and lifestyle and highlighted the importance of vaccines as a countermeasure. COVID-19 vaccines were developed and approved in approximately 350 days, which was 5-10 times faster than ever before. However, even greater speed is needed, and 100-days for vaccine development is the current target. Achieving this goal will require a shift in current vaccine paradigms and development of new flexible, rapid, scalable technologies.

The Sementis viral vector vaccine platform (SCV) has been designed to deliver safe, versatile, and scale-able vaccines and is being positioned for pandemic preparedness. Sementis and the Experimental Therapeutics Laboratory at the University of South Australia (UniSA) have worked together to develop a multi-pathogen Chikungunya and Zika virus vaccine and a first-generation COVID-19 vaccine to demonstrate the immunogenicity of the vaccine platform and utility in infectious disease. UniSA has further collaborated with Sementis, as part of a broader global network to position the vaccine platform for pandemic preparedness. This included supporting the introduction of new technology to enable rapid construction and characterisation of new vaccines.

Dust and soil were collected from three sites with differing soil properties across South Australia. The resulting ITS and 16S sequencing data, generated at the SAGC, showed that it was possible to perform analysis from a single swabbed sample and that there were unique biological and chemical signatures between sites. When modelling bacterial and fungal diversity, Dr. Jennifer Young and team found samples were correctly predicted using dust 67% and 56% of the time and using soil 56% and 22% of the time for bacteria and fungi communities respectively. This proof-of-concept study found that metabarcoding of dust samples can generate bacteria and fungi community profiles that are unique to sites. While there is still a long way to go in this field, both metabarcoding and biogeochemical analyses of dust samples show potential for applications in forensic science.

Nicole R. Foster et. al, 2023, The secret hidden in dust: Assessing the potential to use biological and chemical properties of the airborne fraction of soil for provenance assignment and forensic casework, Forensic Science International: Genetics, Volume 67

Irritable Bowel Syndrome (IBS) is a chronic disorder of the gastrointestinal tract affecting ~11% of the global population. Chronic visceral pain (CVP) is the most debilitating symptom associated with IBS. Despite this burden of disease, effective therapies for CVP associated with IBS are lacking. The first step in finding new treatments is understanding how pain from the gut is generated and transmitted to the brain via the microbiome-gut-brain axis and determining how these processes change in IBS. The Visceral Pain Research Group, led by Professor Stuart Brierley at SAHMRI is working in conjunction with SAGC to fill this knowledge gap. The goal is to identify changes in the molecular profile of precise cell types within the microbiome-gut-brain axis to determine how changes to these molecular profiles drive altered function and generate the abnormal pain signalling characteristic of IBS.

The genes encoding isocitrate dehydrogenase 1 and 2 (IDH1/2) are mutated in brain cancer and leukaemia and drive the production of the oncometabolite (R)-2-hydroxyglutarate (R-2HG). However, specific differences between these two mutations with targetable implications have not been described. Dr Dan Thomas is a clinician and researcher who runs the Myeloid Metabolism Laboratory at SAHMRI. His team has recently discovered a novel susceptibility specifically for IDH1 mutation (mIDH1) by identifying the lipid synthesis enzyme ACC1 (acetyl CoA carboxylase 1) as a synthetic lethal target, published recently in Cancer Discovery and highlighted as one of the most important papers for 2023 by the European Cancer Association. By analyzing the metabolome of primary patient cells, his team identified a mIDH1-specific reduction in fatty acids compared to healthy progenitor cells that was not observed in IDH2-mutant cancer. Moreover, mIDH1 cells exhibited an increase in acyl-carnitine linked fatty acids destined for the mitochondria, indicating a switch to beta-oxidation. Overall, their data show that mIDH1 cancer cells have a higher dependency on both exogenous and de novo fatty acids than mIDH2 cancers.