Queen Elizabeth II Medical Centre

Abstract Mesothelioma is a cancer derived from mesothelial cells, most commonly arising from the pleura or the peritoneum. Immune checkpoint therapy (ICT) has shown survival benefit for pleural mesothelioma, but little is known about the response in peritoneal mesothelioma. Most preclinical mesothelioma models involve subcutaneous cancer cell implantation, which lacks the relevant tumour microenvironment of peritoneal mesothelioma and does not resemble the clinical presentation. We therefore set out to explore the influence of location on the mesothelioma tumour microenvironment, comparing pleural, peritoneal, and subcutaneous models using identical cell line-derived syngeneic mesotheliomas. We found that the peritoneal location conferred an anti-inflammatory tumour microenvironment, characterised by low IFN, TNFα, and STAT signalling activity, low immune cell infiltration and a gene signature associated with non-response to ICT. ICT was effective in subcutaneous models, but the same cell line-derived tumours were irresponsive when inoculated intraperitoneally. Together, these findings show that peritoneal location is associated with an immune suppressive tumour microenvironment.

Third-generation sequencing (TGS), also known as long-read sequencing, has become a promising tool in clinical and research laboratories because it delivers high-resolution results with unmatched throughput. Specialised immunohematology laboratories currently employ sequencing-based methods to characterise rare ABO blood group phenotypes that cannot be identified through serology and genotyping methods. However, routine clinical application of these methods remains elusive due to the absence of validated laboratory protocols and bioinformatics tools. In this study, we have developed and validated a TGS-based workflow for comprehensive determination of the clinically relevant ABO phenotypes from DNA isolated from buccal swabs or whole blood. The region spanning exons 2 to 7 of the ABO gene were amplified and sequenced on MinION 10.4.1 flow cells. Predicted ABO phenotypes were initially determined based on single-nucleotide variants at gDNA261 (rs8176719), gDNA796 (rs8176746), and gDNA803 (rs8176747). However, certain O subtypes lacked the distinguishing deletion (rs8176719) and instead exhibited variations in exon 7 at gDNA802 (rs41302905) and gDNA805, caused by gDNA804 (rs782782485), which differentiate them from A alleles sharing the same nucleotides at gDNA261, gDNA796, and gDNA803. These additional variants were added to the analysis pipeline to identify the additional subtypes. DNA sequence data were sufficient to distinguish between the four clinically relevant ABO blood group phenotypes based on five polymorphic positions. While high sequencing coverage allowed for higher resolution genetic analysis, as few as 20 reads are sufficient for determining the ABO genotype and predicted phenotype of an individual. Typing results generated by this pipeline showed remarkable concordance with both serological results and molecular typing results by an independent laboratory, indicating its accuracy and reliability. This study demonstrates a comprehensive characterisation of clinically relevant ABO blood genotypes and predicted phenotypes using TGS methods. The approach provided a scalable and precise method for routine ABO blood group screening and aided in the development of pioneering bioinformatics tools suitable for clinical and research application.

Resistance to tyrosine kinase inhibitors (TKIs, e.g., sorafenib and lenvatinib) presents a significant hurdle for hepatocellular carcinoma (HCC) treatment, underscoring the need to decipher the underlying mechanisms for improved therapeutic strategies. MicroRNAs (miRNAs) have emerged as critical modulators in HCC progression and TKI resistance. In this study, we report a positive correlation between the expression levels of a tumor suppressor miRNA, miR-142-3p, and increased sensitivity to sorafenib and lenvatinib, supported by clinical data from the BIOSTORM HCC cohort. Overexpression of miR-142-3p in TKI-resistant HCC cells significantly inhibited proliferation and colony formation, induced apoptosis, increased cell cycle arrest at the G2 phase, and reduced migration and invasion by reversing epithelial-mesenchymal transition. Notably, combining miR-142-3p with lenvatinib synergistically inhibited growth in both inherent and acquired TKI-resistant HCC cells by modulating critical signaling pathways, including STAT3, PI3K/AKT, MAPK, YAP1, and by impeding autophagic influx. RNA-sequencing of a TKI-resistant HCC cell line ± miR-142-3p overexpression identified YES1 and TWF1 as direct downstream target genes of miR-142-3p, both of which are key genes associated with drug resistance in HCC. Small interfering RNA (siRNA)-mediated knockdown of these genes mirrored the antitumor effects of miR-142-3p and enhanced TKI sensitivity, with YES1 knockdown decreasing YAP1 phosphorylation, and TWF1 knockdown inhibiting autophagy. Collectively, these findings indicate that restoring miR-142-3p expression or targeting its downstream effectors YES1 and TWF1 offers a promising strategy to overcome drug resistance and improve therapeutic outcome in HCC.

Caesarean section surgical site infections (SSIs) have the potential to cause major complications however there remains a paucity of data on its microbiology and causative organisms. The aim of this study was to identify the organisms causing SSIs in hospitalised patients after lower uterine caesarean section (LUCS). Secondary aims included investigating the differences in organisms isolated from elective versus emergency caesarean sections, and to compare the microbiology of superficial and deep SSIs. This was a large retrospective study reviewing all SSIs after LUCS in Western Australian (WA) public hospitals from 2001 to 2021 with data obtained from the Healthcare Infection Surveillance of Western Australia (HISWA). There were 687 patients identified with an SSI following LUCS over this twenty-year period. The five most common organisms or organism groups isolated were methicillin-susceptible Staphylococcus aureus (MSSA) (30.4%), methicillin-resistant Staphylococcus aureus (MRSA) (14.7%), Enterobacterales (9.5%), Pseudomonas aeruginosa (9.3%), Staphylococcus lugdunensis (5.7%). Group A and B Streptococcus were isolated cumulatively in more than 5% of cases. There were a significantly higher number of superficial SSIs than deep SSIs, and SSIs were significantly more common post emergency LUCS. Current Australian guidelines may not be appropriate in a considerable proportion of patients with LUCS SSIs, and patients with systemic features of infection should receive antibiotics with activity against P. aeruginosa and MRSA. Critically ill patients with features of sepsis or septic shock should be administered a carbapenem agent which has activity against AmpC/ESBL producing Enterobacterales, instead of piperacillin/tazobactam. Staphylococcus lugdunensis may be an under recognised pathogen in LUCS SSIs. This study also highlighted the importance of wound site sampling in SSIs to establish appropriate directed antimicrobial therapy.

Background/Objectives: The current standard of care for assessing chronic kidney disease complicating diabetes (DKD) includes measurement of estimated glomerular filtration rate (eGFR) and urinary albumin:creatinine ratio (uACR) but both tests have limitations. The present study compared the biomarker-based Promarker®D test with conventional biochemical measures for predicting future kidney function decline in adults with type 2 diabetes (T2D). Methods: Baseline concentrations of apolipoprotein A-IV, CD5 antigen-like protein and insulin-like growth factor binding protein 3 were combined with age, serum HDL cholesterol and eGFR to generate PromarkerD risk scores for incident DKD/eGFR decline ≥ 30% (the primary endpoint) in 857 adults with T2D (mean age 65.4 years, 54% males). Logistic regression modelling was used to compare the association of (i) PromarkerD, (ii) eGFR, (iii) uACR, and (iv) eGFR plus uACR with this outcome during 4 years of follow-up. Results: Study participants were classified by PromarkerD as low (63%), moderate (13%), or high risk (24%) for kidney function decline at baseline. Over a mean 4.2 years, 12.5% developed the primary endpoint. PromarkerD scores showed significantly higher predictive performance (area under the receiver operating characteristic curve (AUC) 0.88 (95% confidence interval (CI) 0.85-0.91)) compared to conventional biochemical measures (AUC = 0.63-0.82). There was a progressive increase in risk with moderate and high risk by PromarkerD exhibiting greater odds of the primary endpoint compared to those at low risk (odds ratios (OR) (95% CI) 8.11 (3.99-16.94) and 21.34 (12.03-40.54), respectively, both p < 0.001). Conclusions: PromarkerD more accurately identifies adults with T2D at risk of kidney function decline than current usual care biochemical tests.

Abstract Skin scars are a significant clinical challenge, with poor appearance and increased tissue stiffness affecting both physical and psychological wellbeing. Lysyl oxidases are a family of enzymes that catalyze collagen crosslinking, a key factor in scar pathophysiology. Here we report a randomized, double-blind, placebo-controlled Phase 1 clinical trial to assess the safety and tolerability of PXS-6302, a topical pan-lysyl oxidase inhibitor, in treating mature scars (ACTRN12621001545853). Fifty participants with scars were enrolled and PXS-6302 or placebo cream applied to a 10 cm2area for three months. No severe adverse events were reported. All treatment-related adverse events (AEs) were localized skin reactions. Treatment with PXS-6302 significantly inhibited lysyl oxidase activity (66%). Hydroxyproline (a marker for collagen) and total protein concentration in the scar were significantly reduced in the PXS-6302 treatment group compared to placebo. Optical coherence tomography (OCT) was used to measure vascularity and attenuation (a marker of extracellular matrix composition). PXS-6302 treatment significantly increased vessel density at 3 months compared to baseline. Tissue attenuation was also significantly increased in PXS-6302 treated participants compared to baseline, suggesting extracellular matrix was becoming increasingly similar to normal skin. No significant differences between placebo and PXS-6302 treatment groups were observed in Patient Observer Scar Assessment Scale (POSAS) scores at study conclusion. To our knowledge, this study represents the first demonstration of a safe and effective pharmaceutical intervention that significantly improves the molecular composition of established scar extracellular matrix in humans. Pan-lysyl oxidase inhibition therefore represents a potential paradigm shift for the amelioration of scarring.One Sentence SummaryMature scars remodeled after treatment with a topical cream for three months.

Background/Objectives: Current inflammatory bowel disease (IBD) treatments focus on symptomatic relief, highlighting the need for innovative approaches. Dysregulation of the cannabinoid 1 (CB1) receptor, part of the endocannabinoid system, is linked to colitis. While tetrahydrocannabinol (THC) alleviates colitis via CB1 activation, its psychotropic effects limit clinical use. ZCZ011, a CB1R allosteric modulator, and cannabidiol (CBD), a non-psychoactive cannabinoid, offer alternatives. This study investigated combining sub-therapeutic THC doses with ZCZ011 or CBD in a murine model of dextran sodium sulphate (DSS)-induced colitis. Methods: Acute colitis was induced with 4% DSS for 7 days, followed by 3 days of water. Chronic colitis was modelled over 24 days with alternating DSS concentrations. The combination of 2.5 mg/kg THC with 20 mg/kg ZCZ011 or 10 mg/kg CBD was evaluated. Key markers were assessed to determine efficacy and safety, including disease activity index (DAI), inflammation, cytokine levels, GLP-1, and organ health. Results: DSS-induced colitis resulted in increased DAI scores, cytokines, organ inflammation and dysregulation of GLP-1 and ammonia. THC at 10 mg/kg significantly improved colitis markers but was ineffective at 2.5 and 5 mg/kg. ZCZ011 alone showed transient effects. However, combining 2.5 mg/kg THC with either 20 mg/kg ZCZ011 or 10 mg/kg CBD significantly alleviated colitis markers, restored colon integrity and reestablished GLP-1 homeostasis. This combination also maintained favourable haematological and biochemical profiles, including a notable reduction in colitis-induced elevated ammonia levels. Conclusions: This study demonstrates the synergistic potential of low-dose THC combined with CBD or ZCZ011 as a novel, effective and safer therapeutic strategy for ulcerative colitis.