The effect of three years of vitamin D supplementation on erectile dysfunction: Results from the randomized placebo-controlled D-Health Trial.

BACKGROUND & AIMS: Erectile dysfunction is common among older men and has been associated with low serum 25-hydroxy vitamin D concentration. However, this association may be due to uncontrolled confounding, and there is a paucity of evidence from interventional studies. We aimed to examine the effect of vitamin D supplementation on the prevalence of erectile dysfunction, in an exploratory analysis using data from a large randomized controlled trial. METHODS: The D-Health Trial recruited Australians aged 60-84 years between January 2014 and May 2015 and randomly assigned them to supplementation with 60,000 IU of vitamin D or placebo per month for up to 5 years. Blood samples were collected annually from randomly selected participants (total N = 3943). We assessed erectile dysfunction at the end of the third year of follow-up. We used log-binomial regression to examine the effect of vitamin D on the prevalence of erectile dysfunction overall, and within sub-groups. RESULTS: Of the 11,530 men enrolled, 8920 (77.4 %) completed the erectile dysfunction question and were included in the analysis. After three years of supplementation, the mean serum 25-hydroxy vitamin D concentration was 76 nmol/L (standard deviation (SD) 24.94) in the placebo group and 106 nmol/L (SD 26.76) in the vitamin D group (p < 0.0001). The prevalence of erectile dysfunction was 58.8 % and 59.0 % in the vitamin D and placebo groups, respectively (prevalence ratio 1.00, 95 % CI 0.97, 1.03); there was no evidence of an effect of vitamin D in any subgroup analyses. CONCLUSION: Supplementing older men with vitamin D is unlikely to prevent or improve erectile dysfunction. CLINICAL TRIALS REGISTRY: (ACTRN12613000743763).

Impact of disease recurrence on the supportive care needs of patients with ovarian cancer and their caregivers.

OBJECTIVE: We aimed to explore the supportive care needs of ovarian cancer patients and their caregivers before and after the first cancer recurrence, the top unmet needs after recurrence, and the relationship between patient and caregiver needs at recurrence. METHODS: Participants were 288 patients and 140 caregivers from the Australian Ovarian Cancer Study-Quality of Life (AOCS-QoL) cohort. They completed Supportive Care Needs Surveys (patients: SCNS-SF34, caregivers: SCNS-P&C44) every three-to-six months for up to two years. Linear mixed models tracked changes in needs over time. We calculated the percentage reporting moderate-to-high needs after recurrence. LASSO regression analysed patient-caregiver need relationships. RESULTS: Both patients' and caregivers' psychological, health system/service and information needs increased with recurrence along with patients' support and physical needs. These remained stable at nine months after recurrence. Dominant patient needs post-recurrence included 'fear of recurrence' (38%) and 'concerns about the worries of those close' (34%), while caregivers expressed 'concerns about recurrence' (41%) and 'recovery of the patient not turning out as expected' (31%). Among dyads, when patients had 'fears about the cancer spreading' this was associated with caregivers having a need for help with 'reducing stress in the patients' life'; when caregivers had concerns about 'recurrence' this was associated with patients needing help with 'uncertainty about the future' and 'information about things they can do to help themselves'. CONCLUSIONS: Recurrent ovarian cancer intensifies disease-related fears and concerns for patients and loved ones. Addressing dyadic concerns through supportive care interventions may enhance cohesion during the challenging journey of recurrent disease.

A novel saliva-based miRNA profile to diagnose and predict oral cancer.

Oral cancer (OC) is the most common form of head and neck cancer. Despite the high incidence and unfavourable patient outcomes, currently, there are no biomarkers for the early detection of OC. This study aims to discover, develop, and validate a novel saliva-based microRNA signature for early diagnosis and prediction of OC risk in oral potentially malignant disorders (OPMD). The Cancer Genome Atlas (TCGA) miRNA sequencing data and small RNA sequencing data of saliva samples were used to discover differentially expressed miRNAs. Identified miRNAs were validated in saliva samples of OC (n = 50), OPMD (n = 52), and controls (n = 60) using quantitative real-time PCR. Eight differentially expressed miRNAs (miR-7-5p, miR-10b-5p, miR-182-5p, miR-215-5p, miR-431-5p, miR-486-3p, miR-3614-5p, and miR-4707-3p) were identified in the discovery phase and were validated. The efficiency of our eight-miRNA signature to discriminate OC and controls was: area under curve (AUC): 0.954, sensitivity: 86%, specificity: 90%, positive predictive value (PPV): 87.8% and negative predictive value (NPV): 88.5% whereas between OC and OPMD was: AUC: 0.911, sensitivity: 90%, specificity: 82.7%, PPV: 74.2% and NPV: 89.6%. We have developed a risk probability score to predict the presence or risk of OC in OPMD patients. We established a salivary miRNA signature that can aid in diagnosing and predicting OC, revolutionising the management of patients with OPMD. Together, our results shed new light on the management of OC by salivary miRNAs to the clinical utility of using miRNAs derived from saliva samples.

Human papillomavirus (HPV) DNA methylation changes in HPV-associated head and neck cancer.

Despite the rising incidence, currently, there are no early detection methods for HPV-driven HNC (HPV-HNC). Cervical cancer studies suggest that HPV DNA methylation changes can be used as a biomarker to discriminate cancer patients from HPV-infected individuals. As such, this study was designed to establish a protocol to evaluate DNA methylation changes in HPV late genes and long control region (LCR) in saliva samples of HPV-HNC patients and HPV-positive controls. Higher methylation levels were detected in HPV late genes (L1 and L2) in both tumour and saliva samples of HPV-HNC patients compared with HPV-positive controls. Moreover, methylation patterns between tumours and corresponding saliva samples were observed to have a strong correlation (Passing-Bablok regression analysis; τ = 0.7483, P < 0.0001). Considering the differences between HNC and controls in methylation levels in late genes, and considering primer amplification efficiencies, 13 CpG sites located at L1 and L2 genes were selected for further evaluation. A total of 18 HNC saliva samples and 10 control saliva samples were assessed for the methylation levels in the selected sites. From the CpG sites evaluated statistically significant differences were identified for CpG sites at L2-CpG 6 (P = 0.0004), L1-CpG 3 (P = 0.0144), L1-CpG 2 (P = 0.0395) and L2-CpG 19 (P = 0.0455). Our pilot data indicate that higher levels of DNA methylation in HPV late genes are indicative of HPV-HNC risk, and it is a potential supplementary biomarker for salivary HPV detection-based HPV-HNC screening.

Imetelstat-mediated alterations in fatty acid metabolism to induce ferroptosis as a therapeutic strategy for acute myeloid leukemia.

Telomerase enables replicative immortality in most cancers including acute myeloid leukemia (AML). Imetelstat is a first-in-class telomerase inhibitor with clinical efficacy in myelofibrosis and myelodysplastic syndromes. Here, we develop an AML patient-derived xenograft resource and perform integrated genomics, transcriptomics and lipidomics analyses combined with functional genetics to identify key mediators of imetelstat efficacy. In a randomized phase II-like preclinical trial in patient-derived xenografts, imetelstat effectively diminishes AML burden and preferentially targets subgroups containing mutant NRAS and oxidative stress-associated gene expression signatures. Unbiased, genome-wide CRISPR/Cas9 editing identifies ferroptosis regulators as key mediators of imetelstat efficacy. Imetelstat promotes the formation of polyunsaturated fatty acid-containing phospholipids, causing excessive levels of lipid peroxidation and oxidative stress. Pharmacological inhibition of ferroptosis diminishes imetelstat efficacy. We leverage these mechanistic insights to develop an optimized therapeutic strategy using oxidative stress-inducing chemotherapy to sensitize patient samples to imetelstat causing substantial disease control in AML.

Profiling salivary miRNA expression levels in Fanconi anemia patients - a pilot study.

The overarching goal of this study is to predict the risk of developing oral squamous cell carcinoma (OSCC) in Fanconi anemia (FA) patients. We have compared the microRNA (miRNA, miR) expression levels in saliva samples from FA patients (n = 50) who are at a low-moderate and/or high risk of developing OSCC to saliva samples from healthy controls (n = 16). The miRNA expression levels in saliva samples were quantified using qPCR. We observed that miR-744, miR-150-5P, and miR-146B-5P had the best discriminatory capacity between FA patients and controls, with an area under the curve (AUC) of 94.0%, 92.9% and 85.3%, respectively. Our data suggest that miR-1, miR-146B-5P, miR-150-5P, miR-155-5P, and miR-744 could be used as panel to predict the risk of developing OSCC in FA patients, with a 89.3% sensitivity and a 68.2% specificity (AUC = 81.5%). Our preliminary data support the notion that the expression levels of salivary miRNAs have the potential to predict the risk of developing OSCC in FA patients and in the future may reduce deaths associated with OSCC.

Prognostic utility of serum NT-proBNP (fragments 1-76aa and 13-71aa) and galectin-3 in predicting death and re-hospitalisation due to cardiovascular events in patients with heart failure.

Patients with heart failure (HF) are at a higher risk of rehospitalisation. In this study, we investigated the prognostic utility of galectin-3 (Gal-3) and NT-proBNP fragments (1-76aa and 13-71aa) as biomarkers to predict outcomes for patients with HF. We collected blood samples from patients with HF (n = 101). Gal-3 and NT-proBNP fragments (1-76aa and 13-71aa) concentrations were measured by immunoassay. Survival analysis and Cox proportional regression models were used to determine the prognostic utility of Gal-3 and NT-proBNP fragments. In patients with increased baseline levels of NT-proBNP1-76 the time to primary endpoint (cardiovascular death or re-hospitalisation) was significantly shorter (p = 0.0058), but not in patient with increased baseline levels of Gal-3 or NTproBNP13-71. Patients with increased levels of NT-proBNP13-71aa at 1 month showed reduced time to the primary endpoint (p = 0.0123). Our findings demonstrated that Gal-3 and NT-proBNP can be used as prognostic biomarkers to stratify patients with HF.

The Unmet Needs of Pancreatic Cancer Carers Are Associated with Anxiety and Depression in Patients and Carers.

Pancreatic cancer has one of the lowest survival rates, and patients experience debilitating symptoms. Family carers provide essential daily care. This study determined the prevalence of and risk factors for unmet supportive care needs among carers for pancreatic cancer patients and examined which carer needs were associated with anxiety and depression in carers and patients. Eighty-four pancreatic cancer patients and their carers were recruited. The carers completed a needs survey (SCNS-P&C). Both carers and patients completed the Hospital Anxiety and Depression Scale. Log binomial regression was used to identify associations between carer needs and anxiety and depression among carers and patients. The top 10 moderate-to-high unmet needs reported by ≥28% of carers were related to healthcare (e.g., discussing concerns with doctors) and information need domains (e.g., information about a patient's physical needs), plus one other item related to hospital parking. Being male or caring for a patient within 4 months of their diagnosis were associated with greater unmet needs. Some unmet needs, including 'accessing information about treatments' and 'being involved in patient care', were associated with both carers and patients having anxiety and depression. Carers should be involved in health care consultations and provided with information and opportunities to discuss concerns.

A comparison between mutational profiles in tumour tissue DNA and circulating tumour DNA in head and neck squamous cell carcinoma - A systematic review.

BACKGROUND: Head and neck cancer is the seventh most common malignancy globally. Head and neck squamous cell carcinoma (HNSCC) originates from squamous cells and 90% of HNC are HNSCC. The gold standard for diagnosing HNSCC is tissue biopsy. However, given tumour heterogeneity, biopsies may miss important cancer-associated molecular signatures, and more importantly, after the tumour is excised, there is no means of tracking response to treatment in patients. Captured under liquid biopsy, circulating tumour DNA (ctDNA), may identify in vivo molecular genotypes and complements tumour tissue analysis in cancer management. A systematic search was conducted in PubMed, Embase, Scopus and the Cochran Library between 2012 to early 2023 on ctDNA in HNSCC using publications written in English. We summarise 20 studies that compared mutational profiles between tumour tissue DNA (tDNA) and ctDNA, using a cohort of 631 HNSCC patients and 139 controls. Among these studies, the concordance rates varied greatly and the most mutated and the most concordant gene was TP53, followed by PIK3CA, CDKN2A, NOTCH1 and FAT1. Concordant variants were mainly found in Stage IV tumours, and the mutation type is mostly single nucleotide variants (SNV). We conclude that, as a biomarker for HNSCC, ctDNA demonstrates great promise as it recapitulates tumour genotypes, however additional multi-central trials are needed.

Discrepancies in tumor mutation burden reporting from sequential endobronchial ultrasound transbronchial needle aspiration samples within single lymph node stations - brief report.

Introduction: Tumour Mutation Burden (TMB) is a potential biomarker for immune cancer therapies. Here we investigated parameters that might affect TMB using duplicate cytology smears obtained from endobronchial ultrasound transbronchial needle aspiration (EBUS TBNA)-sampled malignant lymph nodes. Methods: Individual Diff-Quik cytology smears were prepared for each needle pass. DNA extracted from each smear underwent sequencing using large gene panel (TruSight Oncology 500 (TSO500 - Illumina)). TMB was estimated using the TSO500 Local App v. 2.0 (Illumina). Results: Twenty patients had two or more Diff-Quik smears (total 45 smears) which passed sequencing quality control. Average smear TMB was 8.7 ± 5.0 mutations per megabase (Mb). Sixteen of the 20 patients had paired samples with minimal differences in TMB score (average difference 1.3 ± 0.85). Paired samples from 13 patients had concordant TMB (scores below or above a threshold of 10 mutations/Mb). Markedly discrepant TMB was observed in four cases, with an average difference of 11.3 ± 2.7 mutations/Mb. Factors affecting TMB calling included sample tumour content, the amount of DNA used in sequencing, and bone fide heterogeneity of node tumour between paired samples. Conclusion: TMB assessment is feasible from EBUS-TBNA smears from a single needle pass. Repeated samples of a lymph node station have minimal variation in TMB in most cases. However, this novel data shows how tumour content and minor change in site of node sampling can impact TMB. Further study is needed on whether all node aspirates should be combined in 1 sample, or whether testing independent nodes using smears is needed.

Changing Prevalence of Medication Use in People with Cirrhosis: A Retrospective Cohort Study Using Pharmaceutical Benefits Scheme Data.

BACKGROUND: Safe and appropriate use of medicines is essential to improve health outcomes in cirrhosis. However, little is known about the number and type of medicines dispensed to people with cirrhosis in Australia, as this predominantly occurs in the community. We aimed to characterise the prescriptions dispensed to people with cirrhosis and explore changes in the use of medication groups over time. METHODS: Pharmaceutical Benefits Scheme data between 1 January 2016 and 30 June 2020 was extracted for consenting CirCare participants (multi-site, prospective, observational study). Prescriptions dispensed from cirrhosis diagnosis until liver transplant or death were included. Safety classifications for dispensed medicines were defined using published evidence-based recommendations. The pattern of medication use was analysed in 6-monthly time intervals. Generalised estimating equations models were used to estimate the change in consumption of medicines over time. RESULTS: Five hundred twenty-two patients (mean age 60 years, 70% male, 34% decompensated at recruitment) were dispensed 89,615 prescriptions during the follow-up period, representing a median of 136 [interquartile range (IQR) 62-237] prescriptions and a median of 16 (IQR 11-23) unique medicines per patient (total n = 9306 medicines). The most commonly used medicines were proton pump inhibitors (PPIs) (dispensed at least once to 73% of patients), opioids (68%) and antibiotics (89%). Polypharmacy was prevalent, with 59-69% of observed participants in each time period dispensed five or more unique medicines. Prescription medication use increased over time (p < 0.001) independently of age, comorbidity burden and liver disease aetiology. The likelihood of taking PPIs, opioids, antidepressants and inhaled medicines also increased with each successive time period. Use of angiotensin therapies, metformin and statins differed over time between patients with compensated versus decompensated cirrhosis. General practitioners prescribed 69% of dispensed medicines, including a higher proportion of 'unsafe' and 'safety unknown' medicines compared with consultants/specialists (p < 0.001). CONCLUSIONS: Polypharmacy is common in people with cirrhosis and some medication groups may be overused. Pharmacovigilance is required and future medication safety efforts should target high-risk prescribing practices and promote medication rationalisation in the community.

Diabetes mellitus and the progression of non-alcoholic fatty liver disease to decompensated cirrhosis: a retrospective cohort study.

OBJECTIVE: To determine the incidence of decompensated cirrhosis and associated risk factors in people hospitalised with non-alcoholic fatty liver disease (NAFLD) or non-alcoholic steatohepatitis (NASH) with or without cirrhosis. DESIGN: Retrospective cohort study; analysis of linked Queensland Hospital Admitted Patient Data Collection, Queensland Registry of Births, Deaths and Marriages, and Queensland Cancer Register data. SETTING, PARTICIPANTS: Queensland residents aged 20 years or older admitted to Queensland hospitals with NAFLD/NASH during 1 July 2009 - 31 December 2018. MAIN OUTCOME MEASURES: Progression to decompensated cirrhosis (ascites, hepatic encephalopathy, or oesophageal variceal bleeding). RESULTS: We included data for 8006 patients in our analysis (10 082 admissions), including 4632 women (58%) and 2514 people with diabetes mellitus (31%); median follow-up time was 4.6 years (interquartile range, 2.7-7.2 years). Three hundred and fifty-one people (4.4%) experienced decompensated cirrhosis during the follow-up period. Of the 6900 people without cirrhosis, 4.5% (95% confidence interval [CI], 3.6-5.7%) experienced decompensated cirrhosis within ten years (mean, 0.5% per year; 95% CI, 0.4-0.6% per year); risk of progression was greater for people aged 70 years or older (v 20-39 years: adjusted hazard ratio [aHR], 4.7; 95% CI, 2.0-11.0) and those who had extrahepatic cancers (aHR, 5.0; 95% CI, 3.0-8.2), history of major cardiovascular events (aHR, 1.9; 95% CI, 1.2-3.1), or diabetes mellitus (aHR, 2.8; 95% CI, 2.0-3.9). Of the 1106 people with cirrhosis, 32.4% (95% CI, 27.2-38.3%) experienced decompensated cirrhosis within ten years (mean, 5.5% per year; 95% CI, 4.8-6.3% per year); risk of progression was greater for those with portal hypertension (aHR, 1.8; 95% CI, 1.3-2.7), extrahepatic cancer (aHR, 1.8; 95% CI, 1.1-2.9), or diabetes mellitus (aHR, 1.5; 95% CI, 1.1-2.0). Compared with people who had neither cirrhosis nor diabetes mellitus, the risk of decompensation was greater for people with cirrhosis (aHR, 10.7; 95% CI, 7.6-15.0) or cirrhosis and diabetes mellitus (aHR, 14.4; 95% CI, 10.1-20.6). CONCLUSIONS: Given the greater risk of progression to cirrhosis decompensation in people with diabetes mellitus, a disorder common in people with NAFLD/NASH, identifying advanced fibrosis and providing appropriate treatment for averting disease progression is vital.

The Effect of Vitamin D Supplementation on Hypothyroidism in the Randomized Controlled D-Health Trial.

Background: Hypothyroidism is common, and in iodine-sufficient areas, it is primarily caused by autoimmune destruction of the thyroid gland. Observational studies have consistently shown an inverse association between serum 25-hydroxyvitamin D concentration and autoimmune diseases; however, there is a lack of evidence from randomized controlled trials to support a benefit of vitamin D supplementation, particularly for autoimmune thyroid diseases. We, therefore, aimed to assess the effect of vitamin D supplementation on the incidence of hypothyroidism. Methods: We analyzed data from the D-Health Trial (n = 21,315), a randomized double-blind placebo-controlled trial of 60,000 international units per month of supplemental vitamin D3 among Australians aged 60 years and over. Hypothyroidism, a tertiary outcome of the D-Health Trial, was defined by treatment with levothyroxine, ascertained through linkage with the Australian Pharmaceutical Benefits Scheme. The outcome was time to first prescription of levothyroxine. We began follow-up at 12 months after randomization; people who had died or who had been dispensed levothyroxine during the first year were excluded. Flexible parametric survival models were used to assess the effect of vitamin D supplementation on hypothyroidism, overall and within strata defined by age, sex, body mass index, and predicted baseline vitamin D status. Results: We included 17,851 participants in the main analysis (vitamin D = 8939; placebo = 8912). During a median follow-up of 4.1 years (interquartile range 4.1-4.1), 293 participants developed hypothyroidism (vitamin D = 138 [1.5%]; placebo = 155 [1.7%]). Vitamin D supplementation did not significantly reduce the incidence of hypothyroidism (overall hazard ratio [HR] 0.89; 95% confidence interval [CI] 0.71-1.12). There was some suggestion of an effect in females (overall HR 0.78; CI 0.58-1.06) but not in males (overall HR 1.06; CI 0.74-1.50; p interaction 0.20). Conclusions: Vitamin D supplementation did not reduce the incidence of hypothyroidism overall; however, the possible beneficial effect observed in females warrants further investigation. Clinical Trial Registration: Australian New Zealand Clinical Trials Registry: ACTRN12613000743763.

A Decrease in Maternal Iron Levels Is the Predominant Factor Suppressing Hepcidin during Pregnancy in Mice.

In order to supply adequate iron during pregnancy, the levels of the iron regulatory hormone hepcidin in the maternal circulation are suppressed, thereby increasing dietary iron absorption and storage iron release. Whether this decrease in maternal hepcidin is caused by changes in factors known to regulate hepcidin expression, or by other unidentified pregnancy factors, is not known. To investigate this, we examined iron parameters during pregnancy in mice. We observed that hepatic iron stores and transferrin saturation, both established regulators of hepcidin production, were decreased in mid and late pregnancy in normal and iron loaded dams, indicating an increase in iron utilization. This can be explained by a significant increase in maternal erythropoiesis, a known suppressor of hepcidin production, by mid-pregnancy, as indicated by an elevation in circulating erythropoietin and an increase in spleen size and splenic iron uptake. Iron utilization increased further in late pregnancy due to elevated fetal iron demand. By increasing maternal iron levels in late gestation, we were able to stimulate the expression of the gene encoding hepcidin, suggesting that the iron status of the mother is the predominant factor influencing hepcidin levels during pregnancy. Our data indicate that pregnancy-induced hepcidin suppression likely occurs because of reductions in maternal iron reserves due to increased iron requirements, which predominantly reflect stimulated erythropoiesis in mid-gestation and increased fetal iron requirements in late gestation, and that there is no need to invoke other factors, including novel pregnancy factor(s), to explain these changes.

Whole Genome Sequencing in Advanced Lung Cancer can be Performed Using Diff-Quik Cytology Smears Derived from Endobronchial Ultrasound, Transbronchial Needle Aspiration (EBUS TBNA).

INTRODUCTION: Maximising alternative sample types for genomics in advanced lung cancer is important because bronchoscopic samples may sometimes be insufficient for this purpose. Further, the clinical applications of comprehensive molecular analysis such as whole genome sequencing (WGS) are rapidly developing. Diff-Quik cytology smears from EBUS TBNA is an alternative source of DNA, but its feasibility for WGS has not been previously demonstrated. METHODS: Diff-Quik smears were collected along with research cell pellets. RESULTS: Tumour content of smears were compared to research cell pellets from 42 patients, which showed good correlation (Spearman correlation 0.85, P < 0.0001). A subset of eight smears underwent WGS, which presented similar mutation profiles to WGS of the matched cell pellet. DNA yield was predicted using a regression equation of the smears cytology features, which correctly predicted DNA yield > 1500 ng in 7 out of 8 smears. CONCLUSIONS: WGS of commonly collected Diff-Quik slides is feasible and their DNA yield can be predicted.

Development of CRISPR/Cas13a-based assays for the diagnosis of Schistosomiasis.

BACKGROUND: Schistosomiasis is a disease that significantly impacts human health in the developing world. Effective diagnostics are urgently needed for improved control of this disease. CRISPR-based technology has rapidly accelerated the development of a revolutionary and powerful diagnostics platform, resulting in the advancement of a class of ultrasensitive, specific, cost-effective and portable diagnostics, typified by applications in COVID-19/cancer diagnosis. METHODS: We developed CRISPR-based diagnostic platform SHERLOCK (Specific High-sensitivity Enzymatic Reporter unLOCKing) for the detection of Schistosoma japonicum and S. mansoni by combining recombinase polymerase amplification (RPA) with CRISPR-Cas13a detection, measured via fluorescent or colorimetric readouts. We evaluated SHERLOCK assays by using 150 faecal/serum samples collected from Schistosoma-infected ARC Swiss mice (female), and 189 human faecal/serum samples obtained from a S. japonicum-endemic area in the Philippines and a S. mansoni-endemic area in Uganda. FINDINGS: The S. japonicum SHERLOCK assay achieved 93-100% concordance with gold-standard qPCR detection across all the samples. The S. mansoni SHERLOCK assay demonstrated higher sensitivity than qPCR and was able to detect infection in mouse serum as early as 3 weeks post-infection. In human samples, S. mansoni SHERLOCK had 100% sensitivity when compared to qPCR of faecal and serum samples. INTERPRETATION: These schistosomiasis diagnostic assays demonstrate the potential of SHERLOCK/CRISPR-based diagnostics to provide highly accurate and field-friendly point-of-care tests that could provide the next generation of diagnostic and surveillance tools for parasitic neglected tropical diseases. FUNDING: Australian Infectious Diseases Research Centre seed grant (2022) and National Health and Medical Research Council (NHMRC) of Australia (APP1194462, APP2008433).

Salivary micro RNAs as biomarkers for oropharyngeal cancer.

BACKGROUND: Despite the rising incidence, particularly of the human papillomavirus (HPV)-associated fraction of oropharyngeal cancer (OPC), there are no early detection methods for OPC. Considering the close association between saliva and head and neck cancers, this study was designed to investigate salivary micro RNA (miRNAs) associated with OPC, especially focusing on HPV-positive OPC. METHODS: Saliva was collected from OPC patients at diagnosis and patients were clinically followed up ≤5 years. Salivary small RNA isolated from HPV-positive OPC patients (N = 6), and HPV-positive (N = 4) and negative controls (N = 6) were analysed by next-generation sequencing to identify dysregulated miRNAs. Discovered miRNAs were validated by quantitative PCR using two different assays in a separate cohort of patients (OPC = 91, controls = 92). The relative expression was calculated considering SNORD-96A as the normalizer. Candidate miRNAs with diagnostic and prognostic potential were evaluated by generalized logistic regression. RESULTS: A panel consisting of nine miRNAs was identified to have the best diagnostic performance to discriminate HPV-positive OPC from HPV-positive controls (AUC- validation-1 = 94.8%, validation-2 = 98%). Further, a panel consisting of six miRNAs were identified to discriminate OPC from controls regardless of the HPV status (AUC- validation-1 = 77.2%, validation-2 = 86.7%). In addition, the downregulation of hsa-miR-7-5p was significantly associated with poor overall survival of OPC patients (HR = 0.638). A panel consisting of nine miRNAs were identified for the prediction of the overall survival of the OPC patients (log-rank test-p = 0.0008). CONCLUSION: This study highlights that salivary miRNAs can play an essential role in the detection and prognostication of OPC.

The effect of vitamin D supplementation on the gut microbiome in older Australians - Results from analyses of the D-Health Trial.

Observational studies suggest a link between vitamin D and the composition of the gut microbiome, but there is little evidence from randomized controlled trials of vitamin D supplementation. We analyzed data from the D-Health Trial, a randomized, double-blind, placebo-controlled trial. We recruited 21,315 Australians aged 60-84 y and randomized them to 60,000 IU of vitamin D3 or placebo monthly for 5 y. Stool samples were collected from a sample of 835 participants (417 in the placebo and 418 in the vitamin D group) approximately 5 y after randomization. We characterized the gut microbiome using 16S rRNA gene sequencing. We used linear regression to compare alpha diversity indices (i.e. Shannon index (primary outcome), richness, inverse Simpson index), and the ratio of Firmicutes to Bacteroidetes between the two groups. We analyzed between-sample (beta) diversity (i.e. Bray Curtis distance and UniFrac index) using principal coordinate analysis and used PERMANOVA to test for significant clustering according to randomization group. We also assessed the difference in the abundance of the 20 most abundant genera between the two groups using negative binomial regression model with adjustment for multiple testing. Approximately half the participants included in this analysis were women (mean age 69.4 y). Vitamin D supplementation did not alter the Shannon diversity index (mean 3.51 versus 3.52 in the placebo and vitamin D groups, respectively, p = 0.50). Similarly, there was little difference between the groups for other alpha diversity indices, the abundance of different genera, and the Firmicutes-to-Bacteroidetes ratio. We did not observe clustering of bacterial communities according to randomization group. In conlusion, monthly doses of 60,000 IU of vitamin D supplementation for 5 y did not alter the composition of the gut microbiome in older Australians.

Vitamin D supplementation and major cardiovascular events: D-Health randomised controlled trial.

OBJECTIVE: To investigate whether supplementing older adults with monthly doses of vitamin D alters the incidence of major cardiovascular events. DESIGN: Randomised, double blind, placebo controlled trial of monthly vitamin D (the D-Health Trial). Computer generated permuted block randomisation was used to allocate treatments. SETTING: Australia from 2014 to 2020. PARTICIPANTS: 21 315 participants aged 60-84 years at enrolment. Exclusion criteria were self-reported hypercalcaemia, hyperparathyroidism, kidney stones, osteomalacia, sarcoidosis, taking >500 IU/day supplemental vitamin D, or unable to give consent because of language or cognitive impairment. INTERVENTION: 60 000 IU/month vitamin D3 (n=10 662) or placebo (n=10 653) taken orally for up to five years. 16 882 participants completed the intervention period: placebo 8270 (77.6%); vitamin D 8552 (80.2%). MAIN OUTCOME MEASURES: The main outcome for this analysis was the occurrence of a major cardiovascular event, including myocardial infarction, stroke, and coronary revascularisation, determined through linkage with administrative datasets. Each event was analysed separately as secondary outcomes. Flexible parametric survival models were used to estimate hazard ratios and 95% confidence intervals. RESULTS: 21 302 people were included in the analysis. The median intervention period was five years. 1336 participants experienced a major cardiovascular event (placebo 699 (6.6%); vitamin D 637 (6.0%)). The rate of major cardiovascular events was lower in the vitamin D group than in the placebo group (hazard ratio 0.91, 95% confidence interval 0.81 to 1.01), especially among those who were taking cardiovascular drugs at baseline (0.84, 0.74 to 0.97; P for interaction=0.12), although the P value for interaction was not significant (<0.05). Overall, the difference in standardised cause specific cumulative incidence at five years was -5.8 events per 1000 participants (95% confidence interval -12.2 to 0.5 per 1000 participants), resulting in a number needed to treat to avoid one major cardiovascular event of 172. The rate of myocardial infarction (hazard ratio 0.81, 95% confidence interval 0.67 to 0.98) and coronary revascularisation (0.89, 0.78 to 1.01) was lower in the vitamin D group, but there was no difference in the rate of stroke (0.99, 0.80 to 1.23). CONCLUSIONS: Vitamin D supplementation might reduce the incidence of major cardiovascular events, although the absolute risk difference was small and the confidence interval was consistent with a null finding. These findings could prompt further evaluation of the role of vitamin D supplementation, particularly in people taking drugs for prevention or treatment of cardiovascular disease. TRIAL REGISTRATION: ACTRN12613000743763.

Post-treatment 18F-fludeoxyglocuse-positron emission tomography in human papillomavirus-associated oropharyngeal cancer.

BACKGROUND: Human papillomavirus association has changed the landscape of treatment for oropharyngeal squamous cell carcinoma; it remains to be seen whether current post-treatment surveillance schedules are effective. OBJECTIVE: Evaluate whether post-treatment surveillance of oropharyngeal cancer through FDG-PET imaging is modified by human papillomavirus association. METHODS: A prospective cohort analysis of retrospective data was conducted for patients undergoing treatment of oropharyngeal cancer between 2016 and 2018. This study was conducted at a single large tertiary referral center in Brisbane, Australia. RESULTS: Two-hundred and twenty-four patients were recruited for the purposes of the study, 193 (86%) with HPV-associated disease. In this cohort FDG-PET had a sensitivity of 48.3%, specificity of 72.6%, positive predictive value of 23.7%, and negative predictive value of 88.8% in detecting disease recurrence. CONCLUSIONS: FDG-PET in HPV-associated oropharyngeal cancer has significantly lower positive predictive value when compared to non-HPV-associated oropharyngeal cancer. Caution should be used when interpreting positive post-treatment FDG-PET.