Multimodal analysis of methylomics and fragmentomics in plasma cell-free DNA for multi-cancer early detection and localization.

Despite their promise, circulating tumor DNA (ctDNA)-based assays for multi-cancer early detection face challenges in test performance, due mostly to the limited abundance of ctDNA and its inherent variability. To address these challenges, published assays to date demanded a very high-depth sequencing, resulting in an elevated price of test. Herein, we developed a multimodal assay called SPOT-MAS (screening for the presence of tumor by methylation and size) to simultaneously profile methylomics, fragmentomics, copy number, and end motifs in a single workflow using targeted and shallow genome-wide sequencing (~0.55×) of cell-free DNA. We applied SPOT-MAS to 738 non-metastatic patients with breast, colorectal, gastric, lung, and liver cancer, and 1550 healthy controls. We then employed machine learning to extract multiple cancer and tissue-specific signatures for detecting and locating cancer. SPOT-MAS successfully detected the five cancer types with a sensitivity of 72.4% at 97.0% specificity. The sensitivities for detecting early-stage cancers were 73.9% and 62.3% for stages I and II, respectively, increasing to 88.3% for non-metastatic stage IIIA. For tumor-of-origin, our assay achieved an accuracy of 0.7. Our study demonstrates comparable performance to other ctDNA-based assays while requiring significantly lower sequencing depth, making it economically feasible for population-wide screening.

Fragment length profiles of cancer mutations enhance detection of circulating tumor DNA in patients with early-stage hepatocellular carcinoma.

BACKGROUND: Late detection of hepatocellular carcinoma (HCC) results in an overall 5-year survival rate of less than 16%. Liquid biopsy (LB) assays based on detecting circulating tumor DNA (ctDNA) might provide an opportunity to detect HCC early noninvasively. Increasing evidence indicates that ctDNA detection using mutation-based assays is significantly challenged by the abundance of white blood cell-derived mutations, non-tumor tissue-derived somatic mutations in plasma, and the mutational tumor heterogeneity. METHODS: Here, we employed concurrent analysis of cancer-related mutations, and their fragment length profiles to differentiate mutations from different sources. To distinguish persons with HCC (PwHCC) from healthy participants, we built a classification model using three fragmentomic features of ctDNA through deep sequencing of thirteen genes associated with HCC. RESULTS: Our model achieved an area under the curve (AUC) of 0.88, a sensitivity of 89%, and a specificity of 82% in the discovery cohort consisting of 55 PwHCC and 55 healthy participants. In an independent validation cohort of 54 PwHCC and 53 healthy participants, the established model achieved comparable classification performance with an AUC of 0.86 and yielded a sensitivity and specificity of 81%. CONCLUSIONS: Our study provides a rationale for subsequent clinical evaluation of our assay performance in a large-scale prospective study.

Clinical validation of a ctDNA-Based Assay for Multi-Cancer Detection: An Interim Report from a Vietnamese Longitudinal Prospective Cohort Study of 2795 Participants.

The SPOT-MAS assay "Screening for the Presence Of Tumor by Methylation And Size" detects the five most common cancers in Vietnam by evaluating circulating tumor DNA in the blood. Here, we validated its performance in a prospective multi-center clinical trial, K-DETEK. Our analysis of 2795 participants from 14 sites across Vietnam demonstrates its ability to detect cancers in asymptomatic individuals with a positive predictive value of 60%, with 83.3% accuracy in detecting tumor location. We present a case report to support further using SPOT-MAS as a complementary method to achieve early cancer detection and provide the opportunity for early treatment.

Circulating DNA methylation profile improves the accuracy of serum biomarkers for the detection of nonmetastatic hepatocellular carcinoma.

Aim: This study exploited hepatocellular carcinoma (HCC)-specific circulating DNA methylation profiles to improve the accuracy of a current screening assay for HCC patients in high-risk populations. Methods: Differentially methylated regions in cell-free DNA between 58 nonmetastatic HCC and 121 high-risk patients with liver cirrhosis or chronic hepatitis were identified and used to train machine learning classifiers. Results: The model could distinguish HCC from high-risk non-HCC patients in a validation cohort, with an area under the curve of 0.84. Combining these markers with the three serum biomarkers (AFP, lectin-reactive AFP, des-γ-carboxy prothrombin) in a commercial test, µTASWako®, achieved an area under the curve of 0.87 and sensitivity of 68.8% at 95.8% specificity. Conclusion: HCC-specific circulating DNA methylation markers may be added to the available assay to improve the early detection of HCC.

The early detection of liver cancer in high-risk populations can help people with the disease have a higher chance of survival and better quality of life. However, this is still a healthcare challenge. Current commercial blood tests measuring protein signatures in the blood have low accuracy due to increased levels of these proteins being detected in both liver cancer patients and patients with chronic liver diseases. In this study, we identified a set of signatures in DNA released by cancer cells into the bloodstream and used them as biomarkers to distinguish liver cancer patients from high-risk patients. We also demonstrated that adding those signatures to a commercial blood test currently used in clinics could improve the accuracy in detecting liver cancer patients.

Pathogenic Variant Profile of Hereditary Cancer Syndromes in a Vietnamese Cohort.

BACKGROUND: Hereditary cancer syndromes (HCS) are responsible for 5-10% of cancer cases. Genetic testing to identify pathogenic variants associated with cancer predisposition has not been routinely available in Vietnam. Consequently, the prevalence and genetic landscape of HCS remain unknown. METHODS: 1165 Vietnamese individuals enrolled in genetic testing at our laboratory in 2020. We performed analysis of germline mutations in 17 high- and moderate- penetrance genes associated with HCS by next generation sequencing. RESULTS: A total of 41 pathogenic variants in 11 genes were detected in 3.2% individuals. The carrier frequency was 4.2% in people with family or personal history of cancer and 2.6% in those without history. The percentage of mutation carriers for hereditary colorectal cancer syndromes was 1.3% and for hereditary breast and ovarian cancer syndrome was 1.6%. BRCA1 and BRCA2 mutations were the most prevalent with the positive rate of 1.3% in the general cohort and 5.1% in breast or ovarian cancer patients. Most of BRCA1 mutations located at the BRCA C-terminus domains and the top recurrent mutation was NM_007294.3:c.5251C>T (p.Arg1751Ter). One novel variant NM_000038.6(APC):c.6665C>A (p.Pro2222His) was found in a breast cancer patient with a strong family history of cancer. A case study of hereditary cancer syndrome was illustrated to highlight the importance of genetic testing. CONCLUSION: This is the first largest analysis of carrier frequency and mutation spectrum of HCS in Vietnam. The findings demonstrate the clinical significance of multigene panel testing to identify carriers and their at-risk relatives for better cancer surveillance and management strategies.

Probiotic Beverage From Pineapple Juice Fermented With Lactobacillus and Bifidobacterium Strains.

Pineapple is an economically significant plant and the third most important fruit crop in the tropical and subtropical regions of the world. In this study, fermentation of pineapple juice with probiotic bacteria Lactobacillus and Bifidobacterium strains as well as changes of some properties in the beverage during storage were investigated. All tested strains exhibited good growth properties on pineapple juice without supplementation of any nutrient compounds. After 24 h fermentation, the cell counts of lactobacilli passed the level of 5*109 cfu/ml, while the cell number of bifidobacteria reached a level of 109 cfu/ml. The highest volumetric productivity (3.5*108 cfu/ml*h) was observed in L. plantarum 299V. The ratios of lactic acids to acetic acids in the cases of L. plantarum 299V and L. acidophilus La5 were 5.37 and 9.91, respectively. In the case of B. lactis Bb-12, the concentrations of lactic acid and acetic acid were 6 mM and 23 mM in natural juices, and 15 and 21 mM in the case of supplementation with prebiotics at the 16th h of fermentation, respectively. Additionally, supplementation with prebiotics at the initiation of fermentation resulted 7 mM lactic acid and 23 mM acetic acid at the end of fermentation. Fructose was the most preferred sugar for both lactobacilli and bifidobacteria. Both total phenolic content and antioxidant capacity increased slightly during fermentation and dropped during the storage period. The microbial population did not change significantly during the first month of storage. After the storage period (2 months), the probiotic bacteria lost about 0.11 log cfu/ml viability after treatment with 0.3% pepsin for 135 min, and a further 0.1 log cfu/ml after treatment with 0.6% bile salts. These values were 10 times higher than data from the fresh fermented pineapple juice. Our results are very promising and may serve as a good base for developing probiotic pineapple juice.

Enhancing the photoluminescence intensity of conjugated polycationic polymers by using quantum dots as antiaggregation reagents.

The aggregation in conjugated polyelectrolytes (CPs) can be effectively reduced by the formation of CP/nanoparticle assemblies. The photophysical properties of various nanoassemblies were studied by means of UV-visible and fluorescence spectroscopy in solution and as thin films. The dissociation of the polymer chains is caused by favorable electrostatic interactions between the cationic substituents of the CPs and the anionic charges present on the surface of the nanoparticles. Such an efficient displacement of pi-stacking by competitive positive interactions constitutes the first example of positive aggregation modulation.

Selection for aluminium tolerance in embryogenic lines of Citrus sinensis L. and C. aurantium L.

The Citrus cultivars 'Amargo' and 'Pineapple' produce embryogenic cultures on two types of sugars, galactose and lactose. Two percent lactose is optimal for both cultivars; embryogenic callus is produced 8-10 weeks after inoculation. Whereas 2% galactose is optimal for 'Amargo', it is 3% for 'Pineapple'. Three types of embryos are recognized: whitish, greenish and heart shaped. In 8 to 10 week old cultures we do also observe a black spotted type of embryos. A high percentage of plant conversion for both cultivars is observed on a medium containing 30 mg/l of adenine and 2 mg/l of GA3 for 'Amargo'; for 'Pineapple' it is necessary to add BA. We tested two techniques for selection of Al-tolerant lines: one on stagnant liquid medium, and another on shaken liquid medium. The second technique gives a more serious selection pressure; there are almost no surviving calli. In 'Amargo' more calli survived than in 'Pineapple'. Selected calli had to be subcultured nine times to determine their stability. Four callus types of the two cultivars grew normally on selective media containing 0, 400, 1000 or 2000 mumol of aluminium.