TOTEM: a multi-cancer detection and localization approach using circulating tumor DNA methylation markers.

BACKGROUND: Detection of cancer and identification of tumor origin at an early stage improve the survival and prognosis of patients. Herein, we proposed a plasma cfDNA-based approach called TOTEM to detect and trace the cancer signal origin (CSO) through methylation markers. METHODS: We performed enzymatic conversion-based targeted methylation sequencing on plasma cfDNA samples collected from a clinical cohort of 500 healthy controls and 733 cancer patients with seven types of cancer (breast, colorectum, esophagus, stomach, liver, lung, and pancreas) and randomly divided these samples into a training cohort and a testing cohort. An independent validation cohort of 143 healthy controls, 79 liver cancer patients and 100 stomach cancer patients were recruited to validate the generalizability of our approach. RESULTS: A total of 57 multi-cancer diagnostic markers and 873 CSO markers were selected for model development. The binary diagnostic model achieved an area under the curve (AUC) of 0.907, 0.908 and 0.868 in the training, testing and independent validation cohorts, respectively. With a training specificity of 98%, the specificities in the testing and independent validation cohorts were 100% and 98.6%, respectively. Overall sensitivity across all cancer stages was 65.5%, 67.3% and 55.9% in the training, testing and independent validation cohorts, respectively. Early-stage (I and II) sensitivity was 50.3% and 45.7% in the training and testing cohorts, respectively. For cancer patients correctly identified by the binary classifier, the top 1 and top 2 CSO accuracies were 77.7% and 86.5% in the testing cohort (n = 148) and 76.0% and 84.0% in the independent validation cohort (n = 100). Notably, performance was maintained with only 21 diagnostic and 214 CSO markers, achieving a training AUC of 0.865, a testing AUC of 0.866, and an integrated top 2 accuracy of 83.1% in the testing cohort. CONCLUSIONS: TOTEM demonstrates promising potential for accurate multi-cancer detection and localization by profiling plasma methylation markers. The real-world clinical performance of our approach needs to be investigated in a much larger prospective cohort.

Genome-Scale Multimodal Analysis of Cell-Free DNA Whole-Methylome Sequencing for Noninvasive Esophageal Cancer Detection.

PURPOSE: Simultaneous profiling of cell-free DNA (cfDNA) methylation and fragmentation features to improve the performance of cfDNA-based cancer detection is technically challenging. We developed a method to comprehensively analyze multimodal cfDNA genomic features for more sensitive esophageal squamous cell carcinoma (ESCC) detection. MATERIALS AND METHODS: Enzymatic conversion-mediated whole-methylome sequencing was applied to plasma cfDNA samples extracted from 168 patients with ESCC and 251 noncancer controls. ESCC characteristic cfDNA methylation, fragmentation, and copy number signatures were analyzed both across the genome and at accessible cis-regulatory DNA elements. To distinguish ESCC from noncancer samples, a first-layer classifier was developed for each feature type, the prediction results of which were incorporated to construct the second-layer ensemble model. RESULTS: ESCC plasma genome displayed global hypomethylation, altered fragmentation size, and chromosomal copy number alteration. Methylation and fragmentation changes at cancer tissue-specific accessible cis-regulatory DNA elements were also observed in ESCC plasma. By integrating multimodal genomic features for ESCC detection, the ensemble model showed improved performance over individual modalities. In the training cohort with a specificity of 99.2%, the detection sensitivity was 81.0% for all stages and 70.0% for stage 0-II. Consistent performance was observed in the test cohort with a specificity of 98.4%, an all-stage sensitivity of 79.8%, and a stage 0-II sensitivity of 69.0%. The performance of the classifier was associated with the disease stage, irrespective of clinical covariates. CONCLUSION: This study comprehensively profiles the epigenomic landscape of ESCC plasma and provides a novel noninvasive and sensitive ESCC detection approach with genome-scale multimodal analysis.

New records of non-indigenous species from the eastern Mediterranean Sea (Crustacea, Mollusca), with a revision of genus Isognomon (Mollusca: Bivalvia).

We report new data on non-indigenous invertebrates from the Mediterranean Sea (four ostracods and 20 molluscs), including five new records for the basin: the ostracods Neomonoceratina iniqua, Neomonoceratina aff. mediterranea, Neomonoceratina cf. entomon, Loxoconcha cf. gisellae (Arthropoda: Crustacea)-the first records of non-indigenous ostracods in the Mediterranean-and the bivalve Striarca aff. symmetrica (Mollusca). Additionally, we report for the first time Electroma vexillum from Israel, and Euthymella colzumensis, Joculator problematicus, Hemiliostraca clandestina, Pyrgulina nana, Pyrgulina microtuber, Turbonilla cangeyrani, Musculus aff. viridulus and Isognomon bicolor from Cyprus. We also report the second record of Fossarus sp. and of Cerithiopsis sp. cf. pulvis in the Mediterranean Sea, the first live collected specimens of Oscilla galilae from Cyprus and the northernmost record of Gari pallida in Israel (and the Mediterranean). Moreover, we report the earliest records of Rugalucina angela, Ervilia scaliola and Alveinus miliaceus in the Mediterranean Sea, backdating their first occurrence in the basin by 3, 5 and 7 years, respectively. We provide new data on the presence of Spondylus nicobaricus and Nudiscintilla aff. glabra in Israel. Finally, yet importantly, we use both morphological and molecular approaches to revise the systematics of the non-indigenous genus Isognomon in the Mediterranean Sea, showing that two species currently co-occur in the basin: the Caribbean I. bicolor, distributed in the central and eastern Mediterranean, and the Indo-Pacific I. aff. legumen, at present reported only from the eastern Mediterranean and whose identity requires a more in-depth taxonomic study. Our work shows the need of taxonomic expertise and investigation, the necessity to avoid the unfounded sense of confidence given by names in closed nomenclature when the NIS belong to taxa that have not enjoyed ample taxonomic work, and the necessity to continue collecting samples-rather than relying on visual censuses and bio-blitzes-to enable accurate detection of non-indigenous species.