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.

Multimodal analysis of genome-wide methylation, copy number aberrations, and end motif signatures enhances detection of early-stage breast cancer.

Introduction: Breast cancer causes the most cancer-related death in women and is the costliest cancer in the US regarding medical service and prescription drug expenses. Breast cancer screening is recommended by health authorities in the US, but current screening efforts are often compromised by high false positive rates. Liquid biopsy based on circulating tumor DNA (ctDNA) has emerged as a potential approach to screen for cancer. However, the detection of breast cancer, particularly in early stages, is challenging due to the low amount of ctDNA and heterogeneity of molecular subtypes. Methods: Here, we employed a multimodal approach, namely Screen for the Presence of Tumor by DNA Methylation and Size (SPOT-MAS), to simultaneously analyze multiple signatures of cell free DNA (cfDNA) in plasma samples of 239 nonmetastatic breast cancer patients and 278 healthy subjects. Results: We identified distinct profiles of genome-wide methylation changes (GWM), copy number alterations (CNA), and 4-nucleotide oligomer (4-mer) end motifs (EM) in cfDNA of breast cancer patients. We further used all three signatures to construct a multi-featured machine learning model and showed that the combination model outperformed base models built from individual features, achieving an AUC of 0.91 (95% CI: 0.87-0.95), a sensitivity of 65% at 96% specificity. Discussion: Our findings showed that a multimodal liquid biopsy assay based on analysis of cfDNA methylation, CNA and EM could enhance the accuracy for the detection of early- stage breast cancer.

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.

Pulmonary carcinosarcoma initially presenting as invasive aspergillosis: a case report of previously unreported combination.

Carcinosarcoma of the lung is a malignant tumor composed of a mixture of carcinoma and sarcoma elements. The carcinomatous component is most commonly squamous followed by adenocarcinoma. The sarcomatous component commonly comprises the bulk of the tumor and shows poorly differentiated spindle cell features. Foci of differentiated sarcomatous elements such as chondrosarcoma and osteosarcoma may be seen. Aspergillus pneumonia is the most common form of invasive aspergillosis and occurs mainly in patients with malignancy, immunocompromising or debilitating diseases. Patients with Aspergillus pneumonia present with fever, cough, chest pain and occasionally hemoptysis. Tissue examination is the most reliable method for diagnosis, and mortality rate is high.We describe a case of primary carcinosarcoma of the lung concurrently occurring with invasive pulmonary aspergillosis in a 66-year old patient.

Adiponectin and glucose production in patients infected with Plasmodium falciparum.

Infections are often complicated by an increase in glucose production due to stimulation of the secretion of glucose counter-regulatory hormones and cytokines. Adiponectin, a fat-derived hormone with insulin-sensitizing properties, could play a regulatory role in the degree of stimulation of glucose production by the infectious agent. Therefore, we investigated the possible correlation between glucose production and plasma adiponectin levels in 25 subjects: 7 patients with cerebral malaria, 6 with uncomplicated malaria, and 12 matched controls. Glucose production was significantly higher in patients with malaria compared to healthy controls (P < .001). Adiponectin levels were not different between the patients with malaria and the control group. However, patients with cerebral malaria had significantly higher values for adiponectin than the patients with uncomplicated malaria (P < .005). Glucose production and gluconeogenesis were positively correlated to plasma adiponectin in the patients (r = 0.835, P < .001 and r = 0.846, P < .001, respectively), whereas these correlations were absent in the controls (r = -0.329, NS and r = -0.028, NS, respectively). In conclusion, adiponectin levels were not different between patients with malaria and their matched controls. However, patients infected with Plasmodium falciparum who have higher glucose production also have higher adiponectin levels. In healthy subjects such a correlation was not found. As adiponectin is known to inhibit glucose production, stimulation of adiponectin secretion during infection could be intended to restrain the glucose production stimulating properties of hormones and cytokines secreted during infection.

FFAs are not involved in regulation of gluconeogenesis and glycogenolysis in adults with uncomplicated P. falciparum malaria.

In normal subjects, elevation of plasma free fatty acid (FFA) levels stimulates gluconeogenesis (GNG) and inhibits glycogenolysis (GLY). In adults with uncomplicated Plasmodium falciparum malaria, GNG is increased and GLY decreased. To test the hypothesis that FFAs are regulators of GNG and GLY in uncomplicated falciparum malaria, we investigated the effect of inhibition of lipolysis by acipimox in 12 patients with uncomplicated falciparum malaria. Six of them were given acipimox, and six served as controls. Also as controls, six matched healthy subjects were studied on two occasions with and without acipimox. After 16 h of fasting, glucose production and GNG were significantly higher in the malaria patients compared with the healthy controls (P = 0.003 and < 0.0001, respectively), whereas GLY was significantly lower (P < 0.001), together with elevated plasma concentrations of cortisol and glucagon. During the study, glucose production in patients declined over time (P < 0.0001), without a statistically significant difference between the acipimox-treated and untreated patients. In controls, however, with acipimox the decline was less outspoken compared with nontreated controls (P = 0.005). GNG was unchanged over time in patients as well as in healthy controls, and no influence of acipimox was found. In patients, GLY declined over time (P < 0.001), without a difference between acipimox-treated and untreated patients. In contrast, in controls treated with acipimox, no change over time was found, which was statistically different from the decline in untreated controls (P = 0.002). In conclusion, in falciparum malaria, FFAs are not involved in regulation of glucose production, nor of GNG or GLY.