Apostle 技术在 cfRNA （游离RNA）领域的应用案例

Terminal modifications independent cell-free RNA sequencing enables sensitive early cancer detection and classification. Jun Wang, Jinyong Huang, Yunlong Hu, et al. Nature Communications   15, Article number: 156 (2024) 

(Note: Apostle MiniMax technology is used in this study.) 

Abstract Cell-free RNAs (cfRNAs) offer an opportunity to detect diseases from a transcriptomic perspective, however, existing techniques have fallen short in generating a comprehensive cell-free transcriptome profile. We develop a sensitive library preparation method that is robust down to 100 µl input plasma to analyze cfRNAs independent of their 5’-end modifications. We show that it outperforms adapter ligation-based method in detecting a greater number of cfRNA species. We perform transcriptome-wide characterizations in 165 lung cancer, 30 breast cancer, 37 colorectal cancer, 55 gastric cancer, 15 liver cancer, and 133 cancer-free participants and demonstrate its ability to identify transcriptomic changes occurring in early-stage tumors. We also leverage machine learning analyses on the differentially expressed cfRNA signatures and reveal their robust performance in cancer detection and classification. Our work sets the stage for in-depth study of the cfRNA repertoire and highlights the value of cfRNAs as cancer biomarkers in clinical applications.

(Methods section)  cfRNA extraction

Frozen plasma samples were thawed on ice prior to cfRNA extraction. 200 μl of plasma samples were subjected to cfRNA extraction using the Apostle MiniMax High-Efficiency cfRNA Isolation Kit (Apostle), following the manufacturer’s protocol with minor modifications.

Combining cell-free RNA with cell-free DNA in liquid biopsy for hematologic and solid tumors. Maher Albitar, Hong Zhang, Ahmad Charifa, et al. Heliyon  9 (2023) e16261; May 16, 2023 

(Note: Apostle MiniMax technology is used in this study.) 

Abstract Current use of liquid biopsy is based on cell-free DNA (cfDNA) and the evaluation of mutations or methylation pattern. However, expressed RNA can capture mutations, changes in expression levels due to methylation, and provide information on cell of origin, growth, and proliferation status. We developed an approach to isolate cell-free total nucleic acid (cfDNA) and used targeted next generation sequencing to sequence cell-free RNA (cfRNA) and cfDNA as new approach in liquid biopsy. We demonstrate that cfRNA is overall more sensitive than cfDNA in detecting mutations. We show that cfRNA is reliable in detecting fusion genes and cfDNA is reliable in detecting chromosomal gains and losses. cfRNA levels of various solid tumor biomarkers were significantly higher (P < 0.0001) in samples from solid tumors as compared with normal control. Similarly, cfRNA lymphoid markers and cfRNA myeloid markers were all higher in lymphoid and myeloid neoplasms, respectively as compared with control (P < 0.0001). Using machine learning we demonstrate cfRNA was highly predictive of diagnosis (AUC >0.98) of solid tumors, B-cell lymphoid neoplasms, T-cell lymphoid neoplasms, and myeloid neoplasms. In evaluating the host immune system, cfRNA CD4:CD8B and CD3D:CD19 ratios in normal controls were as expected (median: 5.92 and 6.87, respectively) and were significantly lower in solid tumors (P < 0.0002). This data suggests that liquid biopsy combining analysis of cfRNA with cfDNA is practical and may provide helpful information in predicting genomic abnormalities, diagnosis of neoplasms and evaluating both the tumor biology and the host response.

(Methods section)  We used Apostle MiniMax High Efficiency cfRNA/cfDNA isolation kit and followed the recommended protocol. After extraction, half of the cfDNA was treated with DNase to obtain cfRNA and the other half was used for DNA studies.