IABS/DCVMN webinar on next generation sequencing.

Next Generation Sequencing (NGS) is a new technology that could overcome some of the limitations of the current viral testing methods for demonstrating the absence of adventitious agents in biologics. This report is for the webinar that was organized by the International Alliance for Biological Standardization (IABS) and the Developing Countries Vaccine Manufacturers Network (DCVMN), held on July 20, 2022, as an introduction to the technical and bioinformatics concepts of NGS and to some of the strengths and limitations of using the technology for those working in vaccine production or development. The current state of scientific knowledge and readiness of NGS to replace or supplement the current viral tests was further discussed in the 3rd Conference on NGS for Adventitious Virus Detection in Biologics for Humans and Animals that was held in Rockville, Maryland, USA, on September 27-28, 2022. The application of NGS to supplement or replace current in vivo and in vitro assays in adventitious virus testing during vaccine production is promising; however, assay performance (sensitivity, specificity, and reproducibility) needs to be demonstrated, which may include laboratory and bioinformatics work. Efforts from regulatory authorities, industry, and researchers are ongoing to facilitate validation and establishment of NGS as a new method for virus detection.

Surfaceome interrogation using an RNA-seq approach highlights leukemia initiating cell biomarkers in an LMO2 T cell transgenic model.

The surfaceome is critical because surface proteins provide a gateway for internal signals and transfer of molecules into cells, and surfaceome differences can influence therapy response. We have used a surfaceome analysis method, based on comparing RNA-seq data between normal and abnormal cells (Surfaceome DataBase Mining or Surfaceome DBM), to identify sets of upregulated cell surface protein mRNAs in an LMO2-mediated T-ALL mouse model and corroborated by protein detection using antibodies. In this model the leukemia initiating cells (LICs) comprise pre-leukaemic, differentiation inhibited thymocytes allowing us to provide a profile of the LIC surfaceome in which GPR56, CD53 and CD59a are co-expressed with CD25. Implementation of cell surface interaction assays demonstrates fluid interaction of surface proteins and CD25 is only internalized when co-localized with other proteins. The Surfaceome DBM approach to analyse cancer cell surfaceomes is a way to find targetable surface biomarkers for clinical conditions where RNA-seq data from normal and abnormal cell are available.