An extracellular vesicle (EV) is a nanovesicle that shuttles proteins, nucleic acids, and lipids, thereby influencing cell behavior. A recent crop of reports have shown that EVs are involved in infectious biology, influencing host immunity and playing a role in the viral life cycle. In the present work, we investigated the EV-mediated transmission of hepatitis B virus (HBV) infection. We investigated the EV-mediated transmission of HBV infection by using a HBV infectious culture system that uses primary human hepatocytes derived from humanized chimeric mice (PXB-cells). Purified EVs were isolated by ultracentrifugation. To analyze the EVs and virions, we used stimulated emission depletion microscopy. Purified EVs from HBV-infected PXB-cells were shown to contain HBV DNA and to be capable of transmitting HBV DNA to naive PXB-cells. These HBV-DNA-transmitting EVs were shown to be generated through a ceramide-triggered EV production pathway. Furthermore, we showed that these HBV-DNA-transmitting EVs were resistant to antibody neutralization; stimulated emission depletion microscopy showed that EVs lacked hepatitis B surface antigen, the target of neutralizing antibodies. These findings suggest that EVs harbor a DNA cargo capable of transmitting viral DNA into hepatocytes during HBV infection, representing an additional antibody-neutralization-resistant route of HBV infection.

“Liquid biopsy” is an efficient diagnostic tool used to analyse biomaterials in human body fluids, such as blood, saliva, breast milk, and urine. Various biomaterials derived from a tumour and its microenvironment are released into such body fluids and contain important information for cancer diagnosis. Biomaterial detection can provide “real-time” information about individual tumours, is non-invasive, and is more repeatable than conventional histological analysis. Therefore, over the past two decades, liquid biopsy has been considered an attractive diagnostic tool for malignant tumours. Although biomarkers for oral cancer have not yet been adopted in clinical practice, many molecular candidates have been investigated for liquid biopsies in oral cancer diagnosis, such as the proteome, metabolome, microRNAome, extracellular vesicles, cell-free DNAs, and circulating tumour cells. This review will present recent advances and challenges in liquid biopsy for oral cancer diagnosis.

Cancer‐associated fibroblasts (CAFs) are the major component of the tumor stroma. They mediate various attributes of tumor cells, such as cell growth, migration, invasion, angiogenesis, metabolic reprogramming, apoptosis, immune regulation, and extracellular matrix reconstitution, all related to cancer progression and treatment resistance. Although many researchers have recognized CAF heterogeneity, recent technological advances have emphasized the functional and phenotypic diversity of CAFs in cancer progression. Why are these CAF subtypes generated within tumor tissues? And how do cancer cells dictate such heterogeneous subtypes of CAFs? This review will highlight the CAF subtypes within the tumor microenvironment and their role in tumor progression. CAF subtype induction by extracellular vesicles (EVs) and their significance, which we reported previously, is also discussed.

Ribosomes are responsible for the protein synthesis that maintains cellular homeostasis and is required for the rapid cellular division of cancer cells. However, the role of ribosome biogenesis mediators in the malignant behavior of tongue squamous cell carcinoma (TSCC) is unknown. In this study, we found that the expression of RIOK2, a key enzyme involved in the maturation steps of the pre-40S ribosomal complex, was significantly associated with poorer overall survival in patients with TSCC. Further, multivariate analysis revealed that RIOK2 is an independent prognostic factor (hazard ratio, 3.53; 95% confidence interval, 1.19–10.91). Inhibition of RIOK2 expression by siRNA decreased cell growth and S6 ribosomal protein expression in oral squamous cell carcinoma cell lines. RIOK2 knockdown also led to a significant decrease in the protein synthesis in cancer cells. RIOK2 has potential application as a novel therapeutic target for TSCC treatment.

Cancer-associated fibroblasts (CAFs) play a pivotal role in cancer progression by enhancing extracellular matrix remodeling, cancer cell invasion, and mediating inflammatory environment. Therefore, CAFs have become targets for cancer therapies. However, a major hurdle to developing such therapies are the challenges of studying CAFs in culture - specifically it remains unclear which CAF functions and phenotypes are sustained during in vitro culture and the signaling pathways driving distinct CAF phenotypes. We investigated whether CAF phenotypes and functions are sustained or changed during 2D CAF culture under in vitro systems, our analysis included early passage, late passage, and immortalized cultures, as well as CAFs in coculture with cancer cell lines. Transcriptomic analysis revealed that inflammatory-related gene expression programs were suppressed in late passage and immortalized CAFs compared with early passage equivalents. Direct co-culture with lung cancer cells can revert the inflammatory gene expressions in immortalized CAFs such that they partially resemble early passage CAFs. In contrast, indirect coculture with lung cancer cells - in which CAFs could not form cell-cell contacts with cancer cells - had a less significant effect on immortalized CAF inflammatory gene expression but could induce TGFβ regulated genes. Interestingly, inhibition of the TGFβ signaling pathway potentiated the inflammatory gene expression network induced by direct co-culture, suggesting that the signaling mediated by direct cellular interaction is antagonistic to the TGFβ signaling pathway. These data further our understanding of cancer cell – CAF crosstalk and will help to develop in vitro assays that more closely maintain the state of CAFs observed in tissue. Citation Format: Yutaka Naito, Robert Hynds, David Novo, Probir Chakravarty, Gavin Kelly, Charles Swanton, Kazufumi Honda, Erik Sahai, TRACERx Consortium. Tracking the transcriptome of lung cancer-associated fibroblasts within the TRACERx lung study from patient to culture models reveals phenotypic plasticity and instructive cues from cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 674.