Immune checkpoint inhibitors have transformed cancer treatment, but many patients fail to achieve durable benefit because malignant, immune and stromal cells maintain an immunosuppressive tumor immune microenvironment. Extracellular vesicles (EVs), including small EVs often termed exosomes, mediate intercellular communication and can transfer circular RNAs (circRNAs) between defined donor and recipient cells. The clinical relevance of EV‑associated circRNAs extends from understanding basic transfer mechanisms to exploring biomarker and therapeutic potential in oncology.
The pathway linking host cell biology to recipient phenotype includes circRNA biogenesis in donor cells, selection and entry into EV populations, and eventual delivery to recipient cells. Not all reported effects attributed to circRNAs derive from EV‑mediated transfer; experimental contexts such as incomplete transfer experiments, intrinsic tumor circRNA activity and engineered RNA platforms differ from direct EV transfer and must be distinguished in study design and interpretation.
Once delivered to recipient cells, circRNAs can exert multiple intracellular activities. These include regulation of microRNA availability, assembly of RNA‑binding protein complexes, modulation of protein or RNA stability and production of functional peptides. Such mechanisms provide plausible routes by which transferred circRNAs alter cellular behavior and immune phenotypes within the tumor microenvironment.
Direct EV‑mediated circRNA transfer has been linked to several immune cell populations implicated in tumor immunity and immune evasion. Reported recipient cell types include tumor‑associated macrophages (TAMs), myeloid‑derived suppressor cells (MDSCs), natural killer (NK) cells, CD8+ T cells and regulatory T cells (Tregs). The biological impact of transfer is context dependent: donor cell state, recipient identity, anatomical tissue site, EV subpopulation and delivered dose can all alter the resulting phenotype.
Cancer‑associated fibroblasts (CAFs) further link EV‑associated circRNAs to the physical and functional tumor stroma. Through effects on matrix remodeling, CAFs can influence immune‑cell access and contribute to treatment tolerance. Thus, stromal interactions represent an important axis connecting EV cargo to immune modulation and therapeutic resistance.
Biological support for EV‑circRNA transfer is classified along an evidence scale from E0 to E3. EV study methods should be considered separately and must address multiple technical elements: clear definition of EV source, robust separation and characterization of EV populations, demonstration of RNA protection within EVs, appropriate uptake controls to confirm transfer, and quantitative assessment of delivered dose. These methodological standards are essential to distinguish systemic or circulating associations from molecules that are truly transferred and functionally active in recipient cells.
Successful translation of EV‑associated circRNAs as biomarkers or therapeutics will require several specific advances. Key needs reported include identification of full‑length circRNA sequences, absolute measurement of circRNA copies in EVs and in recipient cells, and spatial localization within tissues. In addition, prospective clinical cohorts and repeated‑dose safety testing are necessary to move from associative findings toward therapeutic development. These requirements underscore the difference between detecting circulating associations and establishing transferred, functional molecules suitable for clinical use.
EV‑mediated circRNA biology presents both biomarker and interventional opportunities. Clarifying donor and recipient determinants, EV subpopulation effects and dose–response relationships will inform therapeutic design. Meanwhile, rigorous experimental frameworks—distinguishing direct EV transfer from other sources of circRNA activity—are essential to support development of EV‑based diagnostics or therapeutics and to design prospective treatment cohorts for clinical evaluation.
Exosomal circRNAs offer a mechanistically rich means of shaping the tumor immune microenvironment, with reported links to multiple immune cell types and stromal elements such as CAFs. However, translation into reliable biomarkers or therapeutic agents requires stringent experimental evidence, standardized EV methodologies and clinical validation including safety testing. Addressing full‑length circRNA identification, absolute quantification, spatial mapping and prospective clinical studies will be critical to move the field beyond associations to reproducible, transferable molecular interventions.