The Research Domain Criteria (RDoC) framework offers a dimensional and transdiagnostic approach to understanding complex neuropsychiatric and neurological disorders, facilitating the development of novel therapeutics. However, the integration of cellular and molecular brain processes into the RDoC framework is hindered by the lack of adequate biomarkers. This review explores the potential of brain-derived extracellular vesicles (BDEVs) isolated from biofluids as a source of non-invasive mechanistic biomarkers in RDoC-based drug discovery. We provide an overview of BDEVs, including their classification, biological functions, and current methodologies for isolation and characterization. We then discuss studies that have investigated BDEV cargo as mechanistic biomarkers in CNS drug studies, focusing on target engagement, treatment response, and toxicity. Additionally, we address important considerations and open questions regarding the characterization and validation of BDEVs in neuroscience research. Special emphasis is placed on the use of miRNA cargo within BDEVs as molecular readouts, highlighting their stability, regulatory roles, and potential to reflect dynamic changes in brain function. Finally, the review proposes strategies to further investigate the use of BDEV miRNA cargo as molecular readouts, underscoring its ability to provide insights into the molecular mechanisms underlying drug effects and their relevance to CNS drug discovery within the RDoC framework. Despite existing methodological and conceptual challenges, BDEVs represent a promising tool for advancing RDoC-based drug discovery.