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AAV vector is the most popular viral gene delivery system in clinical delivery systems with unique and multiple advantages, such as tissue tropism, transduction specificity, long\u2010lasting gene expression, low immune responses, and without host chromosome incorporation. Till now, four AAV\u2010based gene therapy drugs have already been approved by the US Food and Drug Administration (FDA) or European Medicines Agency (EMA). Despite the success of AAV vectors, there are still some remaining challenges that limit further usage, such as poor packaging capacity, low organ specificity, pre\u2010existing humoral immunity, and vector dose\u2010dependent toxicity. In the present review, we address the different approaches to optimize AAV vector delivery system with a focus on capsid engineering, packaging capacity, and immune response at the clinical level. The review further investigates the potential of manipulating AAV vectors in preclinical applications and clinical translation, which emphasizes the challenges and prospects in viral vector selection, drug delivery strategies, immune reactions in cancer, neurodegenerative disease, retinal disease, SARS\u2010CoV\u20102, and monkeypox. 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