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While combination strategy mitigates tumor resistance, enhances therapeutic efficacy, and reduces side effects compared to monotherapies, the precise coordination of treatment sequences and timing remains underexplored, particularly in the nanomaterial\u2010based therapies. In this study, this gap is addressed by investigating the interaction between photodynamic therapy (PDT) and nitric oxide (NO) therapy, with a specific focus on the sequence in which these therapies are administered. An upconversion nanoplatform is designed with orthogonal dual\u2010channel fluorescence properties, allowing for non\u2010interfering NO release and PDT, which enables systematic exploration of the effects of treatment sequence on therapy efficacy. The results demonstrate that administering NO therapy prior to PDT yields superior antitumor effects compared to other sequences. 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