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Advances in single-cell and spatial multi-omics technologies enable precise quantification of tumor heterogeneity, supporting detailed investigation of how heterogeneity contributes to chemoresistance and informing the development of personalized therapeutic strategies. In this review, we summarize the evolutionary dynamics underlying the emergence of tumor drug resistance and examine the molecular mechanisms responsible for failure of targeted therapies. We highlight how advances in single-cell and spatial multi-omics have significantly improved our ability to elucidate these processes. We further suggest that addressing tumor drug resistance may require a shift from static, single-target approaches toward dynamic, biology-informed personalized strategies. 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