Increasing evidence suggests that cancer arises from cells that are capable of initiating and sustaining neoplastic tissue growth, termed cancer stem cells (CSCs). Of central scientific and clinical relevance, cells with CSC properties are enriched for chemo- and radiation resistance and therefore may represent a population of cells that must be therapeutically targeted to prevent cancer recurrence/relapse 1. Human CSCs were first isolated in neoplastic hematopoietic tissue that manifests leukemias such as adult acute myeloid leukemia (AML) 2. AML stem cells represent a benchmark model of human CSC biology, ultimately motivating foundational studies leading to the identification of CSCs from solid tumours such as breast and colon 3. Independent of tissue type, a consistent feature of CSCs is their uncontrolled self-renewal capacity and differentiation blockade that have been commonly related to aberrant activation of pro-oncogenic events such as dysregulation of CBP/p300 transcriptional regulation involving -catenin 4. However, the transcriptional networks involving CBP/p300/-catenin complex have been shown to be equally critical to maintain normal stem cell (SCs) self-renewal for tissue homeostasis and regeneration 5. Here, we identify Sam68 as a distinct target that affords the ability to uniquely regulate CBP mediated transcription in human CSCs. Using a small molecule that targets Sam68, we reveal that shifting its affinity for CBP disrupts CBP/-catenin complexes, leading to immediate changes in histone H3 (K14 and K18) acetylation. Chemical targeting of Sam68 induced global changes in transcriptional programs of patient AML cells involving apoptosis and differentiation and was able to uniquely reduce neoplastic self-renewal of human CSCs in an in vivo model of patient specific acute myeloid leukemia (AML). Our study establishes an approach whereby the CBP/-catenin transcriptome can be uniquely targeted via Sam68 based vulnerability of CSCs that impacts neoplastic differentiation and self-renewal.