p97/VCP, an evolutionarily concerned ATPase, partakes in multiple cellular proteostatic processes, including the endoplasmic reticulum (ER)-associated protein degradation (ERAD). Elevated expression of p97 is common in many cancers and is often associated with poor survival. Here we report that the levels of p97 positively correlated with the histological grade, tumor size, and lymph node metastasis in breast cancers. We further examined p97 expression in the stem-like cancer cells or cancer stem cells (CSCs), a cell population that purportedly underscores cancer initiation, therapeutic resistance, and recurrence. We found that p97 was consistently at a higher level in the CD44+/CD24-, ALDH+, or PKH26+ CSC populations than the respective non-CSC populations in human breast cancer tissues and cancer cell lines and p97 expression also positively correlated with that of SOX2, another CSC marker. To assess the role of p97 in breast cancers, cancer proliferation, mammosphere, and orthotopic growth were analyzed. Similarly as p97 depletion, two pharmacological inhibitors, which targets the ER-associated p97 or globally inhibits p97's ATPase activity, markedly reduced cancer growth and the CSC population. Importantly, depletion or inhibition of p97 greatly suppressed the proliferation of the ALDH+ CSCs and the CSC-enriched mammospheres, while exhibiting much less or insignificant inhibitory effects on the non-CSC cancer cells. Comparable phenotypes produced by blocking ERAD suggest that ER proteostasis is essential for the CSC integrity. Loss of p97 gravely activated the unfolded protein response (UPR) and modulated the expression of multiple stemness and pluripotency regulators, including C/EBPδ, c-MYC, SOX2, and SKP2, which collectively contributed to the demise of CSCs. In summary, p97 controls the breast CSC integrity through multiple targets, many of which directly affect cancer stemness and are induced by UPR activation. Our findings highlight the importance of p97 and ER proteostasis in CSC biology and anticancer therapy.