Apoptotic cell death plays a predominant role in histone deacetylase (HDAC) inhibitor-induced cytotoxicity.Nuclear morphological changes and activation of apoptotic executors are involved in CTS203-induced cell death.However, emerging issues of HDAC inhibitor-resistance have been observed in patients. Herein, MCF-7 cellswere continuously exposed to CTS203 until the derived cells could proliferate normally in its presence. Thenewly obtained CTS203-resistant cells were nominated as MCF-7/203R. Compared to MCF-7 original cells, theMCF-7/203R cells were less sensitive to CTS203-induced apoptosis, with a minimal 6-fold higher IC50 value. Incontrast, the expression of Beclin-1 was dramatically up-regulated, positively correlated to the acquisition ofCTS203-resistance. Our results revealed the participation of autophagy in acquired HDAC inhibitor-resistanceand further identified Beclin-1 as a promising target for anti-drug resistance.