In cancer research, autophagy has been revealed as one of the major ways to maintain the metabolism of cancer cells, including glioma cells, through protein degradation. Meanwhile, autophagy is also regarded as a kind of mechanism to protect glioma cells from a harmful stimulus, such as chemical and radiation treatment. So, the inhibition of autophagy may be very helpful in curing glioma. This study aimed to determine the effect of autophagic inhibition on glioma cells using tubacin, a specific inhibitor of histone deacetylase 6(HDAC6). According to the results, tubacin inhibited the growth of both U251 and LN229 cells, which was accompanied by lower HDAC6 activity and accumulated autophagosome. The inhibition of HDCA6 also led to accumulation of autophagosome and death of glioma cells. Moreover, the combined treatment of tubacin and temozolomide, an alkylating agent used to treat glioblastoma, induced more severe glioma cell death. Thus, it can be concluded that inhibition of HDAC6 suppressed growth and drug resistance of glioma cells in-vitro through autophagic suppression and blocking of fusion of autophagosome and lysosome.