N1- (2, 5-dimethoxyphenyl)-N8-hydroxyoctanediamide (N25) is a novel SAHA cap derivative of HDACi,with a patent (No. CN 103159646). This invention is a hydroxamic acid compound with a structural formulaof RNHCO(CH2)6CONHOH (wherein R=2, 5dimethoxyaniline), a pharmaceutically acceptable salt which issoluble. In the present study, we investigated the effects of N25 with regard to drug distribution and moleculardocking, and anti-proliferation, apoptosis, cell cycling, and LD50. First, we designed a molecular approach formodeling selected SAHA derivatives based on available structural information regarding human HDAC8 incomplex with SAHA (PDB code 1T69). N25 was found to be stabilized by direct interaction with the HDAC8.Anti-proliferative activity was observed in human glioma U251, U87, T98G cells and human lung cancer H460,A549, H1299 cells at moderate concentrations (0.5-30μM). Compared with SAHA, N25 displayed an increasedantitumor activity in U251 and H460 cells. We further analyzed cell death mechanisms activated by N25 in U251and H460 cells. N25 significantly increased acetylation of Histone 3 and inhibited HDAC4. On RT-PCR analysis,N25 increased the mRNA levels of p21, however, decreased the levels of p53. These resulted in promotion ofapoptosis, inducing G0/G1 arrest in U251 cells and G2/M arrest in H460 cells in a time-dependent and dosedependentmanner. In addition, N25 was able to distribute to brain tissue through the blood-brain barrier ofmice (LD50: 240.840mg/kg). In conclusion, our findings demonstrate that N25 will provide an invaluable tool toinvestigate the molecular mechanism with potential chemotherapeutic value in several malignancies, especiallyhuman glioma.