Histone deacetylase (HDAC) proteins are epigenetic regulators that deacetylate protein substrates, leading to subsequent changes in cell function. HDAC proteins are implicated in cancers, and several HDAC inhibitors have been approved by the FDA as anticancer drugs, including SAHA (suberoylanilide hydroxamic acid; Vorinostat and Zolinza). Unfortunately, SAHA inhibits most HDAC isoforms, which limits its use as a pharmacological tool and may lead to side effects in the clinic. In this work SAHA analogues substituted at the C2 position were synthesized and screened for HDAC isoform selectivity in vitro and in cells. The most potent and selective compound, C2-n-hexyl SAHA, displayed submicromolar potency with 49- to 300-fold selectivity for HDAC6 and HDAC8 compared to HDAC1, -2, and -3. Docking studies provided a structural rationale for selectivity. Modification of the nonselective inhibitor SAHA generated HDAC6/HDAC8 dual selective inhibitors, which can be useful lead compounds toward developing pharmacological tools and more effective anticancer drugs.