<jats:title>Abstract</jats:title><jats:p>Three new 3,6‐dioxygenated diketopiperazines, lepistamides A–C (<jats:bold>1</jats:bold>–<jats:bold>3</jats:bold>), along with a known compound, diatretol (<jats:bold>4</jats:bold>), were isolated from the mycelial solid cultures of the basidiomycete <jats:italic>Lepista sordida.</jats:italic> Their structures were elucidated by spectroscopic means. The isolated diketopiperazines were evaluated for the cytotoxic activity against Astc‐a‐1, Bel 7402, and HeLa cell lines, and their biogenetic pathway was discussed.