Cyclotides are small disulfide-rich peptides that are characterized by a head-to-tail cyclized peptide backbone and a knotted arrangement of three conserved disulfide bonds. They are present in many plants from the Violaceae, Rubiaceae and Cucurbitaceae families, with individual plants expressing a suite of dozens of cyclotides. So far >140 sequences and 15 three-dimensional structures have been determined but it is estimated that the family probably comprises many thousands of members. Their primary function in plants is thought to be as defense agents, based on their potent insecticidal activity, but they also have a range of other biological activities, including anti-HIV, antimicrobial and cytotoxic activities. Because of their exceptional stability they have attracted interest as templates for protein engineering and drug design applications. This article gives an overview of the discovery of cyclotides, describes their unique structural features and range of bioactivities, and discusses their applications in drug design.