Duocarmycin–pyrrolobenzodiazepine (PBD) dimers are DNA minor groove cross-linking agents formed by linking together an alkylating subunit from the duocarmycin and pyrrolobenzodiazepine classes of natural products. This chapter describes the structures of the 14 known examples, along with representative syntheses, and characterisation of the dimers' DNA alkylating ability (sequence selectivity and cross-linking efficiency) and cytotoxicity in a diverse range of tumour cell lines. One particular dimer example, with a relatively simple structure and concentrations giving 50% of maximum inhibition (IC50) values in the low pM range, has been incorporated into a variety of payload–linkers suitable for ADC application. The synthesis of one such payload–linker is described, along with antibody conjugation strategies, and preclinical anti-tumour activity of the resulting ADCs. The chapter concludes with a summary of closely-related duocarmycin–piperidinobenzodiazepine dimers which have also recently been disclosed as ADC payloads.