The macromolecule that carries genetic information, DNA, is considered as an exceptional target for diseases depending on cellular division of malignant cells (i.e. cancer), microbes (i.e. bacteria) or parasites (i.e. protozoa). To aim for a comprehensive review to cover all aspects related to DNA targeting would be an impossible task and, hence, the objective of the present review is to present, from a medicinal chemistry point of view, recent developments of compounds targeting the minor groove of DNA. Accordingly, we discuss the medicinal chemistry aspects of heterocyclic small-molecules binding the DNA minor groove, as novel anticancer, antibacterial and antiparasitic agents.