The utilization of macromolecules in the therapy of cancer and other diseases is becoming increasingly relevant. Recent advances in molecular biology and biotechnology have made it possible to improve the targeting and design of cytotoxic agents or DNA complexes for clinical applications. In many cases, to achieve the desired biological effect of these macromolecules, internalization to the cell cytosol is crucial. Although new delivery systems have improved the cellular uptake of macromolecules, tissue penetration, cellular uptake and efficient transfer of the molecules into the cytosol of target cells are still fundamental obstacles. At an intracellular level, the greatest obstruction for cytosolic release of the therapeutic molecule is the membrane barrier of endocytic vesicles. Photochemical activation and delivery of drugs and genes are novel technologies to overcome these obstacles. The background theory to these technologies is described in this review and recent data in this field are discussed.