Poly(l-lysine)-based materials are extensively studied for potential biomedical applications. Among them, synthetic polymer–poly(l-lysine) hybrids are of particular interest due to the combination of advantageous properties characteristic of both components. In this chapter, the synthetic routes to poly(l-lysine)-based hybrid copolymers of various architectures are described. Contemporary synthetic methods such as controlled radical polymerization, controlled ring-opening polymerization of N-carboxyanhydrides (NCAs) and “click” chemistry as tools for the preparation of complex macromolecular architectures with poly(l-lysine) segments are presented. The potential applications of these materials in gene and drug delivery, tissue engineering and diagnostics are discussed.