Aminoglycosides are antibiotics commonly used to treat life-threatening gram negative bacterial infections. However, their use is hampered by their severe nephrotoxicity due to accumulation in cells of the renal proximal tubules. Several pathways have been implicated in the renal uptake of aminoglycosides including megalin, an endocytic receptor in proximal tubular cells. Previously, we have demonstrated that uptake of aminoglycosides into the kidney directly correlates with renal megalin activity and is completely eliminated in mice genetically deficient for this receptor. These studies provided unequivocal evidence that megalin is the major pathway responsible for renal aminoglycoside accumulation and that the receptor represents a unique drug target to prevent aminoglycoside-induced nephrotoxicity in patients. Following up on this observation, we now have elucidated the structural basis for megalin-aminoglycoside interaction in detail and we have used rational drug design to develop small molecules that block binding of aminoglycosides but not of physiological ligands to the receptor. Furthermore, we obtained proof of concept in mouse and rat models for the use of these compounds to prevent renal uptake and nephrotoxicty of these antibiotics.