We report the activities of 62 bisphosphonates as inhibitors of the Leishmania major mevalonate/isoprene biosynthesis pathway enzyme, farnesyl pyrophosphate synthase. The compounds investigated exhibit activities (IC(50) values) ranging from approximately 100 nM to approximately 80 microM (corresponding to K(i) values as low as 10 nM). The most active compounds were found to be zoledronate (whose single-crystal X-ray structure is reported), pyridinyl-ethane-1-hydroxy-1,1-bisphosphonates or picolyl aminomethylene bisphosphonates. However, N-alicyclic aminomethylene bisphosphonates, such as incadronate (N-cycloheptyl aminomethylene bisphosphonate), as well as aliphatic aminomethylene bisphosphonates containing short (n = 4, 5) alkyl chains, were also active, with IC(50) values in the 200-1700 nM range (corresponding to K(i) values of approximately 20-170 nM). Bisphosphonates containing longer or multiple (N,N-) alkyl substitutions were inactive, as were aromatic species lacking an o- or m-nitrogen atom in the ring, or possessing multiple halogen substitutions or a p-amino group. To put these observations on a more quantitative structural basis, we used three-dimensional quantitative structure-activity relationship techniques: comparative molecular field analysis (CoMFA) and comparative molecular similarity index analysis (CoMSIA), to investigate which structural features correlated with high activity. Training set results (N = 62 compounds) yielded good correlations with each technique (R(2) = 0.87 and 0.88, respectively), and were further validated by using a training/test set approach. Test set results (N = 24 compounds) indicated that IC(50) values could be predicted within factors of 2.9 and 2.7 for the CoMFA and CoMSIA methods, respectively. The CoMSIA fields indicated that a positive charge in the bisphosphonate side chain and a hydrophobic feature contributed significantly to activity. Overall, these results are of general interest since they represent the first detailed quantitative structure-activity relationship study of the inhibition of an expressed farnesyl pyrophosphate synthase enzyme by bisphosphonate inhibitors and that the activity of these inhibitors can be predicted within about a factor of 3 by using 3D-QSAR techniques.