Abstract Pharmacophore modeling of large, drug- like molecules, such as the dopamine reuptake inhibi- tor GBR 12909, is complicated by their flexibility. A comprehensive hierarchical clustering study of two GBR 12909 analogs was performed to identify repre- sentative conformers for input to three-dimensional quantitative structure–activity relationship studies of closely-related analogs. Two data sets of more than 700 conformers each produced by random search confor- mational analysis of a piperazine and a piperidine GBR 12909 analog were studied. Several clustering studies were carried out based on different feature sets that include the important pharmacophore elements. The distance maps, the plot of the effective number of clusters versus actual number of clusters, and the novel derived clustering statistic, percentage change in the effective number of clusters, were shown to be useful in determining the appropriate clustering level. Six clus- ters were chosen for each analog, each representing a different region of the torsional angle space that determines the relative orientation of the pharmaco- phore elements. Conformers of each cluster that are representative of these regions were identified and compared for each analog. This study illustrates the utility of using hierarchical clustering for the classifi- cation of conformers of highly flexible molecules in terms of the three-dimensional spatial orientation of key pharmacophore elements. Keywords Clustering Æ Cocaine Æ Conformational analysis Æ Dopamine reuptake inhibitor Æ GBR 12909 Æ Hierarchical Æ Molecular mechanics Æ Pharmacophore modeling Æ XCluster Abbreviations %DEff percentage change in the effective num- ber of clusters 3D-QSAR three-dimensional quantitative structure– activity relationship CoMFA Comparative Molecular Field Analysis DA dopamine DAT dopamine transporter FRC fuzzy relational clustering QSAR quantitative structure-activity relation- ship RMSD root mean square deviation RTB number of rotatable bonds SAR structure–activity relationship SERT serotonin transporter Introduction Cocaine addiction is a world-wide problem. The dopamine hypothesis [1] suggests that the biochemical side effects of cocaine are due to the inhibition of dopamine reuptake by the binding of cocaine (Fig. 1a) to the dopamine transporter (DAT), resulting in an excess of dopamine in the synaptic cleft. Chronic use of cocaine leads to depletion of dopamine in the reward centers of the brain, contributing to the potential for relapse from cocaine withdrawal. A treatment agent is sought which would inhibit the binding of cocaine to K. M. Gilbert Æ C. A. Venanzi (&) Department of Chemistry and Environmental Science, New Jersey Institute of Technology, University Heights, Newark, NJ 07102, USA e-mail: venanzi@adm.njit.edu J Comput Aided Mol Des (2006) 20:209–225 DOI 10.1007/s10822-006-9046-2 123 ORIGINAL PAPER Hierarchical clustering analysis of flexible GBR 12909 dialkyl piperazine and piperidine analogs Kathleen M. Gilbert Æ Carol A. Venanzi Received: 2 January 2006 / Accepted: 20 April 2006 / Published online: 20 July 2006 Ó Springer Science+Business Media B.V. 2006