Identification of a new functional target of haloperidol metabolite: implications for a receptor-independent role of 3-(4-fluorobenzoyl) propionic acid Hyeon Soo Kim,* Minseok Song,* Sanatombi Yumkham,* Jang Hyun Choi,* Taehoon Lee,  Joseph Kwon, Sung Jae Lee,à Jong-In Kim,à Kang-Woo Lee,§ Pyung-Lim Han,§ Seung Woo Shin,¶ Ja-Hyun Baik,¶ Yong Sik Kim,** Sung Ho Ryu* and Pann-Ghill Suh* *Department of Life Science, Division of Molecular and Life Science, Pohang University of Science and Technology, Kyungbuk, South Korea  Sigmol Incorporation, Kyungbuk, South Korea àPeptide Library Support Facility, POSTECH Biotech Center, Pohang University of Science and Technology, Kyungbuk, South Korea §Department of Neuroscience, Medical Research Institute, Ewha Womans University School of Medicine, Seoul, South Korea ¶School of Life Sciences and Biotechnology, Korea University, Seoul, South Korea **Department of Psychiatry and Behavioral Sciences and Institute of Human Behavioral Medicine, Seoul National University Hospital, Seoul, South Korea Abstract Haloperidol, a dopamine D2 receptor blocker, is a classical neuroleptic drug that elicits extrapyramidal symptoms. Its metabolites include 3-(4-fluorobenzoyl) propionic acid (FBPA) and 4-(4-chlorophenyl)-4-piperidinol (CPHP). Until now, the biological significance of these metabolites has remained largely unknown. Here, we report that the administration of FBPA to mice effected a suppression of locomotor activity and induced catalepsy in a manner similar to that observed with haloperidol, whereas CPHP had no significant effects. Neither of these two metabolites, however, exhibited any ability to bind to the dopamine D2 receptor. FBPA blocked dopamine- induced extracellular signal-regulated kinase 1/2 phosphory- lation, and it specifically affected mitogen-activated protein kinase kinase (MEK)1/2 activity in hippocampal HN33 cells. Moreover, FBPA was capable of direct interaction with MEK1/ 2, and inhibited its activity in vitro. We demonstrated the generation of haloperidol metabolites within haloperidol-trea- ted cells by mass spectrometric analyses. Collectively, our results confirm the biological activity of FBPA, and provide initial clues as to the receptor-independent role of haloperidol. Keywords: antipsychotic drug, dopamine D2 receptor, ha- loperidol, metabolite, mitogen-activated protein kinase kinase. J. Neurochem. (2006) 99, 458–469. Haloperidol is a classical neuroleptic drug that has been used widely in the treatment of an array of neuropsychiatric disorders. Although this treatment has been associated with a high rate of success, its chronic use has also been reported to induce some extrapyramidal movement disorders, including tardive dyskinesia, resulting in serious limitations on its use as an antipsychotic agent (Jenner et al. 1980). The clinical potency of haloperidol has been correlated directly with its capacity to inhibit agonist–dopamine D2 receptor interaction (Creese et al. 1976). As the inhibitory effect on agonist– receptor interaction appears to occur almost instantaneously, the chronic effects of the drug cannot be explained simply by Received April 6, 2006; revised manuscript received June 3, 2006; accepted June 5, 2006. Address correspondence and reprint requests to Pann-Ghill Suh, Signaling Network Laboratory, POSTECH Biotech Center, Pohang University of Science and Technology, San31 Hyoja-Dong Nam-Gu Pohang, Kyungbuk 790–784, South Korea. E-mail:pgs@postech.ac.kr Abbreviations used: CHO, Chinese hamster ovary; CPHP, 4-(4-chlo- rophenyl)-4-piperidinol; DMSO, dimethylsulfoxide; EGF, epidermal growth factor; ERK, extracellular signal-regulated kinase; FBPA, 3-(4- fluorobenzoyl) propionic acid; GFP, green-fluorescence protein; GST, glutathione S-transferase; HAL, haloperidol; IB, immunoblotting; IP, immunoprecipitation; MAPK, mitogen-activated protein kinase; MEK, mitogen-activated protein kinase kinase; PBS, phosphate-buffered saline; PLC, phospholipase C; SAL, saline; SDS, sodium dodecyl sulfate. Journal of Neurochemistry , 2006, 99, 458–469 doi:10.1111/j.1471-4159.2006.04108.x 458 Journal Compilation Ó 2006 International Society for Neurochemistry, J. Neurochem. (2006) 99, 458–469 Ó 2006 The Authors