BASIC SCIENCES

RADIOCHEMISTRY AND RADIOPHARMACEUTICALS

Neuroleptic Binding Sites: Specific Labeling in Mice with [18]Haloperidol, A

Potential Tracer for Positron Emission Tomography

P. B. Zanzonico, R. E. Bigler, and B. Schmall

Memorial Sloan-Kettering Cancer Center, New York, New York

Haloperidol labeled with fluorine-18 (Ti/a = 110 min, positron emission 97%), prepared yielding .04 Ci/mlllimole by the Balz-Schiemann reaction, was evaluated in a murine model as a potential radiotracer for noninvasive determination, by posi tron-emission tomography, of regional concentrations of brain dopamine receptors in patients. As the haloperldol dose In mice was increased from 0.01 to 1000 fig/kg, the relative concentration of [18F]haloperidol (/¿Ciper g spec¡men/¿tCiper g of body mass), at one hour after injection decreased from 30 to 1.0 in the strl- atum and from 8.0 to 1.0 in the cerebellum. The striata! radioactivity, plotted as rel ative concentration against log of dose, decreased sigmoidally, presumably re flecting competition between labeled and unlabeled haloperidol for a single class of accessible binding sites. Because the cerebellum is relatively deficient in dopa mine receptors, the observed decrease in cerebellar radioactivity may reflect a saturable component of haloperidol transport into brain. The high brain concentra tions and the unexpectedly high striâtum-to-cerebellumconcentration ratios (>4 at haloperidol doses <1 ¿ig/kg)suggest that [18F]haloperidol warrants further in vestigation as a potential radiotracer for dopamine receptors.

J NucíMod 24: 408-416,1983

Aberrant dopaminergic neuronal transmission has conjunction with a suitable radiotracer for dopamine been implicated in the pathogenesis of various neurologic receptors, for the noninvasive determination of regional disorders, including Parkinsonism (1-3), Huntington's concentrations of brain dopamine receptors in vivo. chorea (1,4,5), and tardive dyskinesia (6-10), as well Specific dopamine receptors are present in brain in as of certain neuroses (11) and psychoses (12-15). nanomolar concentrations (22), and therefore will be Characterization of dopamine receptors has been ac saturated at comparably low brain concentrations of a complished largely using radiometrie binding assays of dopaminergic ligand. In contrast, nonspecific binding synaptic membrane preparations derived from animal sites are practically nonsaturable (23-26). A potential or postmortem brain specimens (16-19). The highly dopamine-receptor-binding radiotracer must have a invasive and nonphysiologic nature of these procedures sufficiently high specific activity to yield an adequate severely limits their applicability to man and makes in photon flux for statistically significant radiation counting terpretation of results somewhat problematic. Conse at ligand doses that do not saturate the specific binding quently there is great interest in the potential utilization sites. Because theoretical maximum specific activity is of positron-emission tomography (PET) (20,21), in inversely proportional to half-life, dopaminergic ligands labeled with short-lived radionuclides—most notably carbon-11 (Ti/2 = 20.4 min, positron emission 100%) Received Sept. 24, 1982; revision accepted Dec. 17, 1982. and fluorine-18 (T|/2 = 110 min, positron emission For reprints contact: Dr. Pat Zanzonico, Biophysics Laboratory, Memorial Sloan-Kettering Cancer Center, 1275 York Ave., New 97%)—have great potential as dopamine receptor- York, NY 10021. binding radiotracers. For compounds singly labeled with

408 THE JOURNAL OF NUCLEAR MEDICINE BASIC SCIENCES RADIOCHEMISTRY AND RADIOPHARMACEUTICALS carbon-11 or fluorine-18, the theoretical maximum (Schmall B, Tilbury RS, Nisselbaum, JS, unpublished specific activities are 9.2 X IO6and 1.7 X IO6Ci/mmole, data). Although the resulting specific activity 400 respectively. mCi/mmole appears to be somewhat low for receptor Haloperidol and spiroperidol—potent neuroleptics of studies in vivo, such studies are feasible in the "partial- the butyrophenone series, widely used in the manage body counting facility" a high-sensitivity, low-back ment of schizophrenia—have been shown to bind spe ground radiation counting facility designed and installed cifically, reversibly, and with high affinity to synaptic in our center's biophysics laboratory (45). membrane preparations in vitro (22). Demonstration in vivo of specific spiroperidol binding sites has been ac MATERIALS AND METHODS complished using specifically tritiated spiroperidols of high specific activity, namely l-phenyl-[3-3H]spirop- Chemicals.All commerciallyobtained chemicals and eridol (sp. ac. 8-15 Ci/mmole) (27-30) and 1-phenyl- solventswere of reagent grade and were used as supplied. [4-3H]spiroperidol (30 Ci/mmole) (31,32), and also Haloperidol in crystalline form was a gift.* [G-3H]- />-[77Br]bromospiroperidol (5-10 Ci/mmole) (33,34). Haloperidol (14 Ci/mmole) was obtained commercially. Radiochemical stability of the tritiated ligands in vivo The radiochemical purity (97%) was verified by TLC on has been demonstrated by thin-layer chromatography silica gel* jn chloroform/methanol/ammonium hy (TLC) (27,29,31). In the case of l-phenyl-[3H]spirop- droxide (98:2:1). eridol, the striatum-to-cerebellum radioactivity con Animals. All experiments used male and female B6 centration ratio at 1hr after injection decreased from 8.8 mice, maintained on a 12-hr light-dark cycle, with free to 1.0 as the spiroperidol dose was increased from 44 to access to food and water and housed 8-10 to a cage. The 2000 Mg/kg (27,29). Because dopamine receptors are mice were approximately six months old and weighed abundant in the striatum and relatively deficient in the 20-25 g each. cerebellum (22), this decrease in the striatum-to-cere Preparationandanalysis of "de-fluorinatedhaloper idol". This compound, 4-[4-(4-chlorophenyl)-4-hy- bellum ratio presumably reflects saturation of striatal dopamine receptors and the ensuing dose-dependent droxypiperidino] butyrophenone, appears to be a ligand decrease in the proportion of specifically bound spirop binding to dopamine receptors (46), and was therefore eridol in brain. In contrast, demonstration in vivo of prepared as an analytical standard for high-pressure specific haloperidol binding sites using tritiated halo- liquid chromatography. It was synthesized by the N- peridol has not yet been accomplished. The specific ac alkylation of 4-(4-chlorophenyl)-4-hydroxypiperidine tivities of randomly labeled tritiated haloperidol (0.042 with -y-chlorobutyrophenone, and was purified by re- Ci/mmole) and of 4'-fluorobutyrophenone-[2'-3H]- crystallization from 95% ethanol (mp:125-128°C;Ref. haloperidol (0.1 Ci/mmole), though adequate for 46: 128-130°C). Thin-layer chromatography of the pharmacokinetic studies (35-40), are apparently too low product yielded a single spot (Rf = 0.75). The identity for the discrimination of specifically localized from of the product was verified by chemical ionization mass nonspecifically localized haloperidol in brain (37,40). spectrometry. Though of adequately high specific activity, the unex Preparationandanalysisof [18F]haloperidol.This was pected absence of selective striatal localization of 4- prepared from the corresponding tetrafluoroborate di (4-chlorophenyl)-[3-3H]-haloperidol (8-18 Ci/mmole) azonium salt (chemical yield 20-30%, radiochemical at a haloperidol dose of only 5 Mg/kg (30) suggests, yield 0.5-1.0%) by the Balz-Schiemann reaction using perhaps, that it may undergo spontaneous de-tritiation aqueous [18F]fluoride, as developed by Kook et al. (43) in vivo. and adapted by Schmall and Bigler (44). [I8F]Fluoride We have prepared [l8F]haloperidol by the Balz- was produced in our compact medical cyclotron via the Schiemann reaction and have evaluated it in a murine nuclear reaction 16O(3//e,p)18F by the irradiation of model as a radioligand for the specific labeling in vivo water with 22-MeV helium-3 ions (47,48). Chemical of neuroleptic binding sites. Because of the technical and radiochemical purity was greater that 95%, as de constraints imposed by a short-lived, cyclotron-produced termined by reverse isotope dilution analysis, thin-layer radionuclide such as fluorine-18, we have concurrently chromatography, and high-pressure liquid chromatog evaluated [G-3H]haloperidol as a convenient model raphy. Reverse isotope dilution analysis was accom radioligand for the determination of the dose-dependent plished by chromatographing an admixture of [I8F]- regional concentrations of haloperidol in brain. [G- haloperidol and a large excess of unlabeled haloperidol 3H]Haloperidol is commercially available and has been on neutral alumina (Brockman activity 1, 80-200 mesh) used in receptor binding assays in vitro (41,42). [I8F]- and comparing its specific activity before and after Haloperidol prepared by the Balz-Schiemann reaction chromatography. Analysis of the product by thin-layer includes carrier because fluorine-18 is introduced by chromatography on silica gel in chloroform/ isotopie exchange with stable fluorine in the corre methanol/ammonium hydroxide (98:2:1) and in chlo sponding tetrafluoroborate diazonium salt (43-44). roform/ethyl acetate/formic acid (5:4:1) yielded single

Volume 24, Number 5 409 ZANZONICO, BIGLER, AND SCHMALL

radioactive spots (Rf = 0.75 and 0.36, respectively) 1 Mg/kg or less, each specimen was placed in a small identical to those of haloperidol. High-pressure liquid polyethylene tube and the radioactivity counted by an chromatography of the product was carried out using an nihilation coincidence detection. This utilized the par analytical reverse-phase (PAX 5-1025 ODS-3) column tial-body-counting facility, a radiation counter using two and a solvent system of 0.1 mM ammonium acetate/ heavily shielded Nal(Tl) crystals 23-cm in diameter by methanol (65:35). Mass and radioactivity were mea 13 cm thick. When operated in coincidence (coincidence sured simultaneously using a refractometer and a resolving time: 100 nsec) and with a crystal separation Nal(Tl) well counter with logarithmically scaled ra of 4 cm, the background counting rate is 0.7-0.8 cpm and diation meter, respectively, interfaced to a dual-trace the overall counting efficiency for 0.511-MeV annihi strip-chart recorder. The retention time for defluo- lation photons is ~20%. For tissue specimens from mice rinated haloperidol was 11 min (k' = l .2) and for halo with haloperidol doses of 10 Mg/kg or greater, each peridol 12 min (k' = 1.4). High-pressure liquid chro specimen was placed in a 20-ml glass counting vial and matography yielded a single mass peak and a single ra the radioactivity counted in an automatic gamma dioactive peak, co-eluting with precisely the same re counter with an energy window spanning 450-570 keV. tention time as haloperidol. There was no detectable After correcting for background and for decay (to the defluorinated haloperidol in the [18F]haloperidol prep time of injection), the amount of radioactivity in each aration. The specific activity of [18F]haloperidol, de specimen was expressed as relative concentration. termined using ultra-violet spectrophotometry and a Analysis of the chemical form of radioactivity in vivo. calibrated Nal(Tl) well counter, was 0.2-0.4 Ci/mmole The striata and the cerebella of several mice injected at the time of injection. with [G-3H]haloperidol or with [18F]haloperidol were Injectionanddissectionprocedure.Mice wereinjected combined in chloroform and homogenized manually with under light ether anesthesia with either [G-3H]- or a glass homogenizer. Each of the resulting suspensions [18F]haloperidol in 0.1 ml acidified, ethanolic isotonic was filtered and an aliquot of each of the respective fil saline via the retro-orbital venous plexus. The doses were trates admixed with haloperidol. Each of the resulting 1.0, 10, 25, 100, and 1000/ig/kg for [G-3H]haloperidol solutions was analyzed by TLC. and 0.010, 0.10, 1.0, 10, 25, 100, and 1000 Mg/kg for [18F]haloperidol, adjusted before injection as necessary RESULTS by the admixture of an appropriate amount of unlabeled haloperidol with the radioagent. There were four mice Striata! and cerebellar H-3 concentrations as a func per dosage group. Each mouse was killed at 1 hr by tion of haloperidol dose. Figure 1 shows the relative cervical dislocation and immediately decapitated. The concentrations of tritium in the striatum and cerebellum entire brain was removed and placed on an ice-coldmetal of mice, as a function of dose, 1 hr after the adminis surface. The striatum and the cerebellum were excised tration of [G-3H] haloperidol. As the log dose of halo according to the method of Glowinski and Iversen (49). peridol is increased from 1.0 to 1000 Mg/kg, the relative Each tissue specimen was placed on a tared piece of concentrations of tritium in the striatum and in the weighing paper, the paper and specimen weighed, and cerebellum remain constant at a value of approximately the wet mass of the specimen determined by differ 2.0. ence. Striata! and cerebellar F-18 concentrations as a Counting of tritium in tissue specimens. Each tissue functionof haloperidoldose. Figure 2 shows the relative specimen was placed in a 20-ml glass counting vial. concentrations of F-18 in the striatum and cerebellum Enough tissue solubilizer (typically 0.5 ml) was added of mice as a function of dose 1hr after the administration to each vial to immerse the tissue completely. The of [18F]haloperidol. As the log dose of haloperidol is specimens were incubated for 48 hr at 50°C.After increased from 0.01 to 1000 Mg/kg, the relative con cooling, 15 ml of scintillation cocktail1 were added to centrations of fluorine-18 in the striatum and in the each vial and the radioactivity counted in an automatic cerebellum decrease in a sigmoidal fashion. The dose- beta counter using a preset "tritium" energy window. dependent fall-off in the striatum is much more rapid The counting efficiency for tritium was 58%. Knowing than that in the cerebellum. The striatum-to-cerebellum the mass and the activity per unit mass of the radioactive fluorine-18 concentration ratio, shown in Fig. 3 as a solution injected into each mouse, the injected dose of function of log dose of haloperidol, likewise decreases in activity was determined. After correcting for back sigmoidal fashion. ground, the amount of radioactivity in each tissue Chemical form of radioactivity in vivo. Thin-layer specimen was expressed as relative concentration (¿uCi chromatography of tritium in mouse brain yielded a found per g specimen/MCi injected per g body mass chromatogram on which the radioactivity was diffusely (50). distributed. The absence of a notable radioactive spot Countingof fluorine-18in tissuespecimens. Fortissue co-migrating with haloperidol suggests that [G-3H]- specimens from mice injected with haloperidol doses of haloperidol has undergone substantial de-tritiation in

410 THE JOURNAL OF NUCLEAR MEDICINE BASIC SCIENCES RADIOCHEMISTRY AND RAD1OPHARMACEUTICALS

§

|"F| HALOPERIDOL STRIATUM

CEREBELLUM 001 0.1 1.0 10 100 1000 HALOPERIDOLLXISE(«g/kg) I J_ J_ J FIG. 3. Striatum-to-cerebellum radioactivity concentration ratios 1.0 10 100 1000 in mice at 1 hr after injection of [G-3H]- and of [18F]-haloperidol as HALOPERIDOLDOSE(//g/kg) FK».1. Relative concentrations of [G-3H]haloperidol in striatum a function of administered dose of haloperidol. Error bars represent standard error of the mean. and cerebellum of mice at 1 hr after injection, as a function of ad ministered dose of haloperidol. Error bars represent standard error of the mean. with receptor-mediated localization of [I8F]haloperidol in the striatum (Figs. 2 and 3). At haloperidol doses of vivo. In contrast, TLC of F-18 in mouse brain yielded a < 1 Mg/kg—that is, at doses below those necessary to chromatogram having a single radioactive spot, co- saturate specific binding sites—a relatively large fraction migrating precisely with haloperidol (Rf = 0.75) and of haloperidol in brain is specifically bound and is accounting for at least 80% of the radioactivity on the therefore localized in the striatum. As the haloperidol chromatogram. Because only a single radioactive spot dose is increased (1-100 Mg/kg), approaching and was observed, [I8F]haloperidol probably accounts for eventually exceeding that necessary to saturate the the remaining radioactivity as well. specific binding sites, an increasing proportion of halo peridol is bound nonspecifically. At high haloperidol DISCUSSION doses (> 100 Mg/kg)—that is, at doses that greatly ov- ersaturate the specific binding sites—nonspecific binding In the current study, evidence for the specific labeling in vivo of neuroleptic binding sites in mouse brain with predominates, with a notable absence of selective striata! [18F]haloperidol has been presented. Because dopamine localization of F-18. This is because nonspecific binding receptors are abundant in the striatum and relatively rare is generally considered to be as likely to occur in the in the cerebellum (22), the dose-dependent decrease in cerebellum as it is in the striatum, since neuroleptics the relative concentration in the striatum and in the readily localize in biological membranes because of their striatum-to-cerebellum concentration ratio is consistent high lipid solubility (23,26). Thin-layer chromatography of F-18 in mouse brain suggests that most of the radio activity represented intact [18F]haloperidol. The ap parent absence of any substantial extra-hepatic metab olism of haloperidol (35,39,51,52) and the rapid blood clearance (53) and excretion (35,39,51,52) of halo STRIATUM peridol metabolites, are consistent with this supposition. Because the carbon-fluorine bond is the single strongest bond formed by carbon (54), spontaneous de-fluorina- tion of haloperidol in vivo is highly unlikely. The results obtained with [G-3H]haloperidol are markedly different from those obtained with [1XF]ha loperidol. At all haloperidol doses, the relative concen tration of tritium in the striatum and in the cerebellum maintains a constant low value of approximately 2.0. Even at low haloperidol doses, where demonstrable 001 01 10 10 10 1000 binding to specific receptors is expected to occur, selec DOSE OF HALOPERIOOL FIG. 2. Relative concentrations of | lt!F| haloperidol in striatum and tive striatal localization of tritium is not observed. Thin-layer chromatography of tritium in mouse brain cerebellum of mice at 1 hr after injection, as a function of admin istered dose of haloperidol. Error bars represent standard error of suggests that practically none of the radioactivity rep the mean. resented haloperidol. Moreover, the relative concen-

Volume 24, Number 5 411 ZANZONICO, BIGLER, AND SCHMALL tration of tritium in mouse brain 1 hr after the admin relatively deficient in the cerebellum (22). Hollt et al. istration of 3H2O (1.1) (unpublished results) compares (27), Hollt and Schubert (29), and Kuhar et al. (31), all with that 1 hr after the administration of [G-3H]halo- found a marked dose-dependent decrease in the striata! peridol (1.9). It appears, therefore, that the radioactivity concentration of tritiated spiroperidol, but did not ob in vivo following the administration of [G-3H]haloper- serve any decrease in the cerebellar concentration. It is idol was largely "free" tritium, that is, tritiated water. unlikely, therefore, that the dose-dependent decrease in Thus, [G-3H] haloperidol is not useful for the specific the relative concentration of [18F]haloperidol in the labeling in vivo of neuroleptic binding sites because of cerebellum (Fig. 2) is receptor-mediated. Tewson et al. the lability of the tritium atoms. A critical, but often (59) found that pretreatment of a monkey with a phar times ignored, consideration in such an experiment is macologie dose of haloperidol reduced the fraction of clearly the stability of the tracer to spontaneous radio- no-carrier-added [18F]haloperidolextracted by the brain chemical degradation (e.g., detritiation) under the actual during a single capillary transit following injection into experimental conditions. the internal carotid artery. Because cerebral blood flow [G-3H]haloperidol is prepared commercially by cat was not altered, the observed decrease in extraction alytic exchange in the presence of tritiated water. fraction (i.e., from 0.7 to 0.4) was explained by postu Though nominally "generally labeled," the principal site lating that [I8F]haloperidol competes with excess un- of labeling is presumably the méthylènegroup in the a labeled haloperidol for a carrier-mediated transport position to the carbonyl function. The carbonyl function system at the blood-brain barrier. This postulated sat tends to increase the acidity of the a-hydrogen atoms urable component of haloperidol transport into brain (55). In order to determine whether its lability in vivo could also explain the dose-dependent decrease in the was due simply to the acidity of the a-hydrogen atoms, relative concentration of [I8F]haloperidol in the cere [G-3H]haloperidol was incubated at 37°Cfor 1 hr at bellum. Consistent with this hypothesis is the finding that various hydrogen-ion concentrations (pH 3,7, and 10). the cerebellar concentration of haloperidol in dogs de The radiochemical purity, as determined by TLC, was creased by nearly 50% as the haloperidol dose was in in all instances comparable to the specified value (97%) creased from 40 to 630 Mg/kg (37). for the commercial preparation (maintained at 4°C). The striatal and the cerebellar concentrations of ha Consistent with its utility in receptor-binding assays in loperidol, as indicated in Table 1,are much greater than vitro, conducted under physiologicconditions (37°C,pH those of other neuroleptics, including spiroperidol 7.4), the de-tritiation of [G-3H]haloperidol in vivois not (27-32), /7-bromospiroperido (33,34), and pimozide simply a temperature- or pH-dependent process. (28,30,60). The striatal and the cerebellar concentra Based principally on the differential binding affinities tions are not inconsistent with the whole-brain concen of dopaminergic ligands, the existence of distinct classes, trations determined by tissue distribution studies in rats or subsets, of dopamine receptors has been hypothesized using either 4'-fluorobutyrophenone-[2'-3H]haloperidol (22,56-58). For example, the putative "dopamine-2 (37-39) or [18F]haloperido (53), and by external scin- (D2)" receptor, which exhibits a thousand times the tigraphy in dogs and in monkeys using [18F]haloperidol affinity for antagonists as for agonists, has been impli (45,53, unpublished data). For a given level of statistical cated in the pathogenesis of schizophrenia (13-15,22), accuracy, the greater brain concentrations of haloperidol as well as in the antipsychgtic effect of neuroleptics will permit reduction of the dose of radioactivity, or the (¡5,22,26). The dose-dependent sigmoidal decrease in duration of data acquisition, for the determination by the relative concentration of- [18F]haloperidol in the PET of regional receptor concentrations. Alternatively, striatum and in the striatum-to-cerebellum concentration for a given dose of radioactivity, the greater brain con ratio appears to be conceptually consistent with a single centrations of haloperidol will serve to increase the sta class of accessible binding sites in vivo. Multiple classes tistical accuracy of regional receptor concentration of accessible binding sites in vivo would be expected to measurements. As indicated in Table 2, the brain-to- yield a dose-response curve (i.e., tissue concentration blood concentration ratios are likewise much greater for against log dose) consisting of a number of distinct sig haloperidol (39,53,61 ) than for other neuroleptics, in moidal components. Earlier studies, though suggestive cluding pimozide (61), trifluperidol (67), bromperidol of a single class of binding sites in vivo, have not unam (53,62), and spiroperidol (63). This will serve to increase biguously identified the shape of the dose-response curve further the statistical accuracy of regional receptor because of the limited range (>1 Mg/kg) of neuroleptic concentration measurements by reducing the correction doses studied (27,29,31,32). The current series, which for the vascular component of radioactivity within brain includes haloperidol doses as low as 0.01 Mg/kg, yielded tissue. an unambigously sigmoidal dose-response curve, thus If one assumes that striatal and cerebellar radioac suggesting the existence of a single class of accessible tivities represent total and free ligand, respectively, the binding sites in vivo. maximum total-to-free (i.e., striatum-to-cerebellum) In contrast to the striatum, dopamine receptors are haloperidol concentration ratio in vivo (4.6, Table 1) is

412 THE JOURNAL OF NUCLEAR MEDICINE BASIC SCIENCES RADIOCHEMISTRY AND RADIOPHARMACEUTICALS

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Volume 24, Number 5 413 ZANZONICO, BIGLER, AND SCHMALL

NEUROLEPTICSRadioligand4'-fluorobutyrophenone-[2'-3H]-haloperidol4'-fluorobutyrophenone-[2'-3H]-haloperidol[18F]haloperidol1-(4,4)-bis-

to-bloodconcentrationratio2651t122.1t21t2.20.67Reference396153616153,6263* afterInjection(hr)Ksc)0.5

(iv)1(iv)0.5

(iv)0.5 (iv)1(iv)0.5

(iv)Brain

rats.'Ovariectomized Cerebral cortex-to-plasma ratio.Dose(M9/kg)40500010500050002000.05*Speciesratrat*ratrat*rat'ratmouseTime Based on estimate of specific activity of no-carrier-added [11C]spiroperidol.

greater than that in vitro (1.2-1.4) (41,64). One of the invasive pharmacokinetic studies. For example, the assumptions implicit in this comparison—the utilization distributions of pharmacologie haloperidol doses in a of the ligand concentration in the cerebellum as an es monkey (44,53) and in a dog (unpublished data) have timate of the free ligand concentration in the stri- been determined by two-dimensional longitudinal atum—may perhaps require rigorous validation at some imaging and by PET. Because near-maximal receptor point. In comparison with haloperidol, the maximum binding is attained by several hours after injection striatum-to-cerebellum spiroperidol concentration ratio (27-34,60), the 110-min half-life of F-18 makes it well in vivo (4.5-8.8, Table 1) is not inconsistent with the suited for radiotracers binding to dopamine receptors. maximum total-to-free spiroperidol concentration ratio The striatal concentrations and the brain-to-blood con in vitro (5-15) (41,64). These data suggest, perhaps, that centration ratios are much greater for haloperidol than at least for haloperidol the receptor-binding affinity in for any other neuroleptic studied thus far. Most impor vivo may be somewhat greater than that determined in tantly, perhaps, the maximum striatum-to-cerebellum vitro. concentration ratio in vivo appears to be greater than In contrast to the striatum-to-cerebellum concentra that expected on the basis of receptor-binding assays in tion ratio, the unusually high brain concentrations (Table vitro. No-carrier-added [18F]haloperidol has been pre 1) and brain-to-blood concentration ratios (Table 2) are pared, though in low yield, by the. triazene reaction probably not related to receptor binding. Because tissue (59,69) and by deaminative fiuorination (unpublished localization of a radiotracer is such a complex phe data). The current study suggests that {'8F]haloperidol nomenon, it should not be expected that target tissue warrants further investigation as a potential dopamine concentrations of a putative receptor-binding radiotracer receptor-binding radiotracer. For haloperidol, as well will always be correlated with receptor-binding affinity. as for other dopaminergic ligands, the mechanisms of For example, domperidone—ahighly potent and specific tissue localization and the nature of specific and non dopamine antagonist in vitro (65)—is completely ex specific binding sites require further elucidation. cluded from brain tissue by the blood-brain barrier be cause it is highly basic (pKa =11) and therefore highly FOOTNOTES

charged at physiologic pH (66, P. Seeman, personal * Generously provided by McNeil Laboratories, Fort Washington, communication). Bioavailability to brain (determined PA. by brain extraction efficiency and thus net lipid solubility * 1381 Silica gel Eastman Chromagram Sheet, Eastman Kodak in vivo), localization in and redistribution from other Company, Rochester, NY. tissues [particularly the lungs (67,68)], and metabolism 1 Insta-gel liquid scintillation cocktail, Packard Instrument Com and excretion, are most likely responsible for the ob pany, Inc., Downers Grove, IL. served differences in brain concentrations among neu- ACKNOWLEDGMENTS roleptics. In particular, the postulated carrier-med iated transport system for haloperidol may be largely We express our appreciation to Messrs. Richard Lee and J. Robert Dahl for the production of fluorine-18. responsible for its unusually high brain concentra This investigation was supported in part by the U.S. Department tions. of Energy Contract no. EE-77-S-02-4258 and by the National Cancer [l8F]Haloperidol has already proven useful in non- Institute Grant No. CA-09022.

414 THE JOURNAL OF NUCLEAR MEDICINE BASIC SCIENCES RAD1OCHEMISTRY AND RADIOPHARMACEUTICALS

REFERENCES neuroleptics to synaptosome membranes. J Pharmacol Exp Ther 190:123-130, 1974 /. BERNHEIMER H, BIRKMAYERW, HORNYKIEWICZ O, et 26. SEEMAN P: Anti-schizophrenic drugs-Membrane receptor al: Brain dopamine and the syndromes of Parkinson and sites of action. Biochem Pharmacol 26:1741-1748, 1977 Huntington./yVeuro/Sci 20:415-455, 1973 27. HÖLLTV, CZLONKOWSKI A, HERZ A: The demonstration 2. HORYNKIEWICZ O: Dopamine (3-hydroxytyramine) and in vivo of specific binding sites for neuroleptic drugs in mouse brain function. Pharm Rev 18:925-964, 1966 brain. Brain Res 130:176-183, 1977 3. DuvoiSlN R: Parkinsonism. CIBA Clinical Symposia 28: 28. LADURON P, LEYSEN J: Specific in vivo binding of neuro 1-29,1976 leptic drugs in rat brain. Biochem Pharmacol 26:1003-1007, 4. KLAWANS HL, PAULSON GW, RINGEL SP, et al: The use 1977 of L-DOPA in the presymptomatic detection of Huntington's 29. HÖLLTV, SCHUBERT P: Demonstration of neuroleptic re chorea. Adv Neural 1:295-300,1973 ceptor sites in mouse brain by autoradiography. Brain Res 5. TOLOSA ES, SPARSER SB: Apomorphine in Huntington's 151:149-153,1978 chorea: Clinical observations and theoretical considerations, 30. LADURON PM, JANSSEN PFM, LEYSEN JE: Spiperone. A Life Sci 15:1371-1380,1974 ligand of choice for neuroleptic receptors. 2. Regional distri 6. SCHIELE BC, GALLANT D, SIMPSON G, et al: Neurologic bution and in vivo displacement of neuroleptic drugs. Biochem syndromes associated with antipsychotic-drug use. A' Engl Pharmacol 27:317-321, 1978 J Med 289:20-23, 1973 31. KUHAR MJ, MURRIN LC, MALOUF AT, et al: Dopamine 7. KLAWANS HL, JR: The pharmacology of tardive dyskinesias. receptor binding in vivo: The feasibility of autoradiographic Am J Psychiatry \ 30:82-86, 1973 studies. Life Sci 22:203-207,1977 8. JESTE DV, WYATT RJ: Changing epidemiology of tardive 32. LE FUR G, GUILLOUX F, UZAN A: In vivo blockage of dyskinesia: An over view. Am J Psychiatry 138:297-309, dopaminergic receptors from different rat brain regions by 1981 classical and atypical neuroleptics. Biochem Pharmacol 9. BARBACCIA ML, TRABUCCHI M: Tardive dyskinesia: A 29:267-270, 1980 biological approach. In Neurochemistry and Clinical Neu 33. KULMALA HK, HUANG CC, DINERSTEIN RJ, et al: Spe rology, New York, Alan R. Liss, 1980, pp 181-193 cific in vivo binding of 77Br-p-bromospiroperidol in rat brain: 10. GOETZ CG, WEINER WJ, NAUSIEDA PA, et al: Tardive A potential tool for gamma ray imaging. Life Sci 28:1911- dyskinesia: Pharmacology and clinical implications. ("//// 1916,1981 Neuropharmacol 5:3-22, 1982 34. FRIEDMAN AM, HUANG CC, KULMALA HA, et al:The use //. BARCHAS JD, AKIL H, ELLIOTT GR, et al: Behavioral of radiobrominated p-bromospiroperidol for x-ray imaging neurochemistry: Neuroregulators and behavioral states. of dopamine receptors. Int J NucíMed Biol 9:57-61, 1982 Science 200:964-973, 1978 35. BRAUN GA, Poos GI, SOUDIJN W: Distribution excretion 12. MATTHYSSE S, LIPINSKI J: Biochemical aspects of schizo and metabolism of neuroleptics of the butyrophenone type. phrenia. Ann Rev Med 26:551-565, 1975 Part II. Distribution, excretion and metabolism of haloperidol 13. SNYDER SH: The dopamine hyopthesis of schizophrenia: in Sprague-Dawley rats. Eur J Pharmacol 1:58-62, 1967 Focus on the dopamine receptor. Am J Psychiatry 133: 36. SOUDIJN W, VAN WIJNGAARDEN I:A rapid and convenient 197-202,1976 method for the synthesis of labeled tertiary amines. J Labelled 14. MELTZER HY, STAHL SM: The dopamine hypothesis of Comp 4:159-163, 1968 schizophrenia: A review. Schizophrenia Bull 2:19-76, 37. JANSSEN PAJ, SOUDIJN W, VAN WIJNGAARDEN I: Pi- 1976 mozide, a chemically novel, highly potent and orally long- 15. SNYDER SH: Dopamine receptors, neuroleptics, and schiz acting neuroleptic drug. 3. Regional distribution of pimozide ophrenia. Am J Psychiatry 138:460-464, 1981 and of haloperidol in the dog brain. Arzneim-Forsch (Drug 16. SNYDER SH, BENNETT JP, JR: Neurotransmitter receptors Res) 18:282-287, 1968 in the brain: Biochemical identification. Ann Rev Physiol 38. JANSSEN PA, ALLEWIJN FT: The distribution of the buty- 38:153-175, 1976 rophenones haloperidol, trifluperidol, moperone, and cloflu- 17. WILLIAMS LT, LEVKOWITZ RJ: Receptor Binding Studies perol in rats, and its relationship with their neuroleptic activity. in Adrenergic Pharmacology, New York, Raven Press Books, Arzeim-Forsch (Drug Res) 19:199-208, 1969 Ltd, 1978 39. LEWI PJ, HEYKANTS JJ, ALLEWIJN FT: Distribution and 18. YAMAMURA HI, ENNA SJ, KUHAR MJ (Eds): Neuro metabolism of neuroleptic drugs. I. Pharmacokinetics of ha transmitter Receptor Binding, New York, Raven Press Books, loperidol. Arzeim-Forsch (Drug Res) 20:943-948, 1970 Ltd, 1978 40. NAYLOR RJ, OLLEY JE: The distribution of haloperidol in 19. COOPERJR, BLOOM RE, ROTH RH: The Biochemical Basis rat brain. BrJ Pharmacol 36:208P-209P, 1979 ofNeuropharmacology, New York, Oxford University Press, 41. HOWLETT DR, NAHORSKI SR: A comparative study of Ine, 1978, pp 60-71 [3H]haloperidol and [3H]spiroperidol binding to receptors 20. PHELPS ME: Emission computed tomography. Semin Nucí on rat cerebral membranes. FEBS Letters 87:152-156, Med 7:337-365, 1977 1978 21. RAICHLE ME: Quantitative in vivo autoradiography with 42. DEMENGE P, FEUERSTEIN C, MOUCHET P, et al: Stereo- positron emission tomography. Brain Res Rev 1:47-68, specific binding of 3H-haloperidol in rat dorsal spinal cord. 1979 Eur J Pharmacol 66:117-120, 1980 22. SEEMAN P: Brain dopamine receptors. Pharmacol Rev 32: 43. KOOK CS, REED MF, DIGENIS GA: Preparation of [18F]- 229-313, 1980 haloperidol. J Med Chem 18:533-535, 1975 23. SEEMAN P, BIALY HS: The surface activity of tranquilizers. 44. SCHMALL B, BIGLER RE: Radiolabeled drugs: Use of posi Biochem Pharmacol 12:1181-1191, 1963 tron-emitting radionuclides. In Radiopharmaceuticals: 24. SEEMAN P: The membrane actions of anesthetics and tran Structure-Activity Relationships, RP Spencer (Ed). New quilizers. Pharm Rev 24:583-655 1972 York, Gruñe& Stratton, Ine, 1981, pp 769-800 25. SEEMAN P, STAIMAN A, CHAU-WONG M: The nerve im 45. BIGLER RE, KOSTICK JA, WOODARD HQ, et al: Use of a pulse-blocking action of tranquilizers, and the binding of collimatecisource of fast neutrons to measure malignant tumor

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electrolyte exchange. IEEE Trans NucíSci NS-24:559-562, and antagonist states. Neurosci Symp 1:28-49, 1976 1977 57. KEBABIAN JW, CALNE DB: Multiple receptors for dopa 46. JANSSEN PAJ, VAN DE WESTERINGH C, JAGENEAU mine. Nature 277:93-96, 1979 AHM, et al: Chemistry and pharmacology of CNS depres 55. COSTALL B, NAYLOR RJ: The hypotheses of different do sants related to 4-(4-hydroxy 4-phenyl)-piperidino) butyro- pamine receptor mechanisms, Life Sci 28:215-229, 1981 phenone. Part I: Synthesis and screening data in mice. J Med 59. TEWSON TJ, RAICHLE ME, WELCH MJ: Preliminary Pharm Chem 1:281-297, 1959 studies with [18F]haloperidol: A radioligand for in vivo studies 47. LAUGHLIN JS, MAMOCOS JP, TILBURY RS: Isochronous of dopamine receptors. Brain Res 192:291-295, 1980 cyclotron installation for radionuclide production. Radiology 60. BAUDRY M, MARTRES MP, SCHWARTZ JC: In vivo bind 93:331-337,1969 ing of 3H-pimozide in mouse striatum: Effects of dopamine 48. TILBURY RS, DAHL JR, MAMOCOS JP, et al: Fluorine-18 agonists and antagonists. Life Sci 21:1163-1170, 1977 production for medical use by helium-3 bombardment of 61. GIVANT Y, SHANI (MISHKINSKY) J, GOLDHABER G, et water. Inter J Appi Rad Isotopes 21:277-281, 1970 al: Pharmacology of three mammotropic butyrophenones in 49. GLOWINSKI J, IVERSEN LL: Regional studies of catechol- the rat. Arch Int Pharmacodyn 205:317-327, 1973 amines in the rat brain—I. The disposition of [3H]norepi- 62. VINCENT SH, SHAMBHU MV, DIGENIS GA: Synthesis of nephrine, [3H]dopamine, and [3H]dopa in various regions of [82Br]bromperidol and preliminary tissue distribution studies the brain. J Neurochem 13:655-669, 1966 in the rat. J Med Chem 23:75-79, 1980 50. WOODARD HQ, BIGLER RE, FREED B, et al: Expression of 63. FOWLER JS, ARNETT CD, WOLF AP, et al: [' 'C]Spirop- tissue isotope distribution. J NucíMed 16:958-959, 1975 eridol: Synthesis, specific activity determination, and bio- 5/. SOUDIJN W, VAN WIJNGAARDEN I, ALLEWIJN G: Dis distribution in mice. J NucíMed 23:437-445, 1982 tribution, excretion and metabolism of neuroleptics of the 64. LEYSEN JE, GOMMEREN W, LADURON PM: Spiperone: butyrophenone type. Part I. Excretion and metabolism of A ligand of choice for neuroleptic receptors. 1. Kinetics and haloperidol and nine related butyrophenone-derivatives in the characteristics of in vitro binding. Biochem Pharmacol 27: Wistar rat. Eur J Pharmacol 1:47-57, 1967 307-316, 1978 52. FORSMAN A, ÖHMAN R: Pharmacokinetic studies on ha 65. MARTRES MP, BAUDRY M, SCHWARTZ JC: Character loperidol in man. Curr Ther Res 20:319-336, 1976 ization of 3H-domperidone binding on striatal dopamine re 53. DIGENIS GA, VINCENT SH, KOOK CS, et al: Tissue distri ceptors. Life Sci 23:1781 -1784, 1978 bution studies of [18F]haloperidol, [l8F]-|8-(4-fluorobenzoyl) 66. LADURON PM, LEYSEN JE: Domperidone, a specific in vitro propionic acid, and [82Br]bromperidol by external scintig- dopamine antagonist, devoid of in vivo central dopaminergic raphy. J Pharm Sci 70:985-989, 1981 activity. Biochem Pharmacol 28:2161-2165, 1979 54. SH ARPE AG: The physical properties of the carbon-fluorine 67. BAKHLE YS, VANE JR: Pharmacokinetic function of the bond. In Ciba Foundation Symposium on Carbon-Fluorine pulmonary circulation. Physiol Rev 54:1007-1045, 1974 Compounds: Biochemistry, and Biological Activities, New 68. BROWN EA: The localization, metabolism, and effects of York, Associated Scientific Publishers, 1972, pp 33-54 drugs and toxicants in lung. Drug Metab Rev 3:33-87, 55. MORRISON RT, BOYD RN: Organic Chemistry, 3rd ed, 1974 Boston, Allyn and Bacon, Ine, 1973, pp 701-703 69. TEWSON TJ, WELCH MJ: Preparation of fluorine-18 aryl 56. SNYDER SH, BURT DR, CREESE I: The dopamine receptor fluorides: Piperidyl triazenes as a source of diazonium salts. of the mammalian brain: Demonstration of binding to agonist J Chem Soc Chem Comm 1149-1150, 1979

Hawaii Chapter Society of Nuclear Medicine 6th Annual Meeting May 28-30, 1983 Hyatt Kuilima Hotel Kahuku, Oahu, Hawaii The Hawaii Chapter of the Society of Nuclear Medicine will hold its 6th Annual Meeting on May 28-30, 1983at the Hyatt Kuilima Hotel located on Oahu's magnificent north shore. Guest speakersfor this year's meeting are Wil Nelp, M.D., ErnestGarcia, Ph.D.,Philip Matin, M.D.and Michael Kipper,M.D.

Topicsto be addressed at this Memorial DayWeekendconference include clinical and technical aspects of rotationaltomog raphy,quantification of thallium imaging, uses of monoclonal antibodies, pulmonary, bone and ln-111oxine labeled WBC imaging. Continuing Education and VOICE credits will be available for participants. For further information contact: Patrick McGuigan The Honolulu Medical Group Dept. of Nuclear Medicine 550 South Beretania Street Honolulu, Hawaii 96813

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