BASIC SCIENCES

RADIOCHEMISTRY AND

Hepatic Clearance Mechanism of Tc-99m-HIDA and Its Effect on Quantltation of Hepatobiliary Function: Concise Communication

Elizabeth Harvey, Michael Loberg, James Ryan*, Steven Sikorski, William Faith, and Malcolm Cooper*

University of Maryland, Baltimore, Maryland

Parameters affecting the hepatobiliary clearance of Tc-99m N(2,6-dimethylphenyl carbamoylmethyl) iminodiacetic acid (Tc-HIDA) were evaluated in dogs. Competitive clearance studies were performed with Tc-HIDA after infusion to plasma saturation levels of an anion, sodium sulfobromophthalein (BSP), and a cation, oxyphenonium. The results demonstrated that Tc-HIDA is transported through hepatocytes by a carrier mediated organic-anion pathway. The data are consistent with an alteration of the elimination kinetics of Tc-HIDA induced by elevations in the serum biirubin level, and it is predicted that serum biirubin at some increased concentration will dominate the distribution and elimination kinetics of Tc-HIDA independently of hepatobiiary status. A quantitative description ofliverfunction in terms ofregional distribution and elimination rate constants will require either a pharmacokinetic model that expressly includes the effects of biirubin, the development of new anionic hepatobiiary agents capable of displacing endogenous biirubin from transport binding sites, or the devel opment of new hepatobiiary agents that use a different clearance mechanism from that used by bilirubin.

J NucI Med 20: 310—313, 1979

Technetium-99m N(2,6-dimethylphenylcarba ditional clinical goal for this new agent is the quan moylmethyl) iminodiacetic acid (Tc-HIDA) has titation of certain aspects of function through been developed recently as an intended replace the development of a pharmacokinetic model that ment for 1-131 rose bengal (RB) (1,2) and has dem relates Tc-HIDA's in vivo time-activity curves to onstrated superior kinetic properties for hepatobi regional distribution and elimination rate constants. liary imaging (3). Its value in the clinical evaluation For a pharmacokinetic approach to the quantifica of jaundice in man was shown by Ryan et al. (4) tion of liver function to be successful, the in vivo through the use of sequential . An ad distribution of the should be solely a function of a partient's hepatocellular sta tus. Of the four known carrier-mediated mecha Received Feb. 24, 1978;revisionacceptedNov. 21, 1978. nisms of hepatobiliary clearance, the general an

;@ Present address: Dept. of Radiology, University of Chicago, ionic clearance mechanism used by bilirubin is used Chicago,IL 60637. by most, if not all, of the existing radiotracers (5,6). For reprints contact: Michael D. Loberg, Division of Nuclear , University of Maryland Hospital, 22 S. Greene St., This suggests that in the jaundiced patient bilirubin Baltimore, MD 21201. may competitively displace existing radiopharma

310 THE JOURNAL OF BASIC SCIENCES RADIOCHEMISTRYAND RADIOPHARMACEUTICALS ceuticals from carrier-mediated binding sites, and mm infusion of BSP, each animal was administered that at some point the concentration of serum bill a 10-mCi dose of Tc-HIDA (15 mg) mixed with 70 rubin may become the determinant of radiophar @Ciof 1-131 RB. , bile, and urine were col maceutical distribution and elimination kinetics in lected at the same time intervals as in the first phase dependently of other clinical parameters. To of the experiment. determine the mechanism of the hepatobiliary clear All fluid samples were counted in a NaI(T1) well ance of Tc-HIDA, competitive clearance studies counter with separate counting windows for Tc-99m were performed in dogs with an anion, sulfobrom and 1-131. The radioactivity in each window was ophthalein, and a cation, oxyphenonium. corrected for residual activity from the previous study, and the Tc-99m counts corrected for cross over from 1-131. The counts were compared with METHODS those from a known aliquot of the injectate. The Anionic clearance mechanism. In three female dogs canine blood volume was calculated at 7% of body the distribution and elimination kinetics of I-131RB weight. and Tc-HIDA were simultaneously evaluated be Cation clearance mechanism. The tracer cation, fore and after inducing plasma saturation levels of N([1@C]methyl) lidocaine, was synthesized by the sulfobromophthalein (BSP) (7). The animal model reaction of ‘@CH3Iwith lidocaine (9) and was used was prepared for blood, urine, and bile collection to monitor saturation of the organic cation pathway after i.v. administration ofpentobarbital (30 mg/kg): (/0). The cation clearance mechanism was studied a Cournand needle was inserted upstream into a in six female dogs that were prepared for serial bile femoral artery for blood collection; a Foley catheter collection as previously described except that the was inserted into the bladder; and a midline abdom renal pedicles were ligated to prevent urinary cx inal incision (approximately 15 cm) was made so cretion. Three of these dogs were used for a base that the and biliary tree could be iso line kinetic study of blood and bile levels of lated. The cystic duct was ligated to prevent bile N([―C]methyl)lidocaine. In the other three dogs the flow into the gallbladder. A polyethylene catheter same studies were done after the administration of was introduced through a small incision into the oxyphenonium—a known saturator of the cationic common bile duct and advanced to the junction of pathway (11)—and saturation levels of oxyphen the two hepatic ducts to ensure total bile drainage onium were obtained by i.v. administration of 7.9 into the catheter. mg/kg body weight over a 5-mm period. After 15 After preparation of the basic model, each animal mm Tc-HIDA (1 mCi) and N(['4 C]methyl)lidocaine was given 1 mCi of Tc-HIDA intravenously into a (0.26 —0.82pCi) were injected simultaneously and front cephalic vein. This injection also contained 7 plasma and bile samples were collected over a 2-hr @Ciof 1-131 RB to ensure that initial kinetic values period. The samples from both groups of animals would correlate with literature values for the blood were prepared and counted according to a previ clearance and biliary uptake of RB. One-cubic cen ously published method (12). timeter blood samples were collected at 5, 10, 15, 30, 45, and 60 mm postinjection. Cumulative bile RESULTS samples of known volume were collected at four Table 1 shows the effect of i.v. infusion of BSP 15-mm intervals, and a cumulative urine sample and oxyphenonium on the hepatobiliary clearance was obtained 1 hr postinjection. At 1½hr postin of Tc-HIDA in dogs. A mean cumulative total of jection, blood, bile, and urine samples were again 55.2% ofTc-HIDA was excreted into the bile before taken to ascertain baseline values for the second part of the experiment. At this point, each animal received a 50-mg bolus injection of BSP followed TABLE 1. THE EFFECT OF BSP AND by an i.v. infusion of BSP in saline (7.5 mg/ml), OXYPHENONIUM INFUSION ON BILIARY EXCRETIONOFTc-HIDAIN DOGS* which continued throughout the remainder of the experiment at a rate of 1 ml per minute. Blood (1-hr cum . values) (2-hr cum. values) samples were taken before infusion and ½.hrpost infusion and were analyzed for plasma BSP levels DogBAfter142.50.590.03BeforeSP AfterOxyphenoniumBefore and for the percentage of Tc-HIDA that was protein 92.03265.21 bound. Sodium sulfobromophthalein levels were 92.94358.02.388.22 .684.36 73.9855.21.587.50 determined using the standard colorimetric method (8). The extent of protein binding was measured by Mean 86.30 addition of Tc-HIDA to the plasma samples fol * Percentage of injected dose of Tc-HIDA in bile. lowed by equilibrium dialysis for 18 hr. After a 30-

Volume 20, Number 4 311 HARVEY, LOBERG, RYAN, SIKORSKI, FAITH, AND COOPER

BSP infusion, compared with only 1.5% after BSP Since serum bilirubin levels may be elevated infusion. Additional results, not included in Table through several mechanisms in a variety of 1, showed that the BSP infusion reduced the RB states, competition between Tc-HIDA and bilirubin hepatobiliary excretion from 20% to 6% and in in the transport pathway will complicate any phar creased the 1-hr urinary clearance of Tc-HIDA macokinetic method seeking to quantitate an aspect from 13.3% to 44.0%. Before BSP infusion, 2.7% of liver function. Approaches to the quantitation of of Tc-HIDA and 8.8% of RB remained in the blood liver function, therefore, should expressly include at 30 mm, and after infusion these values were the effects of bilirubin in the model; or, perhaps elevated to 8.6% and 40%, respectively. Postinfu more practically, clinical studies should be under sion serum BSP levels were always greater than 10 taken to determine the serum level at which bili mg/dl , substantially exceeding the 4 mg/dl value rubin dominates Tc-HIDA's in vivo clearance in quoted by Wheeler (7) as sufficient to saturate the man. At some serum bilirubin level the model-de carrier-mediated clearance of BSP. These satura rived rate constants will be more indicative of corn tion levels of BSP were found to exert a minimal petition with bilirubin than of the patient's hepâto effect on the protein binding of Tc-HIDA, decreas bilibary status. Ideally, new radiopharmaceuticals ing it on the average from 64% initially to 56% deSigned to measure hepatobiliary function should postinfusion. The BSP levels achieved in these possess in vivo clearance patterns independent of studies are capable of drastically altering the elim serum bilirubin levels. This could be accomplished ination pattern of Tc-HIDA without significant through the use of either radiolabeled cations or competitive displacement of Tc-HIDA from its pro bile salts or through the development of new lipo tein binding sites. philic, anionic agents that compete more effectively In contrast, saturation levels of the cation oxy with bilirubin for hepatobiliary clearance than do phenonium had no effect on the hepatobiliary clear either Tc-HIDA or I-l3lRB. ance of Tc-HIDA. Ligation of the renal pedicles The difficulties in quantitating liver function resulted in an increase in the hepatobiliary clear using Tc-HIDA and other anionic hepatobiliary ance of Tc-HIDA from 55.2% to 87.5%, and of agents do not preclude their use in the clinical as N(['@C]methyl)Iidocaine from 2% to 11% before ox sessment of the jaundiced patient. Ryan et al (4) yphenonium infusion. Table 1 shows that Tc suggest that larger doses of Tc-HIDA should be HIDA's biliary excretion was not changed by in administered to patients with elevated serum bill fusion of oxyphenonium, whereas the hepatobiliary rubin levels. The increased dose appears necessary clearance of N([―C]methyl)lidocaine decreased in order to keep relatively constant the millicune from 11% to 2% under identical conditions. quantity of Tc-HIDA entering the small intestine. Elevated serum bilirubin levels appear to delay the DISCUSSION blood clearance of Tc-HIDA and to extend the in The inhibition of Tc-HIDA's hepatobiliary clear terval after administration during which sequential ance by BSP demonstrated that Tc-HIDA is elimi scintiphotos must be obtained. It has been shown, nated through the liver's general anionic clearance however, that although the competition between pathway (5), and that this pathway is affected by a Tc-HIDA and bilirubin increases the background competing cation, oxyphenonium. Since both BSP to nontarget tissues, it does not preclude and bilirubin are known to utilize this general an imaging of the gallbladder up to biirubin levels of ionic clearance mechanism (5), it follows that hy 10 mg/dl nor does it preclude imaging of intestinal perbilirubinemia will also impede the clearance of activity 18 hr postinjection if the biliary tract is Tc-HIDA by the liver. patent. The latter is the major determinant of din Technetium-HIDA' s observed carrier-mediated ical significance in severely jaundiced patients. anionic clearance mechanism is consistent with However, the use of Tc-99m as the radiotracer is either facilitated diffusion or active-transport inconvenient in these circumstances, since a long clearance mechanisms and is also in agreement with imaging time (½to 1½hr) is frequently required at previous data (13) indicating that Tc-HIDA exists 18 hr postinjection in order to visualize clearly the as an anionic, bis structure with substantial lipo small amount of Tc-99m activity present in the in philicity and a molecular weight of approximately testinal tract, even when a substantial dose of Tc 683. These molecular features have been associated 99m is injected initially. Hepatobiiary tracers with with rapid hepatobiliary clearance (14). Note that significantly longer physical half-lives than 6 hr may the anions, HIDA and Sn-@HIDA,do not competi be clinically useful in this situation, since imaging tively inhibit the hepatobiliary clearance of Tc studies may be completed more rapidly at 18 hr and HIDA, since their lesser lipophilic quality has been also may be performed at significantly longer inter shown to favor renal clearance (4). vals postinjection than with Tc-99m agents. An

312 THE JOURNAL OF NUCLEAR MEDICINE BASICSCIENCES AND RADIOPHARMACEUTICALS

other approach to hepatobiliary imaging in the jaun An evaluation of @9@c-labeledhepatobiliary agents. J Nuc! diced patient would be the development of a Tc Med 18:455-461,1977 99m hepatocyte imaging agent that does not com 4. RYAN J, COOPERM, LOBERGM, et al: Technetium-99m- labeled N-(2,6-dimethylphenylcarbamoylmethyl) iminodi pete with serum bilirubin for hepatic clearance. Our acetic acid (Tc-99m HIDA): A new radiopharmaceutical for results suggest that imaging studies with this tracer hepatobiliary imaging studies. J Nuc! Med 18: 997-1004, in the jaundiced patient could be performed with a 1977 significantly smaller dose of injected radioactivity 5. JAVITT NB: Hepatic bile formation II. N. Engi N Med 295: than is currently used and also could be completed 1511—1516,1976 6. LOBERG MD Radiopharmaceutical distribution by active in one day, even in the patient with significantly transport: Implications of non-linear kinetics. Proceedings elevated serum bilirubin levels. of the 1st International Symposium on Radiopharmacology: At present it appears that sequential scintigraphic inpress data can provide more clinical information in the 7. WHEELER HO, EPSTEIN RM, ROBINSON RR, et al: Hepatic jaundiced patient than is readily obtainable through storage and excretion of sulfobromophthalein sodium in the dog. J C/in Invest 39: 236-247, 1960 simple pharmacokinetic models and that the quan 8. TIETZN: Fundamentals ofC/inica/ Chemistry. Philadelphia, titation of hepatobiliary function should be re W. B. Saunders Co., 1970, pp. 786-787 stricted to patients with no more than modest ele 9. FAITH WC: M.S. Thesis, University of Maryland, Balti vations of serum bilirubin. A more valid quantitation more, Maryland, 1976 of hepatobiliary function may require new hepato 10. SCHANKERLS, SOLOMONHM Active transport of quater nary ammonium compounds into bile. Am J Physiol 204: biliary agents that either compete more effectively 829-832, 1963 with serum bilirubin for transport binding sites or 11. LEVINE RM, CLARK BB: The physiological disposition of else employ totally different clearance mechanisms. oxyphenonium bromide (Antrenyl) and related compounds. J Pharmacol Exp Ther 121: 63-70, 1957 12. CALLERY PS, FAITH WC, LOBERG MD, et at: Tissue distri bution of technetium-99m and carbon-l4 labeled N-(2,6-di REFERENCES methylphenylcarbamoylmethyl) iminodiacetic acid. J Med 1. LOBERGMD, COOPERM, HARVEYE, et al: Development of Chem 19: 962-964, 1976 new radiopharmaceuticals based on N-substitution of imi 13. Chemical structure of technetium-99m labeled N-(2,6-di nodiacetic acid. J Nuci Med 17: 633-638, 1976 methylphenylcarbamoyl-methyl) iminodiacetic acid (Tc 2. HARVEYE, LOBERGM, COOPERM Tc-99m HIDA: A new HIDA). in: J AppI Radiat: 29: 167-173, 1978 radiopharmaceutical for hepato-biliary imaging. J Nuci Med 14. NIELSONP, RASMUSSENF: Relationships between molecu 16: 533, 1975 (abst) lar structure and excretion of . Life Science 17: 1495- 3. WIST0w BW, SUBRAMANIANG, VAN HEERTUMRL, et al: 1512,1975

MISSOURI VALLEYCHAPTER SOCIETY OF NUCLEAR MEDICINE FALL MEETING

October 12—i4, 1979 Hilton Inn Des Moines, Iowa Abstractsof original papers are invited for presentationat the annual meeting of the Missouri Valley Chapter of the Society of Nuclear Medicine. Papers are to be 10 minutes. Pleasesubmit200word abstractbeforeAugust1, 1979to: DavidF. Preston,M.D.,Division of NuclearMedicine,Departmentof Radiology,KansasUniversityMedicalCenter,39thand RainbowBlvd.,KansasCity,Kansas66103.

For more information contact: @ KarenStuyvesant @ I IowaDepartmentMethodistof NuclearMedicalMedicineCenter 1 1200PleasantStreet I DesMoines,Iowa50308 Telephone: (515)283-6458

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