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Applied Radiation and Isotopes 65 (2007) 301–308

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On the preparation of a therapeutic dose of 177Lu-labeledDOTA–TATE using indigenously produced 177Lu in medium

flux reactor

Tapas Das, Sudipta Chakraborty, Sharmila Banerjee�, Meera Venkatesh

Radiopharmaceuticals Division, Bhabha Atomic Research Centre, Mumbai-400085, India

Received 26 June 2006; received in revised form 1 September 2006; accepted 29 September 2006

Abstract

177Lu could be produced with a specific activity of �23,000mCi/mg (850GBq/mg) by neutron activation using enriched 176Lu (64.3%)

target when irradiation was carried out at a thermal neutron flux of 1� 1014 n/cm2/s for 21 d. 177Lu–DOTA–TATE could be prepared in

high radiochemical yield (�99%) and adequate stability using the 177Lu produced indigenously. The average level of radionuclidic

impurity burden in 177Lu due to 177mLu was found to be 250 nCi of 177mLu/1mCi of 177Lu (9.25 kBq/37MBq) at the end of

bombardment, which corresponds to 0.025% of the total activity produced. The maximum specific activity achievable via careful

optimization of the irradiation parameters was found to be adequate for the preparation of a therapeutic dose of the

radiopharmaceutical. The in-house preparation of this agent using 25mg (17.41 nmole) of DOTA–TATE and indigenously produced177Lu (0.8 mg, 4.52 nmole), corresponding to peptide/Lu ratio of 3.85 yielded 98.7% complexation. Allowing possibility of decay due to

transportation to users, it has been possible to demonstrate that at our end, a single patient dose of 150–200mCi (5.55–7.40GBq) can be

prepared by using 250–333mg of DOTA–TATE conjugate. This amount compares well with 177Lu–DOTA–TATE prepared for a typical

peptide receptor radionuclide therapy (PRRT) procedure which makes use of 100mg of the DOTA–TATE conjugate, which incorporates

50mCi (1.85GBq) of 177Lu activity, thereby implying that in order to achieve a single patient dose of 150–200mCi (5.55–7.40GBq),

300–400 mg of the conjugate needs to be used.

r 2006 Elsevier Ltd. All rights reserved.

Keywords: PRRT; 177Lu; DOTA–TATE; Somatostatin receptors

1. Introduction

Radiometallated peptides which exhibit high specificityfor cognate receptors over-expressed on cancerous lesions,offer important potential as site-directed diagnostic andtherapeutic radiopharmaceuticals (Boerman et al., 2000;Breeman et al., 2001; Britton, 1997; Eckelman and Gibson,1993). The highly specific biochemical and physiologicbinding capabilities of the receptor-avid peptides can beexploited for using these agents as a vehicle to target thedelivery of the radioactivity to the cells over-expressingsimilar type of receptors after radiolabeling the peptideswith the radionuclide of interest. Such peptide analogs

e front matter r 2006 Elsevier Ltd. All rights reserved.

radiso.2006.09.011

ing author. Tel.: +91 22 2559 0616; fax: +91 22 2550 5345.

ess: sharmila@barc.gov.in (S. Banerjee).

when labeled with therapeutic radionuclides are beingactively investigated as agents for use in peptide receptorradionuclide therapy (PRRT) (Kwekkeboom et al., 2000,2003a, 2005; de Jong et al., 2005).Among the several classes of radiolabeled peptides,

radiolabeled somatostatin analogs have been proved to bethe most promising. Octreotide, a metabolically stableanalog of native somatostatin (Pauwels et al., 1998), hasbeen radiolabeled with 111In using a bifunctional chelatingagent and the resultant radiolabeled peptide, 111In–DTPA-Octreotide (commonly known as OctreoScans) has beensuccessfully employed in clinical diagnosis of somatostatinreceptor positive tumors (Pauwels et al., 1998; Krenninget al. 1992, 1993). Efforts are being made to develop suitabletherapeutic analogs having the potential to eradicate thecancerous lesions over-expressing somatostation receptors.

ARTICLE IN PRESST. Das et al. / Applied Radiation and Isotopes 65 (2007) 301–308302

Recently started clinical trials with (177Lu–DOTA–Tyr3)-Octreotide (known as 177Lu–DOTA–TOC) and (177Lu–DOTA–Tyr3)-Octreotate (known as 177Lu–DOTA–TATE)have shown very encouraging results in terms of tumorregression in patients suffering from different types ofneuroendocrine tumors, which are known to over-expresssomatostatin receptors (Kwekkeboom et al., 2003a, b,2005; Forrer et al. 2005). However, it is reported that[DOTA–Tyr3]-Octreotate has a ninefold higher affinity forthe somatostatin receptor subtype 2 as compared with[DOTA–Tyr3]-Octreotide and therefore, the later agent isexpected to be more potent for carrying out PRRT in patientssuffering from neuroendocrine tumors (Kwekkeboom et al.,2003a; Reubi et al. 2000).

177Lu is presently being considered as a potentialradionuclide for use in in vivo therapy, because of itsfavorable decay characteristics. 177Lu decays with a half-life of 6.73 d by emission of b- particles with maximumenergies of 497 keV (78.6%), 384 keV (9.1%), and 176 keV(12.2%) to stable 177Hf (Firestone, 1996). The emission ofsuitable energy g photons of 113 keV (6.4%) and 208 keV(11%) (Firestone, 1996) with relatively low abundancesprovides the opportunity to carry out simultaneousscintigraphic studies, which helps to monitor the properin vivo localization of the injected radiopharmaceutical aswell as to perform dosimetric evaluations. An importantaspect for the countries with limited reactor facility is thecomparatively long half-life of 177Lu which provideslogistic advantage towards facilitating supply to locationsfar away from the reactors (Das et al., 2002). Besides this,the high thermal neutron capture cross-section (s ¼ 2100b)of (176Lu(n,g)177Lu) reaction (Firestone, 1996) ensures that177Lu can be produced with sufficiently high specificactivity using the moderate flux reactors. In fact, thecross-section is the highest encountered among all (n,g)produced radionuclides presently used for therapy. Thesefavorable nuclear parameters ensure that there will be noconstraints with respect to large-scale isotope production.

In connection with our ongoing research on thedevelopment of novel agents for radiotherapy of tumors(Banerjee et al., 2004; Das et al., 2004), production of177Lu, identified as the ideal radioisotope is beingextensively optimized with an aim to achieving themaximum specific activity via a cost-effective route.This is an essential requirement for designing agents forPRRT since the primary criterion is to deliver sufficientnumber of radionuclides to the receptors over-expressed onthe targeted tumor site without saturating the target(Mausner and Srivastava, 1993; Volkert and Hoffman,1999; Breeman et al., 2003).

177Lu can be produced by two different routes, namely,by irradiation of Lu target and by irradiation of Yb targetfollowed by radiochemical separation of 177Lu from Ybisotopes. Though the latter method enables production ofno-carrier-added (NCA) 177Lu, the radiochemical separa-tion of 177Lu activity from irradiated Yb target is a difficulttask owing to the similarity in the chemistry of the two

adjacent members of the lanthanide series. Presence of Ybin substantial amounts, attributable to its low cross-section, besides reducing the effective specific activity ofthe product, will also compete with 177Lu in the complexa-tion procedure unless effectively eliminated from theirradiated target. This is one of the major impediment inthe ready production of NCA 177Lu via 176Yb(n,g,b�)177Luroute. Moreover, use of enriched targets with low activationcross-section (s ¼ 2.4b for 176Yb(n,g)177Yb) (Firestone,1996) is not economical for isotope production, as asignificant part of the target remains unutilized. Though,176Yb recovery is theoretically feasible, the aforementionedpractical problems associated with the cumbersome separa-tion still remains.Our experiences in the production of 177Lu have

provided an insight towards envisaging 177Lu–DOTA–TATE as a radiopharmaceutical, which can be indigen-ously produced and supplied to local users as an agent forPRRT. The primary aim in the designing of this agentattempts to maximize the specific activity of the resultantradiolabeled preparation. Towards this, 177Lu productionis envisaged via (n,g) using an enriched Lu2O3 target andefforts are directed to obtain 177Lu with maximum possiblespecific activity utilizing the moderate flux reactor availablein our country. However, careful optimization of theirradiation parameters is required for obtaining 177Lu inadequate specific activity for PRRT applications, particu-larly when production is envisaged at high flux positionsowing to the high thermal neutron capture cross-section of176Lu which eventually leads to considerable target burnup. The present paper describes optimization of 177Luproduction with sufficiently high specific activity using amoderate flux reactor and successful preparation of patientdose of 177Lu–DOTA–TATE using the 177Lu indigenouslyproduced at our end.

2. Materials and methods

Lutetium oxide (64.3% enriched in 176Lu, spectroscopicgrade, 499.99% pure) was obtained from Isoflex, Russia.DOTA–TATE was obtained from PiChem, Finland,through International Atomic Energy Agency (IAEA) asa part of a coordinated research project (CRP). Allchemicals and solvents used in the experiments were ofAR grade and supplied by reputed chemical manufac-turers. Radionuclidic purity of 177Lu was ascertained byhigh resolution g-ray spectrometry using an HPGe detector(EGG Ortec/Canberra detector) coupled to a 4K multi-channel analyzer (MCA) system after radiochemicalprocessing. 152Eu reference source used for energy andefficiency calibration of the detector was obtained fromAmersham Inc., USA. All other radioactivity measure-ments were carried out using a well type NaI (Tl)scintillation counter after adjusting the 150 keV baselineand keeping a window of 100 keV, thereby utilizing the208 keV g photon of 177Lu. Whatman 3mm chromato-graphy paper (UK) was used for paper chromatography

ARTICLE IN PRESST. Das et al. / Applied Radiation and Isotopes 65 (2007) 301–308 303

studies. The high performance liquid chromatography(HPLC) system used was obtained from JASCO (PU1580), Japan, equipped with a PU 1575 UV/VIS detector.A well-type NaI (Tl) scintillation detector was coupled tothe system for radioactivity measurements in the eluate. Allthe solvents used for HPLC analyses were of HPLC gradeand purchased from reputed local manufacturers, degassedand filtered prior to use.

2.1. Production and radiochemical processing of 177Lu

177Lu was produced by thermal neutron bombardmenton isotopcially enriched (64.3% in 176Lu) Lu2O3 target. Astock solution of enriched target was prepared by dissol-ving enriched Lu2O3 powder in 0.1M HCl (1mg/mLconcentration). A known aliquot of this solution was takenin a quartz ampoule and carefully evaporated to dryness.The ampoule was subsequently flame sealed and irradiatedafter placing inside an aluminum can. Irradiations werecarried out at different available flux positions (1.4�1013–1.0� 1014 n/cm2/s) for different durations (7–21 d) inorder to optimize the irradiation conditions towardsachieving maximum specific activity of 177Lu. The irra-diated target was dissolved in 1M HCl by gentle warminginside a lead-shielded plant. The resultant solution wasevaporated to near dryness and reconstituted in de-ionizedwater. The pH of the 177LuCl3 solution thereby obtainedwas adjusted to�4 prior to complexation studies. A knownaliquot was drawn for assay of radioactivity and determi-nation of radionuclidic purity.

Radioactivity assay was carried out by measuring theionization current obtained when an aliquot of the batchwas placed inside a pre-calibrated well-type ion-chamber.Radionuclidic purity was determined by recording g-rayspectra of the appropriately diluted solution of theirradiated target using an HPGe detector connected to a4K MCA system. Energy as well as efficiency calibration ofthe detector were carried out using a 152Eu reference sourceprior to the recording of g-ray spectra. Several spectra wererecorded for each batch at regular time intervals. Samplesmeasured initially for the assay of 177Lu were preserved forcomplete decay of 177Lu (8–10 T1/2 of

177Lu, i.e. for a periodof 50–70d) and re-assayed to determine the activity of long-lived 177mLu (T1/2 ¼ 160.5 d). Appropriately diluted samplesolutions were counted for 1 h.

2.2. Preparation of 177Lu–DOTA–TATE complex

For the preparation of 177Lu-labeled DOTA–TATEcomplex, a stock solution of the DOTA–TATE was preparedby dissolving DOTA–TATE conjugate in HPLC grade waterwith a concentration of 1mg/mL. To a 25mL aliquot of thisstock solution, 25mL of 177LuCl3 solution was added and thevolume was made upto 200mL using the 0.1M ammoniumacetate buffer of pH 5. The reaction mixture was incubatedat 80 1C for 1h after adjusting the pH to 4.5–5. Variousparameters, such as, ligand concentration, incubation time,

and temperature were varied extensively in order to arrive atthe protocol for maximum complexation.

2.3. Quality control techniques

The characterization of the 177Lu-labeled DOTA–TATEconjugate as well as the determination of the complexationyield was carried out by paper chromatography (PC) aswell as by HPLC techniques.

2.3.1. Paper chromatography (PC)

5 mL portions of the test solutions were applied at 1.5 cmfrom the lower end of the chromatography paper strips.The strips were developed in 50% aqueous acetonitrile,dried, cut into segments of 1 cm each and the radioactivityassociated with each segment was measured in a NaI (Tl)detector.

2.3.2. High performance liquid chromatography (HPLC)

HPLC of the 177Lu-labeled conjugate was carried outusing a dual pump HPLC unit with a C-18 reversed phaseHiQ-Sil (5 mm, 25� 0.46 cm) column. The elution wasmonitored both by detecting UV signals at 270 nm as wellas by radioactivity signal using NaI (Tl) detector. Water(A), and acetonitrile (B) mixtures with 0.1% trifluoroaceticacid were used as the mobile phase and the followinggradient elution technique was adopted for the separation(0–4min 95% A, 4–15min 95% A to 5% A, 15–20min 5%A, 20–25min 5% A to 95% A, 25–30min 95% A). Flowrate was maintained at 1mL/min.

2.4. Maximization of specific activity of177Lu–DOTA–TATE complex

In order to obtain 177Lu-labeled DOTA–TATE con-jugate in highest possible specific activity, experiments werecarried out to achieve maximum complexation yield of177Lu-DOTA–TATE at the minimum possible [L]/[M]ratio. For this, 25 mg of the peptide (already optimizedligand amount) was allowed to react with differentamounts of Lu ranging from 0.2 to 1.6 mg under the samereaction conditions described earlier.

2.5. Stability of 177Lu–DOTA–TATE

The in vitro stability of the 177Lu-labeled conjugate wasascertained by storing the radiolabeled product at roomtemperature and determining the radiochemical purity atdifferent time intervals post-preparation employing thestandard quality control techniques described earlier.

3. Results and discussions

3.1. Production of 177Lu

177Lu was produced by thermal neutron bombardmenton isotopically enriched (64.3% in 176Lu) Lu2O3 target in

ARTICLE IN PRESST. Das et al. / Applied Radiation and Isotopes 65 (2007) 301–308304

Dhruva reactor at our Institute. The radionuclidic purity of177Lu produced was determined by analyzing the g-rayspectrum and found to be 99.975%. A typical g-rayspectrum recorded after the radiochemical processing ofthe irradiated target exhibited major g photopeaks at 72,113, 208, 250, and 321 keV. All of these photopeakscorrespond to that of 177Lu (Firestone, 1996). This wasfurther confirmed from the decay as followed by monitor-ing peak area cps values at those peaks according to thehalf-life of 177Lu. It is well documented in the literaturethat there is a possibility of formation of 177mLu (T1/2

¼ 160.5 d) on thermal neutron bombardment of Lu target(Neves et al., 2002; Pillai et al., 2003; Nir-El, 2004).However, g-ray spectrum of the irradiated Lu target afterchemical processing did not show any significant peakcorresponding to the photopeaks of 177mLu (128, 153, 228,378, 414, 418 keV) (Firestone, 1996). This can be explainedfrom the fact that the radioactivity due to 177mLu producedwill be insignificant compared to that of 177Lu, owing to itslong half-life and comparatively low cross-section(s ¼ 7 barns) (Firestone, 1996) for its formation. The tracelevel of 177mLu produced was determined by recording theg-ray spectrum of a sample aliquot, initially having highradioactive concentration, after complete decay of 177Luactivity (8–10 T1/2 of 177Lu, i.e. for a period of 50–70 d).The average level of radionuclidic impurity burden in 177Ludue to 177mLu determined by this technique was found tobe 250 nCi of 177mLu/1mCi of 177Lu (9.25 kBq/37MBq) atEOB, which corresponds to 0.025% of the total activityproduced.

In order to produce 177Lu with maximum possiblespecific activity using the moderate flux reactor available atour end, Lu2O3 target was irradiated at different availableflux positions for different periods of time. Four differentflux positions with thermal neutron flux ranging between1.4� 1013 and 1.0� 1014 n/cm2/s were available for irradia-tion. Since the normal irradiation schedule of this multi-facility reactor allowed irradiation of the target only in

Table 1

Specific activity of 177Lu obtained at different thermal neutron flux for irradia

Neutron flux (n/cm2/s) Irradiation time (d) Activity obtaine

(mCi/mg) (GBq

1.4� 1013 7 23227102

85.973.8

3.0� 1013 7 46357269

171.579.9

3.0� 1013 14 74607253

276.079.4

6.75� 1013 7 110007782

407.0728.9

6.75� 1013 14 1775071476

656.8754.6

1.0� 1014 21 231857658

857.8724.3

multiples of 7 d, irradiations were carried out for 7, 14, and21 d. The specific activity of the 177Lu obtained onirradiation of enriched target for varied time durationsand at different thermal neutron flux positions aretabulated in Table 1. A maximum specific activity of�23000mCi/mg (850GBq/mg) was achieved when irradia-tion was carried out at a thermal neutron flux of 1� 1014 n/cm2/s for 21 d, which corresponds to �21% of themaximum achievable specific activity. It is evident fromTable 1 that the specific activities of 177Lu obtained weresignificantly higher compared to the theoretically calcu-lated values accounting for only thermal neutron capture.A possible reason for practically obtaining an activityhigher than theoretically calculated values could beattributed to the contribution from epithermal neutrons(resonance integral ¼ 1087b), which is not accounted for intheoretical calculations (Pillai et al., 2003; Nir-El, 2004;Knapp et al., 1995). However, the contribution fromepithermal neutron alone could not provide a satisfactoryexplanation for obtaining 2.5–2.8 times higher specificactivity of 177Lu than that of theoretically calculatedvalues.Apart from the use of enriched target and choosing

the highest available neutron flux position in a reactor,the other factor, which can be optimized for achieving themaximum possible specific activity of a radioisotopeproduced by neutron activation, is the duration ofirradiation. In most cases of radioisotope production using(n,g) route, irradiation of the target 4–6 times of the half-life of the radioisotope being produced results in maximumspecific activity. However, the same does not hold good incase of production of 177Lu by neutron capture, particu-larly, when the production is being carried out at acomparatively higher neutron flux position, owing to thehigh thermal neutron capture cross section of 177Lu. In thiscase, a careful optimization of the time of irradiation needsto be considered in order to obtain the highest specificactivity (Pillai et al., 2003). In high flux positions of a

tion of different durations

d

/mg)

Theoretical activity

(mCi/mg) (GBq/mg)

Experimental/theoretical

846 2.74

31.3

1814 2.56

67.1

2696 2.77

99.8

4081 2.70

151.0

6406 2.77

237.0

8622 2.69

319.0

ARTICLE IN PRESST. Das et al. / Applied Radiation and Isotopes 65 (2007) 301–308 305

reactor, the target burn up will be considerably higherowing to the high thermal neutron capture cross sectionof 176Lu and hence, the usual assumption that the numberof target atoms remains constant during the period ofirradiation will not be valid in this case. Considering thefact that the number of target atoms is not fixed but afunction of time, the commonly used differential equationused for the calculation of the activity produced i.e.

dN2=dt ¼ N1sf�N2l (1)

can be modified as

dN2=dt ¼ N0e�sftsf�N2l, (2)

where N0 is the number of 176Lu atoms used as target (att ¼ 0), N1 the number of 176Lu atoms at any time t, N2 thenumber of 177Lu atoms at any time t, l the decay constantof 177Lu, s the thermal neutron capture cross section of176Lu, f the thermal neutron flux of the reactor and t thetime of irradiation. Solution of the above mentionedmodified differential equation leads to the followingmathematical expression, which enables calculation of the177Lu activity (A) produced at EOB, taking into accountthat the number of the target atoms is also a function oftime of irradiation.

A ¼N0lsfl� sf

½e�sft � e�lt� (3)

Fig. 1 shows the variation of the theoretically calculatedspecific activity of 177Lu obtained at EOB with irradiationtime at 1� 1014 n/cm2/s thermal neutron flux. It is evidentthat the activity of 177Lu produced will be maximum after21 d irradiation, beyond which, the activity will decreaseowing to the high target burn up. Therefore, in order toobtain maximum specific activity of 177Lu using themaximum available flux position in our reactor, the timeof irradiation was fixed as 21 d.

28211470

325

300

275

250

225

200

175

150

125

100

Duration of irradiation (d)

Spec

ific

act

ivity

of17

7 Lu

(GB

q/m

g)

Fig. 1. Variation of theoretically calculated specific activity of 177Lu

obtained at EOB with time of irradiation at thermal neutron flux of

1� 1014 n/cm2/s.

3.2. Characterization of 177Lu–DOTA–TATE

In PC using 50% aqueous acetonitrile, the activitycorresponding to 177Lu–DOTA–TATE complex movedtowards the solvent front with Rf ¼ 0.8–0.9, while un-complexed 177Lu remained at the point of spotting (Rf ¼ 0)under identical conditions. The PC patterns of 177LuCl3and 177Lu–DOTA–TATE complex are shown in Fig. 2.The 177Lu-labeled DOTA–TATE complex was also

characterized by HPLC studies. Fig. 3(a) shows the typicalHPLC pattern of 177Lu-labeled DOTA–TATE conjugateprepared under optimized conditions. Fig. 3(b) shows theHPLC pattern of 177LuCl3 under identical conditions. Theactual extent of complexation achieved was also deter-mined from the HPLC studies.

3.3. Optimization of complexation yield of177Lu–DOTA–TATE

Various parameters such as, conjugate concentration,pH of the reaction mixture, reaction time and temperaturewere varied extensively in order to achieve maximumcomplexation yield. Keeping the reaction volume as200 mL, the amount of DOTA–TATE was varied from 5to 100 mg in order to determine the optimum ligandconcentration required for obtaining maximum complexa-tion. It was observed that a minimum of 25 mg DOTA–TATE was required to obtain a complexation yield of99%. The reaction was carried out by incubating thereaction mixture at three different temperatures (roomtemperature, 50 and 80 1C) for different time periods(5min, 15min, 30min, 1 h, and 2 h) in order to determineoptimum reaction time and temperature. It was observedthat, maximum complexation was achieved when thereaction mixture was incubated at 80 1C for a period of1 h. It is well documented in the literature that the

0

100

80

60

40

20

0

Migration from point of spotting (cm)

% R

adio

activ

ity

177Lu-DOTA-TATE

177LuCl3

1 2 3 4 5 6 7

Fig. 2. Paper chromatography patterns of 177LuCl3 and 177Lu–DOTA–

TATE.

ARTICLE IN PRESS

00 300 600 900 1200 1500 1800

200

400

600

800

1000

CP

S

Retention time (s)

0 300 600 900 1200 1500 18000

100

200

300

400

CP

S

Retention time (s)

(a)

(b)

Fig. 3. HPLC patterns of (a) 177Lu–DOTA–TATE and (b) 177LuCl3.

Table 2

Complexation yield of 177Lu–DOTA–TATE at different [L]/[M] ratio

[DOTA–TATE] [Lu] [L]/[M] Complexation

yield (%)

25mg(17.41 nmole)

0.2 mg(1.13 nmole)

15.41 99.2

25mg(17.41 nmole)

0.4 mg(2.26 nmole)

7.70 99.0

25mg(17.41 nmole)

0.8 mg(4.52 nmole)

3.85 98.7

25mg(17.41 nmole)

1.6 mg(9.04 nmole)

1.92 81.2

T. Das et al. / Applied Radiation and Isotopes 65 (2007) 301–308306

maximum complexation is obtained when 177Lu labeling ofthe biomolecules are carried out at pH�5 (Breeman et al.,2003; Milenic et al., 2002; Stimmel and Kull, 1998). In thepresent case, the maximum complexation yield wasachieved at pH 4.5–5.

3.4. Maximization of the specific activity of177Lu–DOTA–TATE

Bearing in mind the importance of preparing radiola-beled agents intended for targeted use in binding withreceptors over-expressed on tumors, efforts were directedto prepare the 177Lu-DOTA–TATE with maximum possi-ble specific activity, using the 177Lu available at our end.Towards achieving this, the concentration of the peptideconjugate used for a typical labeling procedure needs to beminimum which, in turn, would aim at the minimumpossible ligand/metal ratio, in order that unlabeled peptidewhich would otherwise bind also with the targetedreceptors will be minimum. The ligand/metal ratio can beminimized by keeping the concentration of the peptideconstant and gradually increasing the amount of radio-metal in the reaction mixture. Since 25 mg of the peptidewas found to be the optimum amount required forobtaining �99% complexation, the ligand concentration

is kept constant at this value and complexations werecarried out using different amounts of Lu ranging from 0.2to 1.6 mg under the same reaction conditions describedearlier. The complexation yield in each case was deter-mined by PC as well as by HPLC. The complexation yieldsobtained at different concentrations of Lu are given inTable 2. It is evident from Table 2, that a radiolabelingyield of �99% could be achieved using a ligand/metal ratio�4. This implies that 0.8 mg of Lu can be incorporated in25 mg of the peptide without compromising the extent ofcomplexation. Considering the maximum possible specificactivity of 177Lu achievable at our end to be 23,000mCi/mg(850GBq/mg), 0.8 mg of Lu could provide 18.4mCi(0.68GBq) of activity. Therefore, a single patient dose of150–200mCi (5.55–7.40GBq) can be prepared by using200–275 mg of peptide. In order to carry out a comparativeevaluation of the radiolabeled DOTA–TATE prepared byus, an analysis of a reported case (Kwekkeboom et al.,2003a, 2001) was carried out. It has been shown that177Lu–DOTA–TATE prepared for a typical PRRT pro-cedure uses 100 mg of the DOTA–TATE conjugatewhich incorporates 50mCi (1.85GBq) of 177Lu activity(Kwekkeboom et al., 2001). This implies that to achieve asingle patient dose of 150–200mCi (5.55–7.40GBq),300–400 mg of the conjugate has been used. Consideringthe practical aspects of time required for Post-irradiationradiochemical processing, preparation and quality controlof the radiopharmaceutical and subsequent transportationto the nuclear medicine facilities, �15mCi (0.55GBq) of177Lu activity in 25 mg of DOTA–TATE can be delivered tothe nuclear medicine facilities. Therefore, the single patientdose of 150–200mCi (5.55–7.40GBq) can be prepared byusing 250–333 mg of DOTA–TATE conjugate at our end,which is comparable or possibly less than the reportedvalues.

3.5. Stability of 177Lu–DOTA–TATE

The stability of the 177Lu-labeled DOTA–TATE complexwas studied by employing standard quality control techni-ques mentioned earlier. It was observed that the radi-olabeled conjugate retained its radiochemical purity after7 d of its preparation when stored at room temperature.

ARTICLE IN PRESST. Das et al. / Applied Radiation and Isotopes 65 (2007) 301–308 307

The complex prepared using maximum possible amount ofLu (0.8 mg) also showed excellent stability till 7 d whenstored at identical conditions.

4. Conclusion

Production of 177Lu by the (n,g) route from enriched176Lu target with a specific activity of �23000mCi/mg(850GBq/mg) was achievable when irradiation was carriedout at a thermal neutron flux of 1� 1014 n/cm2/s for 21 d.177Lu–DOTA–TATE could be prepared in high radio-chemical yield and adequate stability using the 177Luproduced indigenously. The maximum specific activityachievable via careful optimization of the irradiationparameters was found to be adequate for the preparationof a therapeutic dose of the radiopharmaceutical. The in-house preparation of this agent using indigenouslyproduced 177Lu was found to be a more cost-effectiveroute over the method involving use of 177Lu fromcommercial sources. Clinical studies with this agent willbe carried out after obtaining clearances from relevantregulatory authorities.

Acknowledgments

The authors sincerely acknowledge the support providedby the International Atomic Energy Agency (IAEA) in theform of a coordinated research project (CRP). The authorsexpress their sincere thanks to Dr. V. Venugopal, Director,Radiochemistry and Isotope Group for his valuablesuggestions and constant encouragement through out thecourse of the work. The sincere help received from ourcolleagues, Dr. S.V. Thakare and Mr. K.C. Jagadeesan forcarrying out irradiations of lutetium oxide target isgratefully acknowledged.

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