[CANCER RESEARCH 61, 2804–2808, April 1, 2001]

Advances in Brief

Highly Specific Tumor Binding of a 213Bi-labeled Monoclonal Antibody againstMutant E-Cadherin Suggests Its Usefulness for Locoregionala-Radioimmunotherapy of Diffuse-Type Gastric Cancer

Reingard Senekowitsch-Schmidtke,1 Christoph Schuhmacher, Karl-Friedrich Becker, Tuomo K. Nikula,Christof Seidl, Ingrid Becker, Matthias Miederer, Christos Apostolidis, Christoph Adam, Roswitha Huber,Elisabeth Kremmer, Kathrin Fischer, and Markus SchwaigerDepartments of Nuclear Medicine [R. S-S., C. Se., M. M., R. H., K. F., M. Sl.] and Surgery [C. Sch., C. Ad.], and Institute of Pathology [K-F. B., I. B.], Klinikum rechts der Isar,Technische Universita¨t Munchen; Institute of Molecular Immunology [E. K.]; and Institute of Pathology [K-F. B.] GSF, National Research Center for Environment and Health,Munchen, European Commission, Institute for Transuranium Elements, Karlsruhe [T. K. N., C. Ap.], 81675 Muenchen, Germany

Abstract

A monoclonal antibody (E-cadherin delta 9–1) directed against a char-acteristic E-cadherin mutation (in-frame deletion of exon 9), found indiffuse-type gastric cancer but not in any normal tissue, was conjugatedwith the high linear energy transfer a-emitter 213Bi and tested for itsbinding specificity in s.c. and i.p. nude mice tumor models. After intra-tumoral application in s.c. tumors expressing mutant E-cadherin, the213Bi-labeled antibody was specifically retained at the injection site asshown by autoradiography. After injection into the peritoneal cavity,uptake in small i.p. tumor nodules expressing mutant E-cadherin was17-fold higher than in tumor nodules expressing wild-type E-cadherin(62% injected dose/g versus 3.7% injected dose/g). 78% of the totalactivity in the ascites fluid was bound to free tumor cells expressingmutant E-cadherin, whereas in control cells, binding was only 18%. Theselective binding of the213Bi-labeled, mutation-specific monoclonal anti-body E-cadherin d 9–1 suggests that it will be successful fora-radioim-munotherapy of disseminated tumors after locoregional application.

Introduction

The use of radiolabeled MAbs2 in radioimmunotherapy is limitedbecause of the lack of tumor-specific antigens. In most cases reportedthus far, tumor antigens that serve as targets are not tumor-specific,being overexpressed by tumor cells and also at a lower level bynormal cells. Thus far, only one tumor-specific MAb has been re-ported that recognizes a mutant form of the epidermal growth factorreceptor (EGFR vIII) that is found on different tumor types but not onnormal human tissue (1). This antibody has been labeled with125I,131I, and the a-emitter 211At, and it appears to be a promisingcandidate for radioimmunotherapy (2, 3).

In-frame deletions of exons 8 or 9 in the mRNA coding for the celladhesion molecule E-cadherin are characteristic of diffuse-type gas-tric carcinomas. These mutations alter cell adhesion functions andcontribute to the diffuse spread of this cancer type (4, 5). A rat MAb,designated d9MAb, was generated that specifically reacts with mutantE-cadherin lacking exon 9 but not with the wild-type protein. d9MAbwas found to react with 13% (22 of 172) of E-cadherin-expressingdiffuse-type gastric cancers (6). Because of this specific tumor celltargeting, d9MAb coupled with thea-emitting radionuclide213Bicould have a significant potential for the locoregional radioimmuno-

therapy of disseminated, diffuse-type gastric carcinoma, which isoften associated with i.p. spread of single malignant cells leading toperitoneal carcinomatosis. The advantages ofa-particles are their highLET and their short range of a few cell diameters. These featuresresult in a high localized energy deposition, even in single target cells,and minimal irradiation of surrounding normal tissue.

We have established an i.p. tumor model using cells expressingE-cadherin with an exon 9 deletion that mimics the clinical situationin human gastric cancer with i.p. tumor spread that is known to be acrucial process in diffuse-type gastric cancer. The free i.p. applicationof a tumor-specific MAb labeled with an appropriate radionuclideseems to be an effective treatment of peritoneal tumor spread. Here wereport that the d9MAb coupled with thea-emitter 213Bi specificallybinds to mutant E-cadherin after locoregional application.

Materials and Methods

Antibody. The rat MAb recognizing mutant E-cadherin lacking exon 9 wasgenerated as described previously (6). Briefly, a 13-amino acid peptide span-ning the fusion junction between exons 8 and 10 of mutant E-cadherin with anexon 9 deletion was injected into Lou/C rats i.p. and s.c. for immunization.After fusion of the immune rat spleen cells with a myeloma cell line, hybri-doma supernatants were tested by a solid phase immunoassay using themutation-specific peptide coupled to BSA. A tumor cell-specific, MAb againstthe delta 9 peptide, referred to as d9MAb (clone 6H8) was selected for thestudies described below.

Conjugation of Chelate to d9MAb and Radiolabeling. d9MAb wasconjugated to SCN-CHX-A0-DTPA as described previously (7, 8). The num-ber of chelates per antibody ranged from 5 to 10 as determined by a standard111In-assay (9). For comparative binding studies, both the MAb chelate con-struct and the MAb without chelate were labeled with125I according to theIodogene method.213Bi (t1/2 5 46 min) was eluted from a225Ac/213Bigenerator provided by the Institute for Transuranium Elements, Karlsruhe,Germany (10), with 0.1M HCl/0.1M NaI as the BiI4

2/BiI 522 anion. The eluant

was adjusted to pH 5.3 with 2M ammonium acetate, and;100 mg of thechelated antibody were added and allowed to react for 5 min. The213Bi-immunoconjugate was purified by size exclusion chromatography (PharmaciaPD-10) with 2 ml of PBS.

The coupling of111In (InCl3, Mallinckrodt) to the chelated d9MAb wascarried out using the213Bi-protocol omitting NaI. The111In-immunoconjugatewas applied for scintigraphic imaging of i.p. retention and biodistribution. Thelabeling efficiency was assayed via TLC with instant thin-layer chromatogra-phy paper (Gelman Sciences).

Cell Lines. The human MDA-MB-435S mammary carcinoma cell line(American Type Culture Collection, Manassas, VA) was transfected with exon9-deleted E-cadherin cDNA and wild-type E-cadherin cDNA, respectively (5).The cells were grown at 37°C in a humidified atmosphere with 5% CO2 inDMEM containing 4.5 g/l glucose supplemented with 10% FCS. For selectionof the transfected MDA-MB-435S cells Geneticin was added to the cell

Received 10/3/00; accepted 2/12/01.The costs of publication of this article were defrayed in part by the payment of page

charges. This article must therefore be hereby markedadvertisementin accordance with18 U.S.C. Section 1734 solely to indicate this fact.

1 To whom requests for reprints should be addressed, at Department of NuclearMedicine, Klinikum rechts der Isar, Technische Universita¨t Munchen, Ismaningerstr. 22,D-81675 Muenchen, Germany.

2 The abbreviations used are: MAb, monoclonal antibody; d9MAb, E-cadherin delta9–1 MAb; LET, linear energy transfer.

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medium. The cells were harvested by rinsing the monolayer with 1 mM EDTAand counted in a hemocytometer.

Determination of Antigen Density of Mutant E-Cadherin TransfectedCells and Binding Characteristics of the Labeled MAb. The antigen den-sity and binding characteristics of the radiolabeled MAb were analyzed byScatchard analysis. Mutant E-cadherin transfected cells (106) were incubatedwith increasing concentrations of d9MAbs labeled with125I or 213Bi. Specificbinding was confirmed by the failure of radiolabeled d9MAb to bind to cellsexpressing wild-type E-cadherin. Antigen density was also determined forunlabeled and chelate-coupled MAbs by indirect immunofluorescence withflow cytometry. The fluorescence signal was quantified by a calibration curveestablished with fluorescence quantitation beads (Quantibrite PE; BectonDickinson).

Animal Models. To investigate the specific binding properties of213Bi-labeled d9Mab, two different tumor models, a s.c. solid tumor model as wellas an i.p. tumor model, were established. For that purpose, 4–5 week oldfemale athymic mice were inoculated s.c. or i.p. with 13 107 cells expressingeither mutant E-cadherin or wild-type E-cadherin as a negative control. Afteri.p. injection, two mice were sacrificed every 2 days until day 12 and thereafterat weekly intervals for histological examination of i.p. tumor progression. Allexperiments with mice were performed in accordance with the guidelines forthe use of living animals in scientific studies and the “German Law for theProtection of Animals.”

Biodistribution Studies. Mice, bearing s.c. tumors of 100–200 mg derivedfrom both mutant and wild-type E-cadherin transfected cells (4–5 weeks aftertumor cell inoculation), were injected i.v. with 3.7 MBq (100mCi) of 213Bi-d9MAb to study the accumulation in tumors and organs at 45 min and at 3 hpostinjection. In addition, 37 kBq (1mCi) of the radioimmunoconjugate wereinjected directly into the tumor at three different sites. After injection, thetumors were removed, immediately frozen, cut into 8-mm sections, and ana-lyzed for intratumoral distribution of the radioimmunoconjugate via exposure(15 min) to a high-sensitive Micro Imager system, which is capable of a spatialresolution of 10–15mm (Biospace Measures, Paris, France).

Animals bearing small i.p. tumor nodules (0.1–2.0 mm in diameter) with orwithout ascites (;3–4 weeks after i.p. tumor cell inoculation) received aninjection of 3 MBq (80mCi) of 213Bi-d9MAb into the peritoneal cavity andwere sacrificed at 45 min and 3 h postinjection. Tumor nodules and variousnormal tissues were removed, washed, and weighed, and the activity wasdetermined by gamma counting using the 440-keVg-emission of213Bi. Theresults were expressed as a percentage of the injected dose/g of tissue (%ID/g).Each reported value represents the mean and the SD of eight animals. Ascitesvolume was measured, and, after centrifugation, the activity in the pellet andthe supernatant was determined.

To obtain scintigraphic images with optimal resolution (Eg111In, 172 and

249 keV; Eg213Bi, 440 keV) and to gain information about the long-term

retention (t1/2111In, 2.6 d;t1/2

213Bi, 46 min) of the immunoconjugate in micebearing i.p. tumor nodules expressing either mutant or wild-type E-cadherinand in mice without tumor, scintigraphic images were taken at 3, 24, and 48 hafter i.p. injection of111In-labeled d9MAb (740 kBq). Immediately after the48 h scintigram, the animals were sacrificed and the distribution of111Inimmunoconjugate in representative organs was determined and expressed as%ID/g.

Statistical Analysis. Unpaired Student’st tests were performed to comparethe mean values.Ps # 0.05 were considered statistically significant.

Results

Antibody Specificity. Specific binding of the rat d9MAb reactingwith mutant, but not with wild-type, E-cadherin was demonstrated byWestern blot and immunohistochemistry. In human tissue, d9MAbreacted only with tumor cells of diffuse-type gastric cancer with anexon 9 deletion and did not show any cross-reaction to any normalhuman tissue similar to the clone 7E6, as described previously (6).

Radiolabeling and Tumor Cell Binding of d9MAb. The labelingefficiency of CHX-A0-DTPA-d9MAb with 213Bi was .90% at aspecific activity of 1.48 GBq/mg (40 mCi/mg). The binding charac-teristics of213Bi- and 125I-labeled d9MAb to MDA-MB-435S cellstransfected with mutant E-cadherin were evaluated by Scatchard anal-

ysis. With both125I-labeled and213Bi-conjugated MAbs, 5.53 104

binding sites/cell and a dissociation constant of 1.9 nmol/l weredetermined. These results were confirmed by flow cytometry usingunconjugated and CHX-A0-DTPA-conjugated MAbs. The resultsdemonstrated that conjugation and213Bi-labeling of the MAb do notinfluence the binding characteristics. The binding of both radiolabeledMAbs to cells expressing wild-type E-cadherin was,4% comparedwith mutant E-cadherin-expressing cells.

Development of the Tumor Models.s.c. tumors of 100–200 mgin weight developed 4–5 weeks after s.c. inoculation of 13 107 tumorcells.

Up to day 6 after i.p. inoculation of mutant and wild-type E-cadherin-transfected cells, single tumor cells and small tumor-cellclusters consisting of;100 cells could be detected histologically inthe peritoneal cavity. After day 10, tumor attachment to the visceralorgans and the peritoneum could be demonstrated microscopically.Beginning from day 20 after tumor cell inoculation, macroscopictumor nodules in the mesenterium ranging from 0.1 to 2 mm indiameter could be observed (Fig. 1a). Histological sections from thesetumor nodules showed tumor cells on the serosa and also invasivetumor cells with a desmoplastic reaction (Fig. 1b). The tumor cells onthe serosa often lost their cell-to-cell contact, and isolated tumor cellsor cell clusters could be detected in the ascites. Forty percent of theanimals developed ascites that contained up to 13 108 tumor cells/mlas single cells and as cell clusters (Fig. 1c). At this stage of tumordevelopment, the animals were used for biodistribution studies afteri.p. injection of the213Bi immunoconjugate.

Biodistribution Studies of 213Bi- and 111In-d9MAb. In the s.c.tumor model, the activity concentration of213Bi-d9MAb 3 h after i.v.injection was lower than in the blood; this was as expected for intactMAbs, which slowly diffuse from the circulation into solid tumortissue. However, binding in tumors expressing mutant E-cadherin was3-fold higher than in tumors expressing wild-type E-cadherin.

After intratumoral injection in s.c. tumors, the specific binding of213Bi-d9MAb to mutant E-cadherin could be demonstrated by auto-radiographic images. Fig. 2ashows local retention of the213Bi im-munoconjugate at the three injection sites in the center of the tumorthat expressed E-cadherin with an exon 9 deletion. Retention is alsoseen at the three puncture sites on the tumor periphery. In contrast,tumors expressing wild-type E-cadherin did not show any specificretention of the213Bi coupled MAb (Fig. 2b).

In the i.p. tumor model,213Bi-d9MAb was injected into the peri-toneal cavity 3–4 weeks after the inoculation of tumor cells express-ing mutant E-cadherin or wild-type E-cadherin, and the biodistribu-tion was quantified at 45 min and 3 h postinjection. The results aresummarized in Table 1. In animals that had not developed ascites, ahigh specific uptake of up to 626 14% ID/g at 45 min and 586 19%ID/g at 3 h was observed in small tumor nodules expressing mutantE-cadherin. In the wild-type E-cadherin model, uptake was only3.7 6 1.0% ID/g at 45 min and 3.46 0.9 at 3 h. In all other tissues,the uptake of213Bi immunoconjugate was low. The low213Bi accu-mulation in the kidneys, which are known to accumulate free bismuth,indicates the stability of the immunoconjugate. In animals bearingtumors expressing wild-type E-cadherin that does not bind the MAb,however, uptake in normal tissue was statistically significantly higherthan in animals expressing mutant E-cadherin. In mice with ascites(up to 5 ml), the concentration of213Bi-d9MAb in tumor nodules andthe other organs was statistically significantly reduced compared withanimals without ascites, depending on the volume of ascites and thenumber of tumor cells in the fluid. This result suggests that theantibody was rapidly and firmly bound to free accessible mutantE-cadherin on tumor cells in the ascites. After centrifugation of asciteswith cells expressing mutant E-cadherin, 78% of the213Bi activity

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was recovered in the cell pellet in contrast with 18% bound in thepellet of ascites from cells expressing wild-type E-cadherin.

Scintigraphic images of mice bearing small tumor nodules, express-ing mutant E-cadherin or wild-type E-cadherin, and of mice withouttumor obtained 48 h after i.p. injection of111In-d9Mab, are shown inFig. 3. In mice without tumor, the activity is mainly distributed in theblood pool of heart, lungs, and liver (Fig. 3a). A similar activitydistribution was found for mice bearing tumors that expressed wild-type E-cadherin (Fig. 3c). This indicates that in both cases most of theactivity was reabsorbed from the peritoneal cavity. Conversely, in themouse with multiple i.p. tumor nodules expressing mutant E-cadherin,a considerable amount of activity was retained in the peritoneal cavity,resulting in a clearly visible lower background activity compared withthe two controls (Fig. 3b). Tissue distribution data of111In were inaccordance with the results of the scintigraphic images. Activityaccumulation in tumor nodules expressing mutant E-cadherin, was

56% ID/g compared with 4.8% ID/g in controls. Activity concentra-tion of 111In in the blood of animals inoculated with wild-typeE-cadherin-expressing cells and of animals without tumor cell inoc-ulation was 11% ID/g compared with 5% ID/g in animals with tumornodules expressing mutant E-cadherin.

Discussion

Early i.p. dissemination of tumor cells is a crucial event in thecourse of gastric carcinoma, resulting in peritoneal carcinomatosis andrapid deterioration of the patient’s clinical status. Apart from a fewexperimental therapeutic strategies, there is currently no specifictreatment for peritoneal cancer spread.

Effective treatment of the i.p. compartment would require locore-gional administration of a cytotoxic substance into the peritonealcavity that could specifically bind to diffusely spread tumor cells and

Fig. 1. An example of the development of i.p. carcinomatosis with ascites 5 weeks after injection of 13 107 cells expressing mutant E-cadherin.a, macroscopic view showingdevelopment of small tumor nodules in the peritoneum (arrow). b, histological section of an i.p. nodule with tumor cells on the serosa (arrow head) and invasive tumor cells in themesenteric adipose tissue (arrow;3200; H&E staining).c, single tumor cells (arrow heads) and small cell clusters (arrow) in the ascites fluid with a nonspecific inflammatory reaction(3400; modified May-Grunwald-Giemsa staining).

Fig. 2.a, autoradiography of an 8-mm tumor section 1 h after intratumoral injection of213Bi-d9MAb in a tumor expressing mutant E-cadherin (activity retention at the three injectionsites in the center of the tumor [arrow heads] and at the puncture sites at the tumor periphery [arrows]).b, autoradiography of a 8mm tumor section, 1 h after intratumoral injectionof a 213Bi-d9MAb in wild type E-cadherin tumors (no specific activity retention).

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tumor cell clusters. Monoclonal antibodies that specifically recognizetumor cell antigens coupled with a radionuclide with high LET arepromising candidates.

Because the d9MAb used in our experiments specifically binds tomutant E-cadherin expressed by diffuse-type gastric carcinoma, it isan ideal vehicle to attach radionuclides to gastric carcinoma cells thathave spread diffusely into the peritoneal cavity.

By choosing the appropriate radionuclide, the range of thecytotoxic effect can be matched to the size of the tumor. For theradioimmunotherapy of malignancies with large tumor masses,b-emitting radionuclides such as131I, 188Re, or 90Y, with meantissue ranges of 0.9 to 3.9 mm, have been coupled to MAbs. Forselective irradiation of single tumor cells or small tumor cellclusters, the new approach of labeling tumor-specific MAbs witha-emitting nuclides seems to be very promising. Thea-particlesemitted by 212Bi, 211At, and 213Bi have short ranges of only50 –100mm and a high LET of;100 keV/mm that deposit a largeamount of energy within a few cell diameters.a-Emitter immuno-conjugates have proven to be powerful therapeutic agents in animalexperiments (11–13), especially for malignancies that spread onthe surface of the body cavities, such as ovarian cancer andmalignant meningitis (14 –16).

We developed a nude mouse model for i.p. tumor spread similar tothat which occurs in patients with diffuse-type gastric cancer. In this

model, d9MAb demonstrated high and specific binding to small tumornodules established from tumor cells expressing mutant E-cadherin,whereas binding of d9MAb to tumors expressing wild-type E-cadherin was comparatively low. In addition, the213Bi-labeledd9MAb bound to the tumor cells in the i.p. cavity within less than onehalf-life of 213Bi (46 min). The binding remained stable at least 3 hafter injection, when 94% of the injected213Bi activity had decayed atthe tumor site.

The number of antigen molecules on the E-cadherin-transfectedtumor cells was calculated to be 5.53 104/cell by Scatchard analysis.At a specific activity of 1.48 GBq/mg,213Bi-labeled d9MAb canattach 40a-particles to a tumor cell. It has been reported in a numberof cell lines that 3–9a-particles bound/cell can reduce clonogenic cellsurvival to as low as 10% (17–19). Because the antigen density onhuman diffuse gastric carcinoma cells as shown by immunohisto-chemistry may exceed that of our tumor model, binding of213Biimmunoconjugates should be sufficient to guarantee destruction ofalmost all of the tumor cells. By increasing the specific activitywithout the loss of immunoreactivity caused by radiolysis, the spec-ificity of binding and the therapeutic efficiency could probably beimproved further.

The beneficial therapeutic effects ofa-emitter-immunoconjugatesare currently being evaluated in two clinical trials. Patients sufferingfrom acute myeloic leukemia are being treated with213Bi-labeledHuM195 MAb recognizing CD33, a differentiation antigen expressedin leukemic cells. More than one-half of the 17 patients treated thusfar have shown a reduction of leukemic cells in the peripheral blood,and a few also have shown decreased numbers of bone marrow blastcells (20). The MAb 81C6 specifically binds to the matrix glycopro-tein tenascin that is expressed by glioma cells but not by normal braintissue. The211At labeled antibody has been applied locally to thesurgical cavity created by the glioma resection with promising results(16).

The results obtained in our experimental model with the d9MAblabeled with213Bi suggest that this radioimmunoconjugate and similarones targeting other E-cadherin mutations (e.g., exon 8 deletion)should be tested in clinical therapeutic trials for a subgroup of diffuse-type gastric carcinoma patients. This would be the first application ofsuch a method in disseminated gastro-intestinal tumors.

Acknowledgments

We thank Dr. Martin W. Brechbiel at the NIH for providing SCN-CHX-A0-DTPA chelate. We acknowledge the kind assistance of Roger Molinet andRamon Carlos-Marquez in the separation of225Ac and in the preparation of213Bi generators. We thank Susanne Daum and Stephanie Alam for technicalassistance and James Mueller for carefully reading the manuscript.

Table 1 Biodistribution of213Bi d9MAb in animals with i.p. tumors expressing mutant or wild-type E-cadherin, 45 min and 3 h after i.p. injectiona

Organ

Mutant E-cadherin without ascites Mutant E-caderin with ascites Wild-type E-cadherin without ascites

45 min 3 h 45 min 3 h 45 min 3 h

Blood 1.16 0.4 2.66 0.9 0.46 0.1 1.46 0.4 2.86 0.6 5.96 1.4Tumor 626 14 586 19 7.16 3.4 9.36 4.2 3.76 1.0 3.46 0.9Heart 0.56 0.1 1.16 0.3 0.16 0.03 0.56 0.1 1.56 0.4 2.16 0.3Lung 0.76 0.2 1.46 0.4 0.26 0.08 0.66 0.2 1.16 0.3 2.06 0.6Spleen 1.06 0.4 1.26 0.5 0.36 0.07 0.96 0.2 1.66 0.6 1.96 0.8Stomach 2.16 1.0 1.76 0.4 0.66 0.1 0.86 0.3 3.36 0.9 4.16 1.3Bowel 1.46 0.3 1.26 0.3 0.86 0.3 0.96 0.3 2.86 0.6 2.96 0.7Peritoneum 3.96 1.8 3.26 1.4 2.56 0.9 3.16 1.2 2.36 0.1 1.86 0.3Kidney 3.66 1.4 5.16 1.9 2.86 1.2 4.06 0.8 4.16 1.3 6.86 2.3Liver 1.26 0.4 2.16 0.8 0.56 0.1 1.26 0.4 2.56 0.3 3.86 0.9Muscle 0.16 0.08 0.46 0.1 0.096 0.03 0.26 0.06 0.46 0.1 0.66 0.1Ascites 14.46 6.3 17.36 5.6

a %ID/g, mean6 SD; n 5 8.

Fig. 3. Scintigrams of mice 48 h after i.p. injection of 740 kBq (20mCi) 111In-d9MAb.a, mouse without tumor cell injection showing blood pool mainly in the heart, lungs, andliver; no visible retention of activity in the peritoneal cavity.b, mouse with tumorsexpressing mutant E-cadherin; besides some blood pool activity, there is a clearly visibleactivity accumulation in peritoneal tumor nodules (arrows).c, mouse with i.p. tumorsexpressing wild-type E-cadherin showing blood pool mainly as ina.

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References

1. Wikstrand C. J., McLendon, R. E., Friedman, A. H., and Bigner D. D. Cell surfacelocalization and density of the tumor-associated variant of the epidermal growthfactor receptor, EGFRvIII. Cancer Res.,57: 4130–4140, 1997.

2. Reist, C. J., Archer, G. E., Wikstrand, C. J., Bigner, D. D., and Zalutsky, M. R.Improved targeting of an anti-epidermal growth factor receptor variant III monoclonalantibody in tumor xenografts after labeling using N-Succinimidyl 5-iodo-3-pyridin-ecarboxylate. Cancer Res.,57: 1510–1515, 1997.

3. Reist, C. J., Foulon, C. F., Alston, K., Bigner, D. D., and Zalutsky, M. R. Astatine-211labeling of internalizing anti-EGFRvIII monoclonal antibody using N-Succinimidyl5-[211At]Astato-3-pyridinecarboxylate. Nucl. Med. Biol.,26: 405–411, 1999.

4. Becker, K-F., Atkinson, M. J., Reich, U., Becker, I., Nekarda, H., Siewert, J. R., andHofler, H. E-cadherin gene mutations provide clues to diffuse type gastric carcino-mas. Cancer Res.,54: 3845–3852, 1994.

5. Handschuh, G., Candidus, S., Luber, B., Reich, U., Schott, C., Oswald, S., Becke, H.,Hutzler, P., Birchmeier, W., Hofler, H., and Becker, K-F. Tumor-associated E-cadherin mutations alter cellular morphology, decrease cellular adhesion, and increasecellular motility. Oncogene,18: 4301–4312, 1999.

6. Becker, K-F., Kremmer, E., Eulitz, M., Becker, I., Handschuh, G., Schuhmacher, C.,Muller, W., Gabbert, H. E., Ochiai, A., Hirohashi, S., and Hofler, H. Analysis ofE-cadherin in diffuse-type gastric cancer using a mutation-specific monoclonal anti-body. Am. J. Pathol.,155: 1803–1809, 1999.

7. Brechbiel, M. W., Pippin, C. G., McMurry, T. J., Milenic, D., Roselli, M., Colcher,D., Gansow, O. A. An effective chelating agent for labeling of monoclonal antibodywith Bi-212 for a-particle mediated radioimmuno-therapy. J. Chem. Soc. Chem.Commun.,1169–1170:1991.

8. Nikula, T. K., McDevitt, M. R., Finn, R. D., Wu, C., Kozak, R. W., Garmestani, K.,Brechbiel, M. W., Curcio, M. J., Pippin, C. G., Tiffany-Jones, L., Geerlings, M. W.,Sr., Apostolidis, C., Molinet, R., Geerlings, M. W., Jr., Gansow, O. A., Scheinberg,D. A. a-Emitting bismuth cyclohexylbenzyl DTPA constructs of recombinant hu-manized anti-CD33 antibodies: pharmacokinetics, bioactivity, toxicity, and chemis-try. J. Nucl. Med.,40: 166–176, 1999.

9. Nikula, T. K., Curcio, M. J., Brechbiel, M. W., Gansow, O. A., Finn, R. D.,Scheinberg, D. A. A rapid, single vessel method for preparation of clinical gradeligand conjugated monoclonal antibodies. Nucl. Med. Biol.,22: 387–390, 1995.

10. Koch, L., Apostolidis, C., Janssens, W., Molinet, R., van Geel, J. Production ofAc-225 and application of the Bi-213 daughter in cancer therapy. Czech. J. Phys.,40:817–822, 1999.

11. Huneke, R. B., Pippin, C. G., Squire, R. A., Brechbiel, M. W., Gansow, O. A., andStrand, M. Effectivea-particle-mediated radioimmunotherapy of murine leukemia.Cancer Res.,52: 5818–5820, 1992.

12. Behr, T. M., Be´he, M., Stabin, M. G., Wehrmann, E., Apostolidis, C., Molinet, R.,Strutz, F., Fayyazi, A., Wieland, E., Gratz, S., Koch, L., Goldenberg, D. M., andBecker, W. High-linear Energy Transfer (LET)a versusLow-LET b emitters inradioimmuno-therapy of solid tumors: therapeutic efficacy and dose-limiting toxicityof 213Bi- versus90Y-labeled CO17–1A Fab9fragments in a human colonic cancermodel. Cancer Res.,59: 2635–2643, 1999.

13. Kennel S. J., Stabin, M., Yoriyaz, H., Brechbiehl, M., and Mirzadeh, S. Treatment oflung tumor colonies with90Y targeted to blood vessels: comparison with thea-particle emitter 213Bi. Nucl. Med. Biol.,26: 149–157, 1999.

14. Vergote, I., Larsen, R. H., de Vos, L., Winderen, M., Ellingsen, T., Bjørgum, J., Hoff,P., Aas, M., Trope, C., and Nustad, K. Distributioon of intraperitoneally injectedmicrosheres labeled with thea-emitter astatine (211At) compared with Phosphorus(32P) and yttrium (90Y) colloids in mice. Gynecol. Oncol.,47: 358–365, 1992.

15. Andersson, H., Lindegren, S., Back, T., Jacobsson, L., Leser, G., and Horvath, G.Radioimmunotherapy of nude mice with intraperitoneally growing ovarian cancerxenograft utilizing211At-labeled monoclonal antibody MOv18. Anticancer Res.,20:459–462, 2000.

16. Zalutsky, M. R., Vaidyanathan, G. Astatine-211-labeled radiotherapeutics: an emerg-ing approach to targeteda-particle radiotherapy. Curr. Pharm. Des.,6: 1433–1455,2000.

17. Humm, J. L., Roeske, J. C., Fisher, D. R., and Chen, G. T. Y. Microdosimetricconcepts in radioimmunotherapy. Med. Phys.,20: 535–541, 1993.

18. Roeske, J. C., and Stinchcomb, T. G. Dosimetric framework for therapeutica-particleemitters. J. Nucl. Med.,38: 1923–1929, 1997.

19. Larsen, R. H., Akabani, G., Welsh, P., and Zalutsky, M. R. The cytotoxicity andmicrodosimetry of Astatine-211-labeled chimeric monoclonal antibodies in humanglioma and melanoma cellsin vitro. Radiat. Res.,149: 155–162, 1998.

20. McDevitt, M. R., Sgouros, G., Finn, R. D., Humm, J. L., Jurcic, J. G., Larson, S. M.,and Scheinberg, D. A. Radioimmunotherapy witha-emitting nuclides. Eur. J. Nucl.Med., 25: 1341–1351, 1998.

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213Bi MAb FOR LOCOREGIONAL a-RADIOIMMUNOTHERAPY

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2001;61:2804-2808. Cancer Res   Reingard Senekowitsch-Schmidtke, Christoph Schuhmacher, Karl-Friedrich Becker, et al.   Gastric Cancer

-Radioimmunotherapy of Diffuse-Typeαfor Locoregional Antibody against Mutant E-Cadherin Suggests Its Usefulness

Bi-labeled Monoclonal213Highly Specific Tumor Binding of a

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