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[Cancer Research 66, 8227-8232, August 15, 2006]
© 2006 American Association for Cancer Research

Experimental Therapeutics, Molecular Targets, and Chemical Biology

The Anti-CD25 Monoclonal Antibody 7G7/B6, Armed with the {alpha}-Emitter 211At, Provides Effective Radioimmunotherapy for a Murine Model of Leukemia

Meili Zhang1, Zhengsheng Yao1, Zhuo Zhang1, Kayhan Garmestani1, Vladimir S. Talanov2, Paul S. Plascjak3, Sarah Yu3, Hyung-Sik Kim3, Carolyn K. Goldman1, Chang H. Paik3, Martin W. Brechbiel2, Jorge A. Carrasquillo3 and Thomas A. Waldmann1

1 Metabolism Branch and 2 Radiation Oncology Branch, Center for Cancer Research, National Cancer Institute, and 3 PET and Nuclear Medicine Department, Clinical Center, NIH, Bethesda, Maryland

Requests for reprints: Thomas A. Waldmann, Metabolism Branch, Center for Cancer Research, National Cancer Institute, NIH, Building 10, Room 4N115, 10 Center Drive, Bethesda, MD 20892-1374. Phone: 301-496-6656; Fax: 301-496-9956; E-mail: tawald{at}helix.nih.gov.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Radioimmunotherapy of cancer with radiolabeled antibodies has shown promise. {alpha}-Particles are very attractive for cancer therapy, especially for isolated malignant cells, as is observed in leukemia, because of their high linear energy transfer and short effective path length. We evaluated an anti-CD25 [interleukin-2 receptor {alpha} (IL-2R{alpha})] monoclonal antibody, 7G7/B6, armed with 211At as a potential radioimmunotherapeutic agent for CD25-expressing leukemias and lymphomas. Therapeutic studies were done in severe combined immunodeficient/nonobese diabetic mice bearing the karpas299 leukemia and in nude mice bearing the SUDHL-1 lymphoma. The results from a pharmacokinetic study showed that the clearance of 211At-7G7/B6 from the circulation was virtually identical to 125I-7G7/B6. The biodistributions of 211At-7G7/B6 and 125I-7G7/B6 were also similar with the exception of a higher stomach uptake of radioactivity with 211At-7G7/B6. Therapy using 15 µCi of 211At-7G7/B6 prolonged survival of the karpas299 leukemia–bearing mice significantly when compared with untreated mice and mice treated with 211At-11F11, a radiolabeled nonspecific control antibody (P < 0.01). All of the mice in the control and 211At-11F11 groups died by day 46 whereas >70% of the mice in the 211At-7G7/B6 group still survived at that time. In summary, 211At-7G7/B6 could serve as an effective therapeutic agent for patients with CD25-expressing leukemias. (Cancer Res 2006; 66(16): 8227-32)


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
The observation that interleukin-2 receptor {alpha} (IL-2R{alpha}; CD25) is not expressed by normal resting cells but is expressed by a high proportion of the abnormal cells in certain forms of lymphoid neoplasia provides the rationale for the use of the IL-2R{alpha} as a target for therapeutic agents (1). Clinical remissions have been observed in patients with adult T-cell leukemia treated with the unmodified humanized anti-Tac monoclonal antibody (daclizumab), which recognizes CD25, at receptor-saturating doses or this antibody labeled at high specific activity with yttrium-90 (90Y; refs. 2, 3). A paradigm is emerging that, for cancer therapy, the addition of two therapeutic agents that function via different mechanisms may be greater than additive in their cytotoxic action leading to malignant cell death (4–8). In our previous therapeutic trials, we obtained improved therapeutic efficacy by combining daclizumab at receptor-saturating doses with pretargeted radioimmunotherapy in a adult T-cell leukemia model (4, 5). However, with a single radiolabeled monoclonal antibody, it is difficult to obtain the complementary actions of receptor-saturating doses of daclizumab to yield antibody-dependent cellular cytotoxicity and IL-2 deprivation–mediated apoptotic leukemic cell death, in conjunction with the tumor cytoreduction provided by irradiation mediated by therapeutic radionuclides (e.g., 90Y or 211At) delivered at high specific activity by the antibody to the leukemic cell surface. To obtain cytokine deprivation–mediated cell death, one must use large receptor-saturating quantities of the monoclonal antibody. However, administration of large quantities of monoclonal antibody armed with a radionuclide leads to a low specific activity and a low proportion of the administered radiolabeled antibody delivered to the tumor cells. Resulting circulating unbound radiolabeled antibody yields unacceptable bone marrow toxicity, which in turn reduces the maximum dose of radioactivity that can be administered. In the present study, we address the limitations inherent in the use of a single radiolabeled monoclonal antibody by employing 7G7/B6, which also recognizes CD25 but binds to a different epitope than daclizumab. These two antibodies are not cross-competing. Our long-term goal is to use daclizumab at receptor-saturating doses in combination with small quantities of 7G7/B6 armed at high specific activity with radionuclides for the treatment of CD25-expressing leukemias and lymphomas.

Monoclonal antibodies directed against tumor-associated antigens armed with diverse radionuclides are being investigated as therapeutic agents for the treatment of malignant disease (3, 9–13). Although encouraging results have been obtained in the treatment of lymphoma with monoclonal antibodies armed with ß-emitting radionuclides, further development is needed before an ideal radioimmunotherapeutic agent is achieved (3, 13). The {alpha}-emitting radionuclides seem to have several advantages when compared with ß-emitting radionuclides in radioimmunotherapy, especially with isolated malignant cells as in leukemia (14–16). The high linear energy transfer makes them highly cytotoxic with a relative biological effectiveness 5 to 20 times that of ß-particles. Another advantage of {alpha}-particles compared with ß-particles is that they exhibit a low dependence on dose rate and oxygen enhancement effects (16, 17). In addition, {alpha}-particles have a relatively short path length in tissue and therefore only deliver low levels of radiation to normal tissues (16, 17). Among {alpha}-emitters currently under investigation for use in radioimmunotherapy, 211At is perhaps the most promising candidate for radioimmunotherapeutic applications on the basis of half-life (t1/2 = 7.2 hours) considerations. In this study, we defined the specificity and stability of the 211At-7G7/B6 and further investigated the therapeutic efficacy of the radiolabeled antibody in both a leukemia model and a lymphoma model. Our findings suggest that 211At-7G7/B6 is a potentially valuable radioimmunotherapeutic agent for the treatment of patients with CD25-expressing leukemias.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Tumor cell lines and mouse models. Kit225IG3 is a leukemic T-cell line. Karpas299 and SUDHL-1 are human anaplastic large-cell lymphoma cell lines. All of these three cell lines, which express CD25 on their cell surfaces, were grown in RPMI 1640 supplemented with 10% heat-inactivated FCS, 100 units/mL penicillin, and 100 µg/mL streptomycin in an atmosphere containing 5% CO2. Severe combined immunodeficient/nonobese diabetic (SCID/NOD) mice were purchased from The Jackson Laboratory (Bar Harbor, ME) and nude mice from National Cancer Institute (NCI)-Frederick (Frederick, MD). The karpas299 model, which was established by i.v. injection of 1 x 107 of karpas299 cells into SCID/NOD mice, served as a leukemia model. The SUDHL-1 model, which was established by s.c. injection of 1 x 107 SUDHL-1 cells in the right flank of nude mice, served as a solid tumor model. All animal experiments were done in accordance with NIH Animal Care and Use Committee guidelines.

Monoclonal antibodies. 7G7/B6 is a mouse immunoglobulin G2a (IgG2a) monoclonal antibody directed toward an epitope of the IL-2R{alpha} subunit other than the IL-2 binding site that is identified by daclizumab (18). The 7G7/B6 was purified from supernatants of a hybridoma (American Type Culture Collection, Manassas, VA) using ImmunoPure Protein A columns (Pierce, Rockford, IL). 11F11, also a mouse IgG2a monoclonal antibody, which recognizes the Shiga-like toxin II of enterohemorrhagic Escherichia (19), was used in this study as an isotype-matched control antibody. The hybridoma producing 11F11 monoclonal antibody was obtained from Dr. Alison D. O'Brien (Department of Microbiology, Uniformed Services University of Health Science, Bethesda, MD). UPC10, a murine IgG2a, which does not recognize resting or activated peripheral blood mononuclear cells or cell lines including T-cell, B-cell, and monocyte populations, was used as an nonspecific agent to block the nonspecific binding of the radiolabeled 7G7/B6 in liver and spleen in both SCID/NOD and nude mice. The plasmocytoma producing UPC10 was obtained from Michael Potter (NCI, Bethesda, MD).

Radiolabeling of monoclonal antibodies. Production and purification of 211At as well as the procedure for the labeling of the antibodies with 211At were recently reported in detail (20, 21). In brief, 211At was produced employing the 209Bi ({alpha}2n) 211At reaction by irradiating a bismuth target with an {alpha} beam from a Cycloton Corporation (Berkeley, CA) CS-30 cyclotron. The 211At was isolated as previously described (21). 7G7/B6 and 11F11 were labeled with 211At using the linker N-succinimidyl N-[4-21astatophenethyl]succinamate as previously described (20). The specific activities of 211At-7G7/B6 and 211At-11F11 were 5 to 10 µCi/µg (185-370 kBq/µg). 7G7/B6 was also labeled with 125I at a specific activity of 3 to 10 µCi/µg (111-370 kBq/µg) using the chloramine-T method.

Immunoreactivity assay. The immunoreactivity of 211At-7G7/B6 was evaluated as previously described (22) using the CD25-positive kit225IG3 cells, and compared with that of 125I-7G7/B6. Briefly, 211At-7G7/B6 (10 ng) was incubated with an increasing number of kit225IG3 cells (2 x 104-1 x 107) with or without unlabeled 7G7/B6 (25 µg/tube) inhibition at 4°C for 1 hour. After centrifugation, the supernatant was aspirated and the radioactivity bound to the cells was quantitated in a gamma counter (Wallac, Turku, Finland).

Stability testing of 211At-7G7/B6. The stability of 211At-7G7/B6 was tested in vitro under conditions similar to that expected in vivo. The 211At-7G7/B6 in PBS containing 0.25% bovine serum albumin was mixed with normal human serum at a volume ratio of 1:10. The mixture was incubated at 37°C and aliquots of the mixed sample were analyzed by size-exclusion high-performance liquid chromatography (Waters, Milford, MA) using a TSK G2000SWXL column (TOSOH Bioscience, Inc., South San Francisco, CA) equipped with an on-line NaI {gamma} detector ({gamma}RAM, IN/US System, Inc., Fairfield, NJ) at 0, 2, and 18 hours after incubation to define the integrity of the radiolabeled antibody.

Pharmacokinetic and biodistribution studies. To define and compare the clearance rates of 211At-7G7/B6 and 125I-7G7/B6 from the circulation, normal SCID/NOD mice were injected with 10 µg of 211At-7G7/B6 or 125I-7G7/B6. Serial blood samples were taken at different time points after injection and counted in a gamma counter. To determine the area under the curve (AUC), the percentage of the injected dose per gram of blood (%ID/g) was calculated and integrated using the trapezoidal rule method (GraphPad Prism version 4p, GraphPad Software, San Diego, CA). At 6 and 24 hours after injection of the radiolabeled antibodies, groups of five mice were sacrificed and the biodistribution of radioactivity was evaluated. The data were expressed as %ID/g. All mice were coinjected with 400 µg of UPC10 to block the nonspecific binding of the radiolabeled 7G7/B6 in the liver and spleen.

Therapy studies. Therapy studies were done on karpas299-bearing SCID/NOD mice at day 7 after the cell inoculation and on SUDHL-1-bearing nude mice when xenografted tumors typically reached ~0.5 cm in maximal diameter.

Groups of karpas299-bearing SCID/NOD or SUDHL-1-bearing nude mice were injected with 15 µCi of 211At-7G7/B6 or 211At-11F11, or 200 µL of PBS. Mice in the 211At-7G7/B6 and 211At-11F11 groups received coinjection of 400 µg of UPC10. Throughout the experiments, body weight and/or tumor size as well as survival of the tumor-bearing mice was monitored.

Monitoring of tumor growth. Karpas299 leukemia growth was monitored by serum levels of soluble IL-2R{alpha} (sIL-2R{alpha}), a surrogate tumor marker that was indicative of the tumor load in the murine model (23) and body weight of the karpas299-bearing mice. Measurement of the serum concentrations of the sIL-2R{alpha} was done using an ELISA. The ELISA kit was purchased from R&D Systems (Minneapolis, MN). The ELISAs were done as indicated in the kit inserts. SUDHL-1 tumor growth was monitored by measuring tumor size twice a week for 2 weeks following treatment, and then once per week. A digital caliper was used to measure the tumor in two orthogonal dimensions. The volume was calculated using the formula [(long dimension)(short dimension)2] / 2. Body weight and survival of both karpas299- and SUDHL-1-bearing mice were monitored throughout the experiments.

Expression of CD25 on karpas299 and SUDHL-1 cell surfaces. The expression of CD25 on karpas299 and SUDHL-1 cell surfaces was analyzed by flow cytometry. Aliquots of 1 x 106 karpas299 or SUDHL-1 cells were stained with a FITC-labeled anti-human CD25 antibody or a FITC-labeled isotype control antibody (PharMingen, San Diego, CA) on ice for 30 minutes. After washing, the cells were analyzed for the expression of CD25 using a FACScan flow cytometry (Becton Dickinson, San Jose, CA).

Statistical analysis. The statistical differences of the AUCs between 211At-7G7/B6 and 125I-7G7/B6 in blood and the tumor size at different time points for the different treatment groups were analyzed using one-way ANOVA with pairwise comparisons done with the Tukey test (Sigma Stat, San Rafael, CA). The statistical significance of differences in survival of the mice in different groups was determined by the log-rank test using the StatView program (Abacus Concepts, Berkeley, CA).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Immunoreactivity of 211At-7G7/B6
For a radiolabeled antibody to be effective, the labeling procedure should not compromise antibody specificity. We tested the bindability of 211At-7G7/B6 and compared it with that of 125I-7G7/B6 in vitro. Both 211At-7G7/B6 and 125I-7G7/B6 bound to kit225IG3 cells similarly with maximum binding of >80% of the added radiolabeled antibodies (Fig. 1 ). The bindings were specifically inhibited by unmodified 7G7/B6 (Fig. 1). 211At-11F11 did not bind to kit225IG3 cells (Fig. 1).


Figure 1
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  		Figure 1. Immunoreactivity of 211At-7G7/B6 and 125I-7G7/B6. The cell binding assay of the radiolabels was done as described in Materials and Methods. Both 211At-7G7/B6 and 125I-7G7/B6 bound to CD25-positive kit225IG3 cells similarly and the bindings were inhibited by the addition of a 1,000-fold greater concentration of unmodified 7G7/B6. 211At-11F11 did not bind to kit225IG3 cells.

 
In vitro Stability of 211At-7G7/B6
In a radiolabeled monoclonal antibody, the radionuclide must be tightly linked to the antibody, eliminating premature release of the radionuclide in vivo, which would lead to unacceptable toxicity and reduced efficacy. We tested the stability of 211At-7G7/B6 in vitro. The high-performance liquid chromatography analysis showed a single peak of radioactivity with a retention time of ~8 minutes, which corresponded to the IgG peak detected by UV at 280 nmol/L for 211At-7G7/B6 at all time points tested (data not shown).

Pharmacokinetics and Biodistribution of 211At-7G7/B6
To test the stability of 211At-7G7/B6 in vivo, we did pharmacokinetic and biodistribution studies with 211At-7G7/B6 and compared the results with those obtained with 125I-7G7/B6 in SCID/NOD mice. The blood clearance rates of 211At-7G7/B6 and 125I-7G7/B6 were very similar in SCID/NOD mice (Fig. 2 ). The AUCs were 691 ± 66 for 211At-7G7/B6 and 730 ± 78 for 125I-7G7/B6 (P = 0.6). The biodistributions of both radiolabels were also similar with the exception of higher stomach uptake of radioactivity with 211At-7G7/B6 (Fig. 3 ).


Figure 2
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  		Figure 2. Comparison of the blood clearance rate of 211At-7G7/B6 with that of 125I-7G7/B6 in SCID/NOD mice. Points, mean %ID/g; bars, SD. The blood clearance rates of 211At-7G7/B6 and 125I-7G7/B6 in SCID/NOD mice were very similar. The AUCs were 691 ± 66 for 211At-7G7/B6 and 730 ± 78 for 125I-7G7/B6 (P = 0.6).

 

Figure 3
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  		Figure 3. Comparison of the biodistribution of 211At-7G7/B6 with that of 125I-7G7/B6 in SCID/NOD mice. Biodistributions of 211At-7G7/B6 and 125I-7G7/B6 were very similar in SCID/NOD mice except for the higher stomach uptake of radioactivity with 211At-7G7/B6 when compared with that of 125I-7G7/B6. Columns, mean %ID/g; bars, SD.

 
Therapy Study
Karpas299 model. The therapeutic study was done in the karpas299 model. The i.v. administration of a single dose of 15 µCi of 211At-7G7/B6 inhibited the leukemia growth significantly as seen by the reduced serum levels of sIL-2R{alpha} in the 211At-7G7/B6 treatment group when compared with the control or radiolabeled nonspecific antibody group, 211At-11F11 (Fig. 4A ). At day 20 after therapy, the concentrations of serum sIL-2R{alpha} were 37,016 and 5,796 pg/mL in the control and 211At-11F11 groups, respectively, whereas it was undetectable in the 211At-7G7/B6 group (Fig. 4A). Furthermore, survival of the mice in the 211At-7G7/B6 group was significantly prolonged as compared with the control and 211At-11F11 groups (Fig. 4B; P < 0.001). All of the mice in the control and 211At-11F11 groups died by day 46. In contrast, 5 of 15 mice in the 211At-7G7/B6 group survived for >5 months when we terminated the experiment. Three of the five mice had no detectable serum sIL-2R{alpha}. The mean survival duration was 24 days in the control group and 32 days in the 211At-11F11 group, whereas the mean survival duration was prolonged to >76 days in the 211At-7G7/B6 group.


Figure 4
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  		Figure 4. Therapeutic study of 211At-7G7/B6 in karpas299 leukemia model. A, serum sIL-2R{alpha} levels of the karpas299 leukemia–bearing SCID/NOD mice at day 20 after therapy. B, Kaplan-Meier survival plot of the karpas299 leukemia–bearing SCID/NOD mice. At day 20 after therapy, serum sIL-2R{alpha} was undetectable in the 211At-7G7/B6 treatment group. In contrast, the mean serum concentrations of sIL-2R{alpha} were 37,016 and 5,796 pg/mL in the control and 211At-11F11 groups, respectively. Survival of the karpas299 leukemia–bearing mice was significantly prolonged in the 211At-7G7/B6 group when compared with those in the control and 211At-11F11 groups (P < 0.001).

 
The specificity of the therapeutic effect was confirmed by comparison of the survival of the mice in the 211At-7G7/B6 and 211At-11F11 groups. Survival of the mice was significantly prolonged with the specific radioimmunotherapy, 211At-7G7/B6, when compared with mice receiving nonspecific radioimmunotherapy, 211At-11F11 (Fig. 4B; P < 0.001).

SUDHL-1 model. The therapeutic study of 211At-7G7/B6 was also done in the SUDHL-1 model, which served as a solid tumor model. Both karpas299 and SUDHL-1 cells express CD25 on their cell surfaces strongly (Fig. 5 ) and they were similarly sensitive to in vitro irradiation (data not shown). However, unlike that in karpas299 model, 15 µCi of 211At-7G7/B6 only showed modest inhibition of the SUDHL-1 tumor growth in vivo, as seen by the tumor size results (Fig. 6A ) and prolongation of the survival of the tumor-bearing mice, although it was statistically significantly different when compared with the control group (Fig. 6B; P < 0.01). The mean survival duration was 11 days in the control group, 21 days in the 211At-7G7/B6, and 23 days in the 211At-11F11 group, respectively. There were no differences in tumor size and survival of the mice between the 211At-7G7/B6 and 211At-11F11 groups. Therefore, the results from this study indicate that, because of the short distance of action and relatively short half-life of 211At as well as the slow tumor uptake and slow penetration of the radiolabeled antibody into solid tumor masses, antibodies armed with {alpha}-emitting radionuclides are superior for treating leukemias.


Figure 5
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  		Figure 5. CD25 expression on the cell surfaces. Both karpas299 and SUDHL-1 cells strongly express CD25 on their cell surfaces.

 

Figure 6
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  		Figure 6. Therapeutic study of 211At-7G7/B6 in the SUDHL-1 solid tumor model. A, tumor volume. B, Kaplan-Meier survival plot of the SUDHL-1-bearing nude mice. Both 211At-7G7/B6 and 211At-11F11 inhibited the SUDHL-1 lymphoma growth as seen by tumor size and prolonged survival (P < 0.01) of the SUDHL-1-bearing mice when compared with the control group. However, there were no differences in tumor size and survival of the SUDHL-1-bearing mice between 211At-7G7/B6 and 211At-11F11 groups (P > 0.4). *, P < 0.05; **, P < 0.001, when control group was compared with either the 211At-7G7/B6 or 211At-11F11 group.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
The hybridoma technique rekindled interest in the use of antibodies targeted to cell-surface antigens to treat cancer patients. However, such monoclonal antibodies have been largely ineffective with only a few exceptions in this arena. Monoclonal antibodies armed with toxins or radionuclides to specifically target these cytotoxic agents to tumor cells provide a valuable augmentation of their therapeutic efficacy (24–26).

One pivotal issue to be addressed in all systemic radioimmunotherapy trials is the selection of a monoclonal antibody that targets the tumor and thereby defines the type of malignancy chosen as the target for radioimmunotherapy. In the present study, we have chosen an epitope of the human IL-2R{alpha} identified by 7G7/B6 as our target for radioimmunotherapy. The scientific basis for this choice of the {alpha} subunit of the IL-2R is that virtually no normal resting cells express this receptor subunit whereas this receptor is expressed by a high proportion of the abnormal cells in certain forms of lymphoid neoplasms, including adult T-cell leukemia and anaplastic large-cell lymphoma (1, 2, 27). In addition, 7G7/B6 recognizes a different epitope of IL-2R{alpha} other than that identified by daclizumab; therefore, these two antibodies potentially can be combined for the treatment of CD25-expressing leukemias and lymphomas. A series of modifications of the anti-Tac monoclonal antibody have been made to increase its effector function, to reduce its immunogenicity, and to improve its pharmacokinetics. To increase its therapeutic effect, daclizumab has been armed with the ß-emitting radionuclide, 90Y (3). Whereas daclizumab armed with this radionuclide provided meaningful therapy for human T-cell lymphotrophic virus I–associated adult T-cell leukemia, only 2 of the 16 patients treated manifested a complete remission (3). Therefore, more effective therapeutic approaches are needed to be developed.

As noted above, a second component of an optimal radioimmunotherapeutic regimen is the choice of chelating agent or linker to couple the radionuclide to the monoclonal antibody. An ideal agent should not alter the specificity of binding of the monoclonal antibody to its antigenic target, nor should it damage the antibody and thus alter its rate of catabolism or pattern of tissue distribution. It should hold the radionuclide tightly so that there is no premature loss of the radionuclide from the monoclonal antibody in vivo. The agent N-succinimidyl N-[4-21astatophenethyl]succinamate used in the present study for linking 211At fulfills each of these requirements (20, 21).

A third pivotal issue in defining an optimal radioimmunotherapeutic agent is to consider the nature of the radionuclide used in relationship to the nature of the diseases being treated. A ß-emitting radionuclide such as 90Y, which acts through crossfire, may be preferable in the treatment of large tumor masses as occurs with the SUDHL-1 model (5). In the clinical situation, this agent may eliminate nontargeted tumor cells through the crossfire effect emanating from neighboring antigen-bearing cells that have been targeted by the radiolabeled monoclonal antibody (26). Nevertheless, the use of ß-emitting radionuclides is limited because as the target mass decreases, the benefit of the crossfire effect also decreases while the potential for normal tissue damage increases. With small tumors including micrometastases, individual tumor cells, and leukemic cells, the therapeutic efficacy of ß-emitters may be limited. This is because high-energy ß-emitting radionuclides such as 90Y deliver a high dose of irradiation to normal tissues due to the long range of ß irradiation. For such cellular populations, the development of radiolabeled monoclonal antibody–mediated approaches may focus on {alpha}-emitting radionuclides, which may be the most effective agents at killing isolated leukemic cells without damaging normal tissues. Radionuclides emitting {alpha}-particles have a high linear energy transfer (LET 6-9 MeV particles), which act over 10 to 80 µm and are effective at killing individual target cells (28). A phase I clinical trial with 213Bi-HuM195 in the treatment of patients with relapsed and refractory acute or chronic myelogenous leukemia was reported to show promising results (29). The present study focused on 211At of which the physical half-life of 7.2 hours may be adequate to obtain effective leukemic cell targeting. This radionuclide is of special value for antibodies, such as 7G7/B6, that have been shown to stay on the lymphocyte cell surface and to not be rapidly internalized by the target cell, a process that might lead to loss of the 211At from its target arena. In the present study, we showed that 211At linked to the 7G7/B6 monoclonal antibody provided effective therapy for the karpas299 leukemia. In contrast, as predicted from its short distance of action and relatively short half-life, as well as the short time required for antibody to reach the tumor, 211At linked to 7G7/B6 did not provide effective therapy for the SUDHL-1 model.

In the present study, we considered an additional issue involved in systemic radioimmunotherapy. We addressed the limitation inherent in the use of a single monoclonal antibody to simultaneously saturate receptors for antibody-dependent cellular cytotoxicity and cytokine deprivation and to deliver a high proportion of the administered radionuclide 211At at high specific activity to the tumor cells. To this end, we used 7G7/B6 to carry the radionuclide. This is a required initial study for the ultimate use of 7G7/B6 at a low concentration as the agent that carries the radionuclide at high specific activity to be used with simultaneously administered saturating doses of the non-cross-competing antibody, daliczumab, to provide the receptor saturation required for antibody-dependent cellular cytotoxicity and cytokine deprivation. In the present study, as just noted, 211At-7G7/B6 showed its specificity and stability for use as a stand-alone therapeutic. Furthermore, the therapeutic results with a single dose of 211At-7G7/B6 in the CD25-expressing karpas299 model were encouraging. These results provide the scientific basis for the subsequent combination therapy using 211At-7G7/B6 in association with receptor-saturating doses of daliczumab to provide the desired two independent cytotoxic actions. Thus, the results of the present study suggested that 211At-7G7/B6 is a potential therapeutic agent for the treatment of patients with CD25-expressing leukemias.


   Acknowledgments
 
Grant support: Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 4/ 1/06. Revised 5/25/06. Accepted 6/15/06.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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M. Zhang, Z. Yao, H. Patel, K. Garmestani, Z. Zhang, V. S. Talanov, P. S. Plascjak, C. K. Goldman, J. E. Janik, M. W. Brechbiel, et al.
Effective therapy of murine models of human leukemia and lymphoma with radiolabeled anti-CD30 antibody, HeFi-1
PNAS, May 15, 2007; 104(20): 8444 - 8448.
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