Astatine-211 Conjugated To Anti-CD20 Monoclonal Antibody Eliminates B-Cell Lymphoma In a Mouse Model

Stem cell rescue after myeloablative doses of beta particle emitting radiolabeled monoclonal antibodies targeting CD20 antigen can lead to remissions in up to 95% of lymphoma patients who previously failed conventional combination chemotherapy. While encouraging, toxicities with beta particle radioimmunotherapy (RIT) are significant and ∼50% of patients ultimately relapse. Higher doses of absorbed radiation to tumors have correlated with a reduced risk of disease recurrence but dose limiting toxicities prevent escalation. A potential advantage of alpha particle emitting radionuclides is the release of large amounts of energy linearly over a few cell diameters (∼50-80 mm) resulting in irreparable double-strand DNA breaks that overwhelm cellular repair mechanisms. As a result, alpha particles confer a unique capacity to kill individual targeted cells while causing minimal radiation damage to surrounding tissues.

To explore alpha-emitter conjugated anti-CD20 RIT, a closo-decaborate(2-) [B10] labeling reagent was developed as a radiolabeling platform capable of providing critical stability to alpha particle-labeled biomolecules. ²¹¹At was chosen as a therapeutic radionuclide based on its high linear energy transfer, biologically relevant t_1/2 (∼7.2 hr) and absence of toxic daughter radionuclide decay byproducts. Cell binding assays using CD20-expressing Ramos tumor cells demonstrated that B10 conjugated to the anti-CD20 monoclonal antibody (mAb) 1F5 (B10-1F5) was specific and that astatination did not impair antibody binding (²¹¹At-B10-1F5 binding was equivalent to ¹²⁵I-B10-1F5). Blood clearance was similar for ¹²⁵I-1F5, ²¹¹At-B10-1F5 and ¹²⁵I-B10-1F5 in athymic nude- Foxn1^nu mice (23.87 ±5.13 % injected dose/gram [ID/g], 19.41 ±3.27 %ID/g and 19.80 ±1.44% ID/g respectively) at 17 hr post infusion. In tissue biodistribution studies using nude mice bearing Ramos flank tumor xenografts (10 x10⁶ cells injected) radioactivity was measured in blood, tumor and nonspecific organs harvested 24 hours after injection (n=5/group). Animals received either ¹²⁵I-1F5, ²¹¹At-B10-1F5 and ¹²⁵I-B10-1F5 (co-injected) or isotype matched control mAb ²¹¹At- B10-HB8181 and¹²⁵I-B10-HB8181(co-injected). Measured activity in tumors was three-fold higher for ¹²⁵I-1F5, ¹²⁵I-B10-1F5 and ²¹¹At-B10-1F5 (7.62 ± 2.09%ID/g, 7.53 ± 1.59%ID/g and 9.28 ± 1.85%ID/g respectively) than for ¹²⁵I-B10-HB8181 and ²¹¹At-B10-HB8181 controls (2.87 ± 0.35%ID/g and 3.45 ± 0.58%ID/g respectively). In non-target organs no appreciable difference in measured activity was seen with either ²¹¹At- or ¹²⁵I-labeled B10-1F5 and their respective controls.

Subsequent therapy studies performed in nude mice bearing Ramos flank tumor xenografts demonstrated only a moderate survival advantage after ²¹¹At-B10-1F5 [data not shown]. This finding was consistent with the hypothesis that the alpha particle's short path length may not be ideally suited to models of “bulky” disease. In contrast, therapy studies using disseminated Ramos and Granta tumor cells introduced into NOD-SCID mice suggest a role for alpha particle based RIT in “non-bulky” disease models. In these studies NOD-SCID animals received 1x10⁶ tumor cells iv 48 hours prior to ²¹¹At-B10-1F5 (10 μCi or 15 μCi) or control ²¹¹At-B10-HB8181 (n=10/group). Stem cell rescue was performed 48 hours after the RIT. Eighty days after treatment 80% of animals receiving 15 μCi of ²¹¹At-B10-1F5 and 70% of animals in the 10 μCi treatment group were alive and tumor free while no animals in the non-binding ²¹¹At-B10-HB8181 (10 μCi or 15 μCi) control groups survived beyond day 47 (Figure). Blood counts, serum creatinine and transaminase levels measured in ²¹¹At-B10-1F5 treated animals ∼180 days after RIT demonstrated no significant long-term bone marrow, renal or liver toxicity.

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²¹¹At-B10-1F5 can eliminate CD20 expressing tumor cells in this mouse model and further study appears to be warranted.