astatine and Breast-Neoplasms

The sigma-2 receptor is a protein with a Heme binding region and is capable of receptor-mediated endocytosis. It is overexpressed in many cancers making it a potential vector for therapeutic drug delivery. Our objective was to introduce an alpha-emitting radionuclide, astatine-211, into a selective sigma-2 ligand moiety to provide cytotoxic capabilities without adversely altering the pharmacological characteristics. In this study we investigated the in vitro/in vivo tumor targeting and estimated dosimetry of alpha-emitting sigma-2 ligand, 5-(astato-(211)At)-N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2,3-dimethoxybenzamide ((211)At-MM3), in a pre-clinical human breast cancer model.. Astatine-211 was produced in a cyclotron and isolated by dry distillation. Radiosynthesis of (211)At-MM3 was performed using a tin precursor through radioastatodestannylation. In vitro sigma-2 binding experiments using (211)At-MM3 were carried out in live EMT6 and MDA-MB-231 breast cancer cells and liver homogenate tissue. In vivo biodistribution experiments were performed using EMT6 mouse breast cancer cells in BALB/c female mice. Approximately 370 kBq of (211)At-MM3 was administered intravenously and at time points of 5 min, 1, 2, 4, 8, and 24 h organs/tissue were harvested. Estimated human dosimetry was extrapolated from biodistribution data using OLINDA/EXM (VU e-Innovations).. Astatine-211 was successfully produced and isolated in quantities suitable for in vitro and small animal in vivo experiments. Radiosynthesis of (211)At-MM3 was reproducible with high radiochemical purity. Astatine-211-MM3 exhibited picomolar affinity to the sigma-2 receptor in contrast to the iodinated analog that had nanomolar affinity. Prolonged tumor targeting was measured through biodistribution studies with a maximal tumor to muscle ratio of 9.02 at 4h. Estimated human dosimetry revealed doses of up to 370 MBq in an adult female patient were below organ radiation limits with the potential to provide a high therapeutic dose to tumors.. The sigma-2 receptor could serve as a suitable targeting platform for designing radiotherapeutics. (211)At-MM3 showed tumor targeting properties in vitro/in vivo and favorable estimated human dosimetry establishing the proof of concept for future development as a radiotherapeutic for the treatment of breast cancer.

Topics: Alpha Particles; Animals; Astatine; Benzamides; Breast Neoplasms; Cell Line, Tumor; Cell Transformation, Neoplastic; Female; Humans; Mice; Molecular Targeted Therapy; Radiometry; Receptors, sigma; Tissue Distribution

Nonuniform dose distributions among disseminated tumor cells can be a significant limiting factor in targeted α therapy. This study examines how cocktails of radiolabeled antibodies can be formulated to overcome this limitation.. Cultured MDA-MB-231 human breast cancer cells were treated with different concentrations of a cocktail of 4 fluorochrome-conjugated monoclonal antibodies. The amount of each antibody bound to each cell was quantified using flow cytometry. A spreadsheet was developed to "arm" the antibodies with any desired radionuclide and specific activity, calculate the absorbed dose to each cell, and perform a Monte Carlo simulation of the surviving fraction of cells after exposure to cocktails of different antibody combinations. Simulations were performed for the α-particle emitters (211)At, (213)Bi, and (225)Ac.. Activity delivered to the least labeled cell can be increased by 200%-400% with antibody cocktails, relative to the best-performing single antibody. Specific activity determined whether a cocktail or a single antibody achieved greater cell killing. With certain specific activities, cocktails outperformed single antibodies by a factor of up to 244. There was a profound difference (≤16 logs) in the surviving fraction when a uniform antibody distribution was assumed and compared with the experimentally observed nonuniform distribution.. These findings suggest that targeted α therapy can be improved with customized radiolabeled antibody cocktails. Depending on the antibody combination and specific activity of the radiolabeled antibodies, cocktails can provide a substantial advantage in tumor cell killing. The methodology used in this analysis provides a foundation for pretreatment prediction of tumor cell survival in the context of personalized cancer therapy.

Topics: Actinium; Algorithms; Alpha Particles; Antibodies; Astatine; Bismuth; Breast Neoplasms; Cell Line, Tumor; Cell Survival; Drug Combinations; Female; Humans; Radioimmunotherapy; Radioisotopes; Radiopharmaceuticals

Carcinomatous meningitis (CM) is a devastating disease characterized by the dissemination of malignant tumor cells into the subarachnoid space along the brain and spine. Systemic treatment with monoclonal antibody (mAb) trastuzumab can be effective against HER2-positive systemic breast carcinoma but, like other therapies, is ineffective against CM. The goal of this study was to evaluate the therapeutic effect of alpha-particle emitting (211)At-labeled trastuzumab following intrathecal administration in a rat model of breast carcinoma CM.. Athymic rats were injected intrathecally with MCF-7/HER2-18 breast carcinoma cells through a surgically implanted indwelling intrathecal catheter. In Experiment 1, animals received 33 or 66 muCi (211)At-labeled trastuzumab, cold trastuzumab or saline. In Experiment 2, animals were inoculated with a lower tumor burden and received 46 or 92 muCi (211)At-labeled trastuzumab or saline. In Experiment 3, animals received 28 muCi (211)At-labeled trastuzumab, 30 muCi (211)At-labeled TPS3.2 control mAb or saline. Histopathological analysis of the neuroaxis was performed at the end of the study.. In Experiment 1, median survival increased from 21 days for the saline and cold trastuzumab groups to 45 and 48 days for 33 and 66 muCi (211)At-labeled trastuzumab, respectively. In Experiment 2, median survival increased from 23 days for saline controls to 68 and 92 days for 46 and 92 muCi (211)At-labeled trastuzumab, respectively. In Experiment 3, median survival increased from 20 days to 29 and 36 days for animals treated with (211)At-labeled TPS3.2 and (211)At-labeled trastuzumab, respectively. Long-term survivors were observed exclusively in the (211)At-trastuzumab-treated groups.. Intrathecal (211)At-labeled trastuzumab shows promise as a treatment for patients with HER2-positive breast CM.

Topics: Alpha Particles; Animals; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Astatine; Breast Neoplasms; Cell Line, Tumor; Female; Humans; Injections, Spinal; Isotope Labeling; Meningeal Carcinomatosis; Mice; Radioimmunotherapy; Rats; Receptor, ErbB-2; Survival Rate; Trastuzumab; Xenograft Model Antitumor Assays

Successful radioimmunotherapy strategies depend on selecting radioisotopes with physical properties complementary to the biological properties of the targeting vehicle. Small, engineered antitumor antibody fragments are capable of rapid, highly specific tumor targeting in immunodeficient mouse models. We hypothesized that the C6.5 diabody, a noncovalent anti-HER2 single-chain Fv dimer, would be an ideal radioisotope carrier for the radioimmunotherapy of established tumors using the short-lived alpha-emitting radioisotope (211)At.. Immunodeficient nude mice bearing established HER2/neu-positive MDA-MB-361/DYT2 tumors treated with N-succinimidyl N-(4-[(211)At]astatophenethyl)succinamate ((211)At-SAPS)-C6.5 diabody. Additional cohorts of mice were treated with (211)At-SAPS T84.66 diabody targeting the carcinoembryonic antigen or (211)At-SAPS on a diabody specific for the Müllerian inhibiting substance type II receptor, which is minimally expressed on this tumor cell line.. A single i.v. injection of (211)At-SAPS C6.5 diabody led to a 30-day delay in tumor growth when a 20 muCi dose was administered and a 57-day delay in tumor growth (60% tumor-free after 1 year) when a 45 muCi dose was used. Treatment of mice bearing the same tumors with (211)At-SAPS T84.66 diabody at the same doses led to a delay in tumor growth, but no complete responses, likely due to substantially lower expression of this antigen on the MDA-MB-361/DYT2 tumors. In contrast, a dose of 20 muCi of (211)At-SAPS on the anti-Müllerian-inhibiting substance type II receptor diabody did not affect tumor growth rate, demonstrating specificity of the therapeutic effect.. These findings indicate that diabody molecules can be effective agents for targeted radioimmunotherapy of solid tumors using powerful, short-lived alpha-emitting radioisotopes.

Topics: Animals; Astatine; Breast Neoplasms; Female; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Inbred ICR; Mice, Nude; Mice, SCID; Peptide Fragments; Radioimmunotherapy; Receptor, ErbB-2; Tissue Distribution; Transplantation, Heterologous

The anti-ErbB2 antibody trastuzumab is used for the treatment of patients with advanced breast cancer, resulting in a response rate of 40-60%. Coupling with a cytotoxic nuclide, e.g. alpha-emitting 211At, may further increase tumour response. The tumour-targeting properties of trastuzumab, astatinated using N-succinimidyl-para-(tri-n-methylstannyl)-benzoate, were evaluated and compared with those of radioiodinated trastuzumab in this study. We found that astatinated trastuzumab retains high specificity towards ErbB2. While the immunoreactive fraction of radioiodinated trastuzumab was higher than that of astatinated trastuzumab (76+/-9% versus 54+/-28%), both radioconjugates showed high affinity (KD 0.75+/-0.16 nM versus 1.8+/-0.3 nM). A growth inhibition study indicated a dose-dependent cell deactivation, in which approximately 74 cell-associated astatine decays per cell gave a survival fraction of 4.5+/-0.8x10(-4). Results of a comparative animal study on normal mice gave no indication that astatination would have any adverse effects on the biodistribution of the antibody. In conclusion, the results of the study suggest that astatinated trastuzumab is a promising candidate for treating ErbB2-expressing tumours.

Topics: Animals; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Astatine; Binding, Competitive; Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Cell Survival; Female; Humans; Liver; Mice; Mice, Inbred Strains; Radioimmunotherapy; Receptor, ErbB-2; Spleen; Time Factors; Tissue Distribution; Trastuzumab

Radioimmunotherapy with anti-HER2 monoclonal antibodies (mAbs) such as trastuzumab is a promising strategy for treating HER2-positive breast and ovarian carcinoma patients. The objective of this study was to determine the cytotoxic effectiveness of trastuzumab labeled with the 7.2-h half-life alpha-particle emitter 211At.. Experiments were performed on SKBr-3, BT-474 and the transfected MCF7/HER2-18 human breast carcinoma cell lines. Intrinsic radiosensitivity was determined after exposure to external beam irradiation. The cytotoxicity of 211At-labeled trastuzumab was measured by clonogenic assays. The distribution of HER2 receptor expression on the cell lines was measured using fluorescence-activated cell sorting. A pharmacokinetic (PK)/microdosimetric model was established to assess the effects of specific activity (SA), HER2 receptor expression and absorbed dose on survival fraction (SF).. With external beam irradiation, the 2-Gy SF for BT-474, SKBr-3 and MCF7/HER2-18 cells was 0.78, 0.53 and 0.64 Gy, respectively. Heterogeneous HER2 expression was observed, with a subpopulation of cells lacking measurable receptor (14.5%, SKBr-3; 0.34%, MCF-7/HER2; 1.73%, BT-474). When plotted as a function of activity concentration, SF curves were biphasic and inversely proportional to SA; however, when the model was applied and absorbed doses calculated, the SF curve was monoexponential independent of SA. Thus, the PK model was able to demonstrate the effects of competition between cold and labeled mAb. These studies showed that the relative biological effectiveness of 211At-labeled trastuzaumab was about 10 times higher than that of external beam therapy.. These in vitro studies showed that 211At-labeled trastuzumab mAb is an effective cytotoxic agent for the treatment of HER2-positive tumor cells. The SA of the labeled mAb and the homogeneity of HER2 receptor expression are important variables influencing the efficiency of cell killing.

Topics: Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Astatine; Breast Neoplasms; Cell Survival; Colony-Forming Units Assay; Humans; Immunoconjugates; In Vitro Techniques; Receptor, ErbB-2; Trastuzumab; Tumor Cells, Cultured