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Background: Internal mammary lymph nodes (IMNs) are a potential site of metastasis for breast cancer. Targeting IMNs as part of a comprehensive regional nodal irradiation comes at the cost of higher unwanted doses to critical nearby organs at risk. Thus, the efficacy and safety of different radiotherapy techniques for the coverage of this area remain elusive. Objectives: We present a dosimetric comparison between partially wide tangents (PWT) and direct photon/electron (P/E) portals in terms of target volume coverage and normal tissue sparing. Methods: Patients with left-sided breast cancer, who were referred to our clinic for post-lumpectomy or post-mastectomy radio-therapy, underwent computed tomography (CT) simulation. The left breast and IMNs, heart, lung, right breast, and esophagus were contoured. Dosimetric comparisons were based on dose-volume histograms (DVHs) generated for all of the aforementioned organs. A subgroup analysis was also performed based on patients' type of surgical treatment. Results: A total of 30 patients (10 with breast conserving surgery and 20 with modified radical mastectomy) were included. The P/E plan provided a higher coverage of the left breast (P-value of CTV V105%: < 0.001) and IMNs with a P-value of 0.087 regarding the mean dose received by IMNs, and also less volume of the heart (P-value of Heart V30Gy: 0.021), and lungs (P-value of Lung V20Gy: 0.003) were irradiated. However, these advantages came at the cost of a higher dose to the esophagus and right breast and more hotspots compared to the PWT technique. Conclusions: The P/E technique had advantages regarding target volume coverage and toxicity regardless of the type of surgery. Based on the results of this study, overall, the P/E portal is superior to the PWT for radiotherapy of breast cancer with internal mammary node inclusion. However, the appropriate treatment plan should be decided on a case-by-case basis. [ABSTRACT FROM AUTHOR]

Abstract: Aim: PR81 is a monoclonal antibody that binds with high affinity to MUC1 antigen that is over expressed in 80% of breast cancers. In this study, we developed a method for indirect labeling of PR81 with lutetium-177 and performed all preclinical qualifications in production of a biologic agent for radioimmunotherapy of breast cancer. Materials and Methods: The radiochemical purity and in vitro stability of 177Lu labeled PR81 was determined by instant thin layer chromatography. The immunoreactivity and cell toxicity of the complex were tested on MCF7 cell line. The biodistribution and scintigraphy studies were performed in BALB/c mice with breast tumor. Results: The radiochemical purity was 91.2±3.8% after 2 h. The in vitro stabilities in phosphate buffer and human blood serum were 83.1±3.4% and 76.2±3.6% at 96 h, respectively. The immunoreactivity of the complex was 83.4±2.4%. The cell toxicity study showed that the complex inhibited 85.2±3.4% growth of MCF7 cells at a concentration of 2500 ng/ml after 96 h. The biodistribution and scintigraphy studies showed the accumulation of the complex at the site of tumors with high sensitivity and specificity. Conclusion: The results showed that one may consider 177Lu-DOTA–PR81 as a potential radiopharmaceutical for therapy of human breast cancer, which needs further investigations. [Copyright &y& Elsevier]

Objective: We digested anti-MUC1 monoclonal antibody PR81 to produce F(ab′) fragments. A comparison was performed between the two radiolabeled PR81 and F(ab′) fragments for breast tumor imaging in a mouse model. Methods: The optimum conditions for pepsin digestion of PR81 were investigated in terms of enzymes: antibody ratio, digestion time duration and preserved immunoreactivity of the produced fragments. The F(ab′) fragments were labeled with Technetium-99m using HYNIC as a chelator and tricine as a co-ligand. The immunoreactivity of the complexes was assessed by radioimmunoassay using MCF7 cells. Biodistribution and imaging studies were performed in female BALB/c mice with breast tumor xenograft at 4, 8 and 24 h post-administration. The PR81 was labeled with technetium-99m in the same way for comparison. Results: The optimum time duration for PR81 digestion was found to be 28 h at an enzyme:antibody weight ratio of 1:20 that resulted in 95.2 ± 4.7% purity. The labeling of intact PR81 and its F(ab′) fragments were 87.6 ± 4.2 and 76.1 ± 3.3% after 1 h, respectively ( p value <0.05). The percentage of immunoreactivity of F(ab′) fragments and intact PR81 were 75.4 ± 2.1% and 85.7 ± 2.9%, respectively ( p value <0.05). The biodistribution and imaging studies demonstrated localization of the fragments at 4 h post-administration with high sensitivity and specificity. Conclusion: The results showed that F(ab′) fragment of PR81 is more suitable than intact PR81 for safer and more rapid detection of human breast cancer. [ABSTRACT FROM AUTHOR]

PR81 is a monoclonal antibody that binds with high affinity to MUC1, which is over expressed on breast and other tumors. The objective of this study was to compare the two labeling methods (direct and indirect radioiodination) for application of this antibody against MUC1 as a radioimmunotherapeutical agent.Monoclonal antibody (PR81) against the tandem repeat of the core protein (MUC1) was prepared, characterized, purified, and labeled with ^{131}I using the direct (chloramin-T) and indirect (Fmoc-D-Tyr (tBu)-D-Tyr (tBu)-D-Lys (Boc)-OH (YYK) attached to N-hydroxysuccinimide as a linker between PR81 and ^{131}I) methods. The immunoreactivity of ^{131}I-PR81 and ^{131}I-TP-PR81 complexes with MUC1 (the native protein), BSA-P_{20} (a 20 amino acid corresponding the tandem repeat of MUC1) and MCF7 cell line were performed by RIA. In vitro stability of ^{131}I-PR81 and ^{131}I-YYK-peptide-PR81 complexes in human serum was determined by thin layer chromatography (TLC). Cell toxicity and in vitro internalization studies were performed with the MCF7 cell line, and the tissue biodistribution of the ^{131}I-PR81 and ^{131}I- YYK-peptide -PR81 complexes was evaluated in normal BALB/c mice at 4, 24 and 48 hrs. The labeling efficiency was determined by measuring the percentage recovery of radioactivity in the final product relative to the initial activity in the shipment vial, was found to be 59.9% ± 7.9% for direct and 50% ± 3.2% for indirect methods. ^{131}I-PR81 and ^{131}I- YYK- peptide -PR81 complexes showed high immunoreactivity towards MUC1 protein, BSA-P_{20} and MCF7 cell line. In vitro stability of the labeled products in human serum which was measured by thin layer chromatography (TLC) was found to be more than 50% over 24 hr for ^{131}I-PR81 and 70% for ^{131}I- YYK-peptide -PR81 complexes. Cell toxicity and in vitro nternalization studies showed that the ^{131}I-PR81 and ^{131}I- YYK-peptide -PR81 complexes inhibited 80% growth of the MCF7 cultured cell lines in vitro in a high concentration and up to 40% of the ^{131}I-PR81 and 60% of the ^{131}I- YYK-peptide -PR81 complexes internalized after 24 h. Biodistribution studies were performed in normal BALB/c mice at 4, 24 and 48 hrs post-injection. Thyroid and stomach levels from PR81 labeled with ^{131}I- YYK-peptide were two- to three- fold less than those with directly labeled ^{131}I-PR81, suggesting low recognition of its D-iodotyrosine residue by endogenous deiodinase. These results show that the indirect labeling was better than the indirect labeling and ^{131}I- YYK-peptide -PR81 may be considered as a promising candidate for therapy of breast cancer. [ABSTRACT FROM AUTHOR]

Radioimmunoscintigraphy (RIS) has found widespread clinical application in tumor diagnosis. The antibody (Ab) PR81 is a new murine anti-MUC1 monoclonal antibody (MAb) against human breast carcinoma. In this study a very simple, rapid and efficient method for labeling of this MAb with 99mTc, particularly suitable for development of a 'kit'is described. The reduction of Ab was performed with 2- mercaptoethanol (2-ME) at a molar ratio of 2000:1 (2-ME:MAb) and the reduced Ab was labeled with 99mTc via methylene diphosphonate (MDP) as a transchelator. The labeling efficiency which was determined by instant thin layer chromatography (ITLC) was 94.2%±2.3. Radiocolloides measured by cellulose nitrate electrophoresis were 2.5%±1.7. In vitro stability of the labeled product in human serum which was measured by gel filtration chromatography (FPLC) was 70%±5.7 over 24 hr. The integrity of labeled MAb was checked by means of SDS-PAGE and no significant fragmentation was observed. The results of the cell-binding studies showed that both labeled and unlabeled PR81 were able to compete for binding to MCF 7 cells. Biodistribution studies were performed in normal BALB/c mice at 4 and 24 hrs post-injection and no important accumulation was observed in vital organs. These results show that the new radiopharmaceutical may be considered as a promising candidate for imaging of breast cancer. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: In the present study, Herceptin was labeled with lutetium-177 via DOTA, and the necessary preclinical quality control tests (in vitro and in vivo) were performed to evaluate its use as a radioimmunotherapy agent. Material and Methods: Herceptin was conjugated to DOTA as a chelator in three different conjugation buffers (ammonium acetate, carbonate and HEPES buffer); each of the resulting conjugates was compared with respect to in vitro characteristics such as number of chelates per antibody, incorporated activity, immunoreactivity and in vitro stability in PBS buffer and blood serum. The biodistribution study and gamma camera imaging were performed in mice bearing breast tumors. To assess the therapeutic effects of 177Lu-Herceptin, cytotoxicity was investigated for 7 days in a SKBr3 breast cancer cell line. Results: Carbonate buffer was the best conjugation buffer (number of chelates per antibody: 6; incorporated activity: 81%; immunoreactivity: 87%; buffer stability: 86%; serum stability: 81%, after 4 days). The efficient tumor uptake observed in the biodistribution studies was consistent with the gamma camera image results. At a concentration of 4 μg ml−1, 177Lu-Herceptin (surviving cells: 5±0.6% of the total cells) of the total cells corresponded to an approximately eightfold increase in cytotoxicity in comparison to unmodified Herceptin (surviving cells: 43±3.9%). Conclusion: The new complex described herein could be considered for further evaluation in animals and potentially in humans as a radiopharmaceutical for use in the radioimmunotherapy of breast cancer. These results may be important for patients who cannot tolerate the therapeutic dosage of Herceptin currently used because of heart problems. [Copyright &y& Elsevier]

Abstract: Aim: Trastuzumab is a monoclonal antibody that is used in treating breast cancer. We labeled this monoclonal antibody with lutetium-177 and performed in vitro quality control tests as a first step in the production of a new radiopharmaceutical. Material and Methods: Trastuzumab was labeled with lutetium-177 using DOTA as chelator. Radiochemical purity and stability in buffer and human blood serum were determined using thin layer chromatography. Immunoreactivity and toxicity of the complex were tested on MCF7 breast cancer cell line. Results: The radiochemical purity of the complex was 96±0.9%. The stabilities in phosphate buffer and in human blood serum at 96 h postpreparation were 93±1.2% and 85±3.5%, respectively. The immunoreactivity of the complex was 89±1.4%. At a concentration of 1 nM, the complex killed 70±3% of MCF7 cells. At 1.9 nM, 90±5% of the cells were killed. Conclusions: The results showed that the new complex could be considered for further evaluation in animals and possibly in humans as a new radiopharmaceutical for use in radioimmunotherapy against breast cancer. [Copyright &y& Elsevier]

Abstract: Human epithelial mucin, MUC1, is commonly overexpressed in adenocarcinoma that includes more than 80% of breast cancers. The PR81 is a murine anti-MUC1 monoclonal antibody (MAb) that was prepared against the human breast cancer. We developed an indirect method for labeling of this antibody with 99mTc in order to use the new preparation in immunoscintigraphy studies of BALB/c mice bearing breast tumors. The 99mTc–PR81 complex was prepared using the HYNIC as a chelator and tricine as a coligand. The labeling efficiency determined by instant thin-layer chromatography (ITLC) was 89.2%±4.7%, and radiocolloides measured by cellulose nitrate electrophoresis were 3.4%±0.9%. The in vitro stability of labeled product was determined at room temperature by ITLC and in human serum by gel filtration chromatography — 88.3%±4.6% and 79.8%±5.7% over 24 h, respectively. The integrity of labeled MAb was checked by means of sodium dodecyl sulfate polyacrylamide gel electrophoresis, and no significant fragmentation was seen. The results of cell binding studies showed that both labeled and unlabeled PR81 were able to compete for binding to MCF 7 cells. Biodistribution studies performed in female BALB/c mice with breast tumor xenografts at 4, 16 and 24 h after the 99mTc-HYNIC-PR81 injection demonstrated a specific localization of the compound at the site of tumors and minimum accumulation in non target organs. The tumor imaging was performed in BALB/c mice with breast xenograft tumors at 4, 8, 12, 16, 20, 24, 28, 32 and 36 h after the complex injection. The tumors were visualized with high sensitivity after 8 h. The findings showed that the new radiopharmaceutical is a promising candidate for radioimmunoscintigraphy of the human breast cancer. [Copyright &y& Elsevier]

Abstract: In this study, we labeled trastuzumab with 177Lu to synthesize a new radiopharmaceutical for therapy of breast cancer and at the first stage investigated its therapeutic effects on SKBr3 and MCF7 breast cancer cell lines. Trastuzumab-177Lu showed very good in-vitro characteristics such as high radiochemical purity (91±0.9%), good stability in PBS buffer (86±2.3%) and blood serum (81±2.7%) up to 96h, appropriate immunoreactivity (85.4±1.1%) and high cytotoxicity in HER2 expression cells. 5 fold increase in toxicity of trastuzumab-177Lu was observed when compared with unlabeled trastuzumab on SKBr3 cells. [ABSTRACT FROM AUTHOR]