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Context: Targeting MUC1 antigens which are overexpressed in 80% of breast cancers can be widely used in the field of radioimmunoscintigraphy (RIS) of breast cancer.Aims: The aim of this study was to develop a new diagnostic labeled compound for breast cancer RIS.Settings and Design: In this study, an efficient indirect labeling method of PR81 with Indium-111 was developed and preliminary preclinical qualifications were reported.Subjects and Methods: 111In-DTPA-PR81 was prepared and its radiochemical purity and stabilities in human serum and in phosphate-buffered saline (PBS) buffer were surveyed. Furthermore, cellular studies including complex reactivity, binding specificity, cell toxicity, etc., were examined. Finally, biodistribution and scintigraphy of the complex were studied in normal and tumoral animals.Statistical Analysis Used: Statistical analyses were performed using SPSS 10.0.Results: 111In-DTPA-PR81 was prepared with a radiochemical purity of >99% at optimized conditions. Stability studies showed the radiochemical purity of >90% in PBS buffer after 96 h, while the stability in human serum showed decrement to 81% after 96 h. Reactivity of the complex with MUC1 was significantly (P < 0.005) higher than bovine serum albumin (BSA) (about 7-8 times), even though BSA concentration was about twice the MUC1. The binding specificity of the complex to the MUC1 antigen was confirmed by means of immunoreactivity assay. Cell toxicity examination showed no significant lethal effect of the radiolabeled compound on the cells. Biodistribution studies of the complex in normal rats were consistent with the biodistribution of antibodies and high accumulation was observed in the tissues expressing MUC1 antigen. The results of 111In-DTPA-PR81 scintigraphy in tumoral female BALB/c mice at 24 and 48 h after injection showed an increasement of the accumulation in the tumor site.Conclusions: 111In-DTPA-PR81 can be considered as a potential agent for imaging of the MUC1 +breast tumors. [ABSTRACT FROM AUTHOR]

In this study, [111In]In‐DOTA‐PR81 was developed, and its preliminary preclinical qualifications were assessed for single photon emission computed tomography (SPECT) imaging of breast cancer. DOTA‐NHS‐ester was practiced and successively purified by molecular filtration. The chelate:mAb ratio was determined by spectrophotometry. DOTA‐PR81 was radiolabeled with In‐111 and its radiochemical yield, in vitro stability, in vitro internalization, and immunoreactivity tests were performed. SPECT imaging and tissue counting were applied to evaluate the tissue distribution of [111In]In‐DOTA‐hIgG and [111In]In‐DOTA‐PR81 in BALB/c mice bearing breast tumors. The radiochemical yield of [111In]In‐DOTA‐PR81 complex was >95.0 ± 0.5% (ITLC), and the specific activity was 170 ± 44 MBq/mg. Conjugation reaction resulted in the average number of chelators attached to a mAb (c/a) of 3.4 ± 0.3:1. The radioimmunoconjugate showed immunoreactivity towards MCF7 cell line and MUC1 antigen while its significant in vitro and in vivo stability were investigated over 48 h, respectively (93.0 ± 1.2% in phosphate‐buffered saline (PBS) and 84.0 ± 1.3% in human serum). The peak concentration of internalized activity of [111In]In‐DOTA‐PR81 was between 4 to 6 h. In comparison with control probes, the complex was accumulated with high specificity and sensitivity at the tumor site. Achieved results indicated that [111In]In‐DOTA‐PR81 could be contemplated as an appropriate radiotracer for prognostic imaging of antigens in oncology. [ABSTRACT FROM AUTHOR]

With PR81 as a murine monoclonal antibody (mAb) that was prepared against the human breast cancer, the MUC1 receptor specific targeting is possible. In this study, PR81-conjugated bovine serum albumin (BSA) nanoparticles loaded with anticancer drug 5-fluorouracil (5-FU) were developed. Enzyme linked immunosorbant assay (ELISA) results showed high immunoreactivity of PR81 mAb conjugated to nanoparticles towards MUC1 related peptide or native cancerous MUC1 and almost no cross-reaction to non-specific proteins. In vitro experiments were performed to determine the ability of this new drug delivery system on overcoming MCF-7 breast cancer cells in comparison with four other systems. The results revealed that these cell-type specific drug loaded nanoparticles could achieve more cell death as compared to when the 5-FU was used with no carriers. Stability studies of produced drug delivery system proved high immunoreactivity of conjugated PR81 even after 11 days of storage in room temperature. [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]

A simple, sensitive, rapid and specific enzyme linked immunosorbent assay (ELISA) for quantitative measurement of aflatoxin B1 (AFB1) in urine samples was developed in this study. Polyclonal antibodies were raised against a C1-carboxymethyl oxime (CMO) derivative of AFB1 conjugated bovine serum albumin (BSA). AFB1-C8-penicillinase (AFB1-C8-P) and AFB1-C1-carboxymethyl oxime-penicillinase (AFB1-CMO-P) were prepared and used as tracer molecule. A heterologous combination of antibody and enzyme conjugates (AFB1-C1-CMO-BSA and AFB1-C8-P) proved to work better with respect to specificity and sensitivity. Ig purified antibody (4 üg/well) was coated onto the pre-coated (BSA) wells of microtiter plate. The assay procedure was completed within 3 hr and the sensitivity was calculated to be from 200 pg/ml. The standard curve was linear up to 10 ng/ml so was able to detect high concentration of AFB1 in sample. Affinities were calculated for homologous and heterologous system in which the heterologous system showed better affinities (1.9 × 108 M-1). The antibody prepared in this study showed minimal cross-reaction with structurally related molecules being affected by homology and heterology of the assay system that is the site of conjugation of carrier protein for antibody production using the hapten BSA conjugate and the site of enzyme conjugated on the hapten molecule used as tracer as well as direct and indirect coating of antibody on the surface of microtiter plat. The results reported here indicated that the heterologous combination of antibody and enzyme conjugate performs better in assay qualities in general. More than 90% recovery of AFB1 added to stripped urine samples were observed in this type of assay. Inter and intra-assay percent of coefficient of variations for ten successive assays were found to be 10.2 and 6.9% respectively. Logit -log transformation of standard curve and sample dilution with urine sample containing no AFB1 in a serial manner exhibited parallel line with the slope of -1.03 and -1.03 respectively. A correlation of 0.90 was found between the ELISA reported in this study and radioimmunoassay (RIA) of AFB1 in urine samples. [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]

Context: Targeting MUC1 antigens which are overexpressed in 80% of breast cancers can be widely used in the field of radioimmunoscintigraphy (RIS) of breast cancer., Aims: The aim of this study was to develop a new diagnostic labeled compound for breast cancer RIS., Settings and Design: In this study, an efficient indirect labeling method of PR81 with Indium-111 was developed and preliminary preclinical qualifications were reported., Subjects and Methods: 111 In-DTPA-PR81 was prepared and its radiochemical purity and stabilities in human serum and in phosphate-buffered saline (PBS) buffer were surveyed. Furthermore, cellular studies including complex reactivity, binding specificity, cell toxicity, etc., were examined. Finally, biodistribution and scintigraphy of the complex were studied in normal and tumoral animals., Statistical Analysis Used: Statistical analyses were performed using SPSS 10.0., Results: 111 In-DTPA-PR81 was prepared with a radiochemical purity of >99% at optimized conditions. Stability studies showed the radiochemical purity of >90% in PBS buffer after 96 h, while the stability in human serum showed decrement to 81% after 96 h. Reactivity of the complex with MUC1 was significantly (P < 0.005) higher than bovine serum albumin (BSA) (about 7-8 times), even though BSA concentration was about twice the MUC1. The binding specificity of the complex to the MUC1 antigen was confirmed by means of immunoreactivity assay. Cell toxicity examination showed no significant lethal effect of the radiolabeled compound on the cells. Biodistribution studies of the complex in normal rats were consistent with the biodistribution of antibodies and high accumulation was observed in the tissues expressing MUC1 antigen. The results of 111 In-DTPA-PR81 scintigraphy in tumoral female BALB/c mice at 24 and 48 h after injection showed an increasement of the accumulation in the tumor site., Conclusions: 111 In-DTPA-PR81 can be considered as a potential agent for imaging of the MUC1 +breast tumors., Competing Interests: None