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Biomedical subjects

C F Meares

Publications and source records attributed to C F Meares.

At least 55 records · Page 3Linked to original sources
PubMed1996

Binding of the sigma 70 protein to the core subunits of Escherichia coli RNA polymerase, studied by iron-EDTA protein footprinting.

We have used a nonspecific protein cleaving reagent to map the interactions between subunits of the multisubunit enzyme RNA polymerase (Escherichia coli). We developed suitable conditions for using an untethered Fe-EDTA reagent, which does not bind significantly to proteins. Comparison of the cleaved fragments of the subunits from the core enzyme (alpha 2 beta beta') and the holoenzyme (core+sigma 70) shows that absence of the sigma 70 subunit is associated with the appearance of several cleavage sites on the subunits beta (within 10 residues of sequence positions 745, 764, 795, and 812) and beta' (within 10 residues of sequence positions 581, 613, and 728). A cleavage site near beta residue 604 is present in the holoenzyme but absent in the core, demonstrating that a conformational change occurs when sigma 70 binds. No differences are observed for the alpha subunit.

Amino Acid Sequence↗
PubMed1996

Lymphoma-selective antibody Lym-1 recognizes a discontinuous epitope on the light chain of HLA-DR10.

The selectivity of Lym-1 for malignant B lymphocytes makes this monoclonal antibody a promising candidate for the delivery of toxic agents to malignant B cells. The original immunogen used for the development of Lym-1 was Raji Burkitt's lymphoma cell nuclei [Epstein A. L., Marder R. J., Winter J. N., Stathopoulos E., Chen F. M., Parker J. W., Taylor C. R. (1987) Cancer Res 47: 830]. The Lym-1 antigen was characterized at that time as a polymorphic HLA-DR variant. We prepared an affinity column using immobilized Lym-1 to isolate the Lym-1 antigen from Raji cell lysate. Immunological characterization of the immunoaffinity-purified Lym-1 antigen on Western blots led to the conclusion that the antigen is the beta chain of HLA-DR10. This was confirmed by Edman sequencing of the isolated polypeptide chain. Western blots further show that the Lym-1 epitope is only recognized if the beta chain disulfide bonds are intact. These results imply that Lym-1 binds a discontinuous epitope on the beta chain of HLA-DR10.

Amino Acid Sequence↗
PubMed1996

Antibody responses to macrocycles in lymphoma.

UNLABELLED: Metallic radioimmunoconjugates have promise for radioimmunoimaging and therapy. Macrocyclic chelating agents allow formation of stable metallic radioimmunoconjugates but have been reported to be immunogenic. This study assesses human antibody responses in patients that were imaged or treated with radiolabeled Lym-1 containing the macrocyclic chelators 1,4,8,11-tetraazacyclotetradecane-N,N',N",N"'-tetraacetic acid (TETA) or 1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid (DOTA). METHODS: One to six doses (median 1) and 6 to 285 mg (median 33) 67Cu-2IT-BAT (2-iminothiolane bromoacetamidobenzyl TETA)- or 111In-2IT-BAD (2-iminothiolane bromoacetamidobenzyl DOTA)-Lym-1 were administered to each of 18 patients with lymphocytic malignancies. Solid-phase ELISA, utilizing unchelated Lym-1 or human serum albumin conjugated to DOTA, TETA or 2IT-bromoacetamidobenzyl-ethylenediaminetetraacetic acid (BABE) as coating antigens, was used to characterize antibody responses against 67Cu-2IT-BAT-Lym-1 and 111In-2IT-BAD-Lym-1 by quantitating antibodies against the Lym-1, DOTA, TETA or 2IT moieties, respectively. RESULTS: None of the patients had evidence for serum sickness. No patient that received 111In-2IT-BAD-Lym-1 developed antibodies to Lym-1 or DOTA. Two (15%) of the 13 patients that received 67Cu-2IT-BAT-Lym-1 developed antibodies against both TETA and Lym-1, and an additional patient developed antibodies against Lym-1 only. No patient developed an antibody response solely against the macrocycle, nor did any of the patients generate antibodies against the 2IT molecule. HAMA levels were many times greater in amount than HATA levels even when their relative molecular masses were considered. CONCLUSION: Although macrocycles such as DOTA and TETA, and other chelates, can be haptens and thus potentially immunogenic, our findings do not support the view that macrocycles are more immunogenic than other radiometal chelating agents.

Animals↗
PubMed1995

Prelabeling of chimeric monoclonal antibody L6 with 90yttrium- and 111indium-1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid (DOTA) chelates for radioimmunodiagnosis and therapy.

90Y and 111In have been attached to chimeric monoclonal antibody L6 with a bifunctional chelating agent (DOTA-peptide isothiocyanate). The bifunctional chelating agent was prelabeled with either radiometal and then conjugated to the antibody. Studies in human patients showed excellent 111In single-photon emission computed tomography images of breast cancer lesions 24 h after injection.

Animals↗
PubMed1995

Proximity mapping the surface of a membrane protein using an artificial protease: demonstration that the quinone-binding domain of subunit I is near the N-terminal region of subunit II of cytochrome bd.

A novel experiment has been used to show proximity relationships between sites on the surface of the cytochrome bd quinol oxidase of Escherichia coli. The artificial protease iron (S)-1-[p-(bromoacetamido)benzyl]-EDTA (Fe--BABE) was conjugated to selected reactive cysteines placed in subunit I or subunit II, with the aim of identifying amino acid residues within approximately 12 A of each site of attachment. The protease was activated with H2O2 and ascorbate for a few seconds, and hydrolysis products were isolated and analyzed by N-terminal sequencing. Among other results, we found that residue 39 of subunit II is near residue 255 of subunit I in the putative quinone-binding domain (Q loop) of the oxidase. Since this technique is insensitive to the nature of the amino acid side chains, it should prove generally valuable in revealing spatial relationships both within and between subunits in complex proteins where high-resolution structural information is not available.

Amino Acid Sequence↗
PubMed1995

Effect of the extent of chelate substitution on the immunoreactivity and biodistribution of 2IT-BAT-Lym-1 immunoconjugates.

Trial therapy for lymphoma with the radiolabeled chelate-antibody conjugate 67Cu-2IT-BAT-Lym-1 has been promising. It is desirable to deliver therapeutic doses of radiometal using a minimum amount of 2IT-BAT-Lym-1 to minimize the risks of adverse patient reaction and antigenic response to antibody. This is readily accomplished by increasing the number of metal-binding sites (i.e., chelating agents) conjugated per antibody, but the ability of the antibody to bind antigen and target tumor cells in vivo must not be impaired by the conjugation reaction. To determine the maximum chelator:antibody ratio (c/a) of 2IT-BAT-Lym-1 at which functional integrity is preserved, immunoconjugates with a c/a of 1.3-23 were prepared and examined by radioimmunoassay and competitive antigen binding versus lightly iodinated Lym-1. The biodistribution in tumored mice of conjugates with c/a of 2.1, 4.3, 8.4, and 11.4 also was examined. Conjugates with c/a up to 5 exhibited no loss of immunoreactivity, and conjugates with c/a up to 11 retained 75% or greater immunoreactivity relative to unmodified Lym-1. All conjugates examined competed less effectively than did unmodified Lym-1 for antigen binding, but the effect at c/a 5 was slight. Tumor uptake declined with increasing c/a, but the effect was insignificant at c/a 2.1 and 4.3. Conjugates of c/a 4-5 were found to be optimal for the preparation of radioimmunoconjugate of high specific activity with minimal, if any, loss of functional integrity.

Animals↗
PubMed1995

Directed hydroxyl radical probing of 16S rRNA using Fe(II) tethered to ribosomal protein S4.

Localized hydroxyl radical probing has been used to explore the rRNA neighborhood around a unique position in the structure of the Escherichia coli 30S ribosomal subunit. Fe(II) was attached to ribosomal protein S4 at Cys-31 via the reagent 1-(p-bromoacetamidobenzyl)-EDTA. [Fe-Cys31]S4 was then complexed with 16S rRNA or incorporated into active 30S ribosomal subunits by in vitro reconstitution with 16S rRNA and a mixture of the remaining 30S subunit proteins. Hydroxyl radicals generated from the tethered Fe resulted in cleavage of the 16S rRNA chain in two localized regions of its 5' domain. One region spans positions 419-432 and is close to the multihelix junction previously placed at the RNA binding site of S4 by chemical and enzymatic protection (footprinting) and crosslinking studies. A second site of directed cleavage includes nucleotides 297-303, which overlap a site that is protected from chemical modification by protein S16, a near neighbor of S4 in the ribosome. These results provide useful information about the three-dimensional organization of 16S rRNA and indicate that these two regions of its 5' domain are in close spatial proximity to Cys-31 of protein S4.

Escherichia coli↗
PubMed1995

RadioimmunoPET: detection of colorectal carcinoma with positron-emitting copper-64-labeled monoclonal antibody.

UNLABELLED: Detection of tumor foci may be improved by combining the selective tumor-targeting properties of a monoclonal antibody with the superior sensitivity and contrast resolution of PET. METHODS: An anti-colorectal carcinoma monoclonal antibody (MAb 1A3) was labeled with 64Cu, a positron-emitting radionuclide, by use of a bifunctional chelate (bromoacetamidobenzyl-TETA) and evaluated in 36 patients with suspected advanced primary or metastatic colorectal cancer. After radiopharmaceutical injection (5-20 mg protein, 10 mCi 64Cu), PET was performed once or twice, 4 to 36 hr later. All patients had CT scans and 18 patients were also studied with [18F]fluorodeoxyglucose (FDG) PET. RESULTS: In 29 patients, one or more tumor sites (n = 56) were proven, in 5 patients the absence of active tumor was confirmed and in the remaining 2, tumor status is not yet confirmed. Of the 56 confirmed tumor sites, 40 were detected by MAb-PET as foci of increased activity (sensitivity 71%). The positive predictive value of MAb-PET was excellent, ranging from 89% (40/45) to 96% (43/45), depending on the ultimate classification of three image-positive, but as yet unconfirmed tumor sites. Also, MAb-PET detected 11 new occult tumor sites, including 9 small abdominopelvic foci less than 2.0 cm in diameter that were not detected by CT or MRI. There were no complications, but significantly elevated HAMA titers were found in 28% of the 29 patients tested 1 to 12 mo after injection. There was no apparent dose-related effect from 5 to 20 mg MAb 1A3. CONCLUSION: These Phase I/II results suggest that PET with radiolabeled MAbs (radioimmunoPET) may have important applications in clinical oncology, particularly for detecting smaller colorectal tumor foci in the abdomen or pelvis and for determining accurate dosimetry.

Adenocarcinoma↗
PubMed1995

Pharmacokinetics of chimeric L6 conjugated to indium-111- and yttrium-90-DOTA-peptide in tumor-bearing mice.

UNLABELLED: A bifunctional chelating agent, DOTA-Gly3-L-(p-isothiocyanato)-phenylalanine amide (DOTA-peptide-NCS), was studied in nude mice bearing human breast cancer xenographs (HBT 3477) to determine its potential for radioimmunoconjugate therapy. METHODS: Indium-111 and yttrium-90 were attached to an anti-adenocarcinoma chimeric L6 (ChL6) monoclonal antibody (MAb) after pre-chelation to the DOTA-peptide-NCS and the desired neutral radiochelates were obtained by purification. The unique characteristic of the DOTA-peptide-NCS to form neutral complexes with trivalent metals was utilized to separate the resulting 111In and 90Y radiochelates from excess chelating agent and other anionic by-products resulting from metal impurities. The purified radiochelates were then conjugated to ChL6. The pharmacokinetics of 111In- and 90Y-DOTA-peptide-ChL6 were obtained for 5 days after injection in nude mice bearing HBT 3477 xenographs. The results were compared with the pharmacokinetics of 125I-ChL6 obtained in the same mouse model. RESULTS: The whole-body clearance of 125I-ChL6, 90Y- and 111In-DOTA-peptide-ChL6 was monoexponential with biologic half-times of 92, 104 and 160 hr, respectively. Blood clearances of the three radiopharmaceuticals were biphasic. The radiometal immunoconjugates had greater tumor uptake and slower clearances. CONCLUSION: Indium-111- and 90Y-DOTA-peptide-ChL6 can be produced at high specific activity with fewer than one chelate per MAb by using a pre-labeling method that permits radiochelate purification by charge selection. Studies in mouse xenografts indicate that tumor uptake is enhanced and a favorable therapeutic index is achieved using these agents.

Animals↗
PubMed1995

Preparation, biodistribution and dosimetry of copper-64-labeled anti-colorectal carcinoma monoclonal antibody fragments 1A3-F(ab')2.

UNLABELLED: Antibody fragments labeled with a radiometal using bifunctional chelates generally undergo renal clearance followed by trapping of the metabolites, leading to high radiation doses to the kidneys. Copper-64-labeled BAT-2IT-1A3-F(ab')2 was recently reported to accumulate in colorectal tumors in an animal model, however, kidney uptake was also high. In this study, the preparation of 64Cu-BAT-2IT-1A3-F(ab')2 was optimized to reduce the renal uptake. METHODS: The bifunctional chelate 6-bromoacetamidobenzyl-1,4,8,11-tetraazacyclotetradecane-N,N ',N",N'"-tetraacetic acid (BAT) was conjugated to 1A3-F(ab')2 using the linking agent 2-iminothiolane (2IT). The conjugation reaction produced 20% of a lower molecular weight (molecular wieght) impurity found to be TETA-1A3-Fab'. The conjugation procedure was optimized to include FPLC purification of the BAT-2IT-1A3-F(ab')2 from TETA-1A3-Fab' after conjugation prior to labeling with 64Cu. The biodistribution of 64Cu-labeled FPLC-purified and unpurified conjugates was determined in normal Sprague-Dawley rats and tumor bearing Golden Syrian hamsters. Human absorbed doses were calculated from rat biodistribution data and PET imaging of a baboon. RESULTS: Upon FPLC purification of the BAT-2IT-1A3-F(ab')2, the immunoreactivity of 64Cu-labeled 1A3-F(ab')2 was significantly improved over that of non-FPLC-purified 64Cu-BAT-2IT-1A3-F(ab')2, and the kidney uptake was decreased in normal rats. The biodistribution in hamsters showed some improvement in both tumor uptake and kidney clearance with FPLC-purified 64Cu-BAT-2IT-1A3-F(ab')2. CONCLUSION: The improved dosimetry of 64Cu-labeled FPLC purified BAT-2IT-1A3-F(ab')2 should more readily allow this agent to be investigated clinically to image colorectal cancer using PET.

Animals↗
PubMed1994

Pharmacokinetics of pretargeted monoclonal antibody 2D12.5 and 88Y-Janus-2-(p-nitrobenzyl)-1,4,7,10-tetraazacyclododecanetetraacetic acid (DOTA) in BALB/c mice with KHJJ mouse adenocarcinoma: a model for 90Y radioimmunotherapy.

Three-step pretargeting for radioimmunotherapy in BALB/c mice with KHJJ tumors was done with monoclonal antibody (mAb) 2D12.5, which is specific for yttrium-1,4,7,10-tetraazacyclododecanetetraacetic acid (DOTA) but nonspecific for the tumor. Tumor uptake was by passive diffusion of mAb through leaky neovasculature in the tumor. The three steps were: (a) anti-hapten mAb 2D12.5 (0 h); (b) polyvalent haptenprotein conjugate chase (20 h); and (c) 88Y-labeled monovalent DOTA or bivalent Janus-DOTA haptens (21 h) and organ and tumor bioassay (24 h). Rapid tumor (T) uptake and high tumor:blood ratio (T:BL) was seen 3 h after injection after step c. For monovalent 88Y-DOTA, T = 1.7%/g* and T:BL = 16:1; for bivalent 88Y-Janus-DOTA, T = 4.41%/g* and T:BL = 21:1 at 3 h (*, P < 0.001). Blood and bone plus marrow were << 1%/g, and liver was < 1%/g. The 24-h whole body retention was approximately 5% of injected dose with 1% in tumor (20% of total), 1.8% in other organs, and 2.2% in carcass; the 24-h whole body retention of covalent nonspecific antibody conjugates was > 80% of injected dose. The biological half-life in the tumor of 0.9 microCi 88Y-Janus-DOTA was approximately 24 h, measured daily for 5 days. Activity in microCi/g of tumor and blood for 90Y equimolar to the amount of 88Y injected (0.9 microCi 88Y = 0.744 pmol = 36.47 microCi 90Y) was used for calculating the area under the curve of tumor and blood in microCi-h/g of 90Y. The 90Y radiation absorbed dose (RAD) from multiplying microCi-h/g x the 90Y absorbed dose constant, 1.99 RAD-g/microCi-h, gave T = 89 RAD and BL = 3.7 RAD. The therapeutic ratio from RAD T:RAD BL = 24:1. These results indicate that pretargeting 90Y hapten-specific mAb for radioimmunotherapy has considerable promise.

Acetates↗
PubMed1994

Immunogenicity in rabbits and mice of an antibody-chelate conjugate: comparison of (S) and (R) macrocyclic enantiomers and an acyclic chelating agent.

The macrocyclic bifunctional chelating agent 2-(p-bromoacetamidobenzyl)-1,4,7,10-tetraazacyclododecanetetraa cetic acid (BAD), forms inert metal complexes ideal for radioimmunotherapy. Kosmas et al. (Cancer Res., 52: 904-911, 1992) found 2-imminothiolane linker-(S)-BAD monoclonal antibody HMFG1 highly immunogenic in patients. We studied the immunogenicity of (S) and (R) enantiomers of 2-imminothiolane linker-BAD rabbit IgG, monoclonal antibody Lym-1, and Lym-1 2-imminothiolane linker-(S)-bromoacetamidobenzyl-EDTA in 15 rabbits. Five groups of three each were given 0.1, 1.0, or 10 mg of 111In conjugate i.v., blood samples were taken daily for 14 days and biweekly for 70 days, and the plasma T1/2 was calculated. A drop in plasma 111In at 6-8 days coincided with the appearance of antibody on enzyme-linked immunosorbent assay. Specific anti-(S)-BAD, anti-(R)-BAD, anti-(S)-bromoacetamidobenzyl-EDTA, and anti-mouse IgG were measured. Rabbit IgG conjugates did not elicit an immune response. Mouse IgG conjugates were immunogenic on the first exposure, with both anti-1,4,7,10-tetraazacylododecane N,N',N'',N'''-tetraacetic acid and anti-mouse responses. Anti-1,4,7,10-tetraazacylododecane N,N',N'',N'''-tetraacetic acid was specific for the (S) or (R) enantiomer, but cross-reaction appeared with reboosting. A second injection of the opposite enantiomer gave a response to that enantiomer. Lym-1 bromoacetamidobenzyl-EDTA produced anti-bromoacetamidobenzyl-EDTA and anti-mouse response.

Animals↗
PubMed1994

Comparative toxicity studies of yttrium-90 MX-DTPA and 2-IT-BAD conjugated monoclonal antibody (BrE-3).

BACKGROUND: BrE-3 is monoclonal antibody that has promise for imaging and therapy of human adenocarcinoma. Because of observations in therapeutic trials of yttrium-90 (90Y) escape from radioimmunoconjugates and uptake by the skeleton with resultant bone marrow toxicity, the authors attempted to evaluate the importance of this factor by a comparison of the LD50 in healthy mice treated with 90Y that had been chelated with either of two high affinity chelators, methylbenzyldiethylene-triaminepentaacetic acid (MX-DTPA) or bromoacetamidobenzyl-1,4,7,10-tetraazocyclododecane- N,N',N'',N'''-tetraacetic acid (BAD). METHODS AND RESULTS: Bone marrow hematopoietic toxicity was dose-limiting and the source of death for both chelators. The LD50 for 90Y-BrE-3-MX-DTPA was 220.9 microCi, and that for 90Y-BrE-3-2IT-BAD and was 307.8 microCi. Whole-body autoradiography revealed substantially greater uptake of 90Y in the skeleton when MX-DTPA was used as the chelator. CONCLUSIONS: These observations suggest that 90Y escape to bone is a significant factor in the maximum tolerated dose of radioimmunoconjugate that can be used in therapeutic trials. These results probably underestimate the importance of 90Y escape since 90Y in the skeleton of patients is likely to be more significant than in mice because more of the 90Y energy is absorbed in the marrow of larger species.

Animals↗
PubMed1994

Pharmacokinetics and toxicity of an yttrium-90-CITC-DTPA-HMFG1 radioimmunoconjugate for intraperitoneal radioimmunotherapy of ovarian cancer.

BACKGROUND: The intracavitary route for the administration of monoclonal antibodies is used in a variety of locally spreading cancers. The authors have been treating patients with ovarian cancer in Phase I and II studies assessing toxicity and response to improved radioimmunoconjugates. METHODS: Nineteen patients, 34-65 years of age, were treated with a new radioimmunoconjugate, 90Y-CITC-DTPA-HMFG1, instilled in the peritoneal cavity after second-look laparoscopy. Activity was increased in a stepwise fashion. RESULTS: Following the intraperitoneal administration of 90Y-CITC-DTPA-HMFG1, levels of the radioimmunoconjugate in the blood increased, reaching a peak of about 30% of injected activity at around 54 hours posttreatment. Approximately 18% of the radiolabel was excreted in the urine within 96 hours. Bone-marrow toxicity was the dose-limiting factor. Grade III platelet and granulocyte toxicity was observed at 19.3 mCi/m2. A type III immunologic response was observed in a number of patients. CONCLUSIONS: A dose of 18.5 mCi/m2 for subsequent treatments is recommended, based on a linear correlation of activity dose-to-body surface area. The clinical profile of a mild to moderate hypersensitivity syndrome is presented and hypotheses regarding its etiology are suggested.

Adult↗
PubMed1994

A comparative study of copper-67 radiolabeling and kinetic stabilities of antibody-macrocycle chelate conjugates.

BACKGROUND: The development of new chelating agents and radiolabeling protocols is essential to progress in radioimmunotherapy with antibody-chelate conjugates. METHODS: Immunoconjugates of four polyazamacrocycles with N-bonded acetate groups were prepared by conjugation via 2-iminothiolane to Lym-1, a murine antilymphoma immunoglobulin G2a MoAb. To optimize 67Cu radiolabeling, complexation conditions were explored. The kinetic stabilities in vitro in human serum of four 67Cu labeled immunoconjugates were investigated. RESULTS: Lym-1-2IT-6-BAT-67Cu, the chelate conjugate of 6-[p-(bromoacetamido)benzyl]-1,4,8,11-tetraazacyclotetradecane- N,N',N''N'''-tetraacetic acid, exhibited excellent kinetic stability in human serum, while Lym-1-2IT-2-BAT-67Cu, prepared from the structural isomer 2-[p-(bromoacetamido)benzyl]-1,4,8,11- tetraazacyclotetradecane-N,N',N'',N'''-tetraacetic acid, exhibited a markedly higher rate of loss of radiometal. It was observed that the radiolabeling ratio of Lym-1-2IT-6-BAT-67Cu, in mCi per mg immunoconjugate, was limited solely by the specific activity of the radiometal, which varied significantly from lot to lot. This ratio for a given lot of 67Cu can be predicted by a preliminary titration. CONCLUSIONS: The preparation of 67Cu labeled immunoconjugates of therapeutic quality has been improved by the determination of optimum radiolabeling conditions, and by development of a titration protocol which rapidly and accurately predicts the radiolabeling ratio in mCi per mg immunoconjugate. The surprising difference in the properties of 6-BAT and 2-BAT shows the exquisite dependence of kinetic stability on structure.

Antibodies, Monoclonal↗
PubMed1992

Development of humoral immune responses against a macrocyclic chelating agent (DOTA) in cancer patients receiving radioimmunoconjugates for imaging and therapy.

The development of stable immunoconjugates by the advent of macrocyclic metal chelating agents (DOTA) has enabled us to study the ability of 111In-DOTA-labeled monoclonal antibodies to detect tumor lesions in a pilot radioimmunolocalization study, as well as to evaluate the kinetics, toxicity, and efficacy of i.p. administered 90Y-DOTA-labeled murine monoclonal antibody in a Phase I/II clinical trial of advanced ovarian cancer. The development of serum sickness-like reactions in three of six treated patients, in the absence of previous monoclonal antibody administration, led us to study the potential immunogenicity of the new chelate. Six patients with ovarian cancer received 25 mg of HMFG1 monoclonal antibody coupled with 90Y-DOTA (doses of radioactivity, 15 to 25 mCi), administered i.p. Eight patients with various malignant tumors received low doses (220 micrograms to 1 mg) of monoclonal antibodies, labeled with 111In-DOTA, i.v. for imaging studies. Using a solid-phase enzyme-linked immunosorbent assay method, the immunogenicity of DOTA was evaluated. Serial dilutions of patients' sera, before and after imaging or therapy with DOTA-coupled monoclonal antibodies, as well as sera from patients who did not receive DOTA-coupled antibody, were screened on enzyme-linked immunosorbent assay plates coated with human serum albumin (HSA), HSA-2-iminothiolane, and HSA-2-iminothiolane-benzyl-DOTA. All patients treated with i.p. monoclonal antibody developed anti-DOTA antibodies. Four of eight patients who received i.v. "imaging" doses of DOTA-coupled monoclonal antibody developed antibodies against DOTA. The levels of anti-DOTA response correlated with the amount of injected radioimmunoconjugate (r = 0.889, P less than 0.001). None of the patients who received DOTA-coupled antibody had detectable antibodies against the macrocycle before immunoconjugate administration. We then addressed further the restriction of the immune response against the macrocycle. We found that there was no or very low response against the aromatic ring attached to DOTA. Most, if not all, of the immune response is directed against the DOTA ring structure. Affinity purification of anti-DOTA antibody from serum enabled quantitation of these antibodies in the serum of patients. An inverse, statistically significant correlation was observed between the percentage of binding inhibition of a patient's serum to DOTA, by HSA-2-iminothiolane-DOTA (100 micrograms/ml) and the level of anti-DOTA immunoglobulin in the serum.(ABSTRACT TRUNCATED AT 400 WORDS)

Aged↗
PubMed1992

Pretargeted immunoscintigraphy: effect of hapten valency on murine tumor uptake.

A method of radioimmunoscintigraphy using bivalent "Janus" haptens with an apparent enhanced affinity ("avidity") for the antibody is described. Janus with 50 micrograms pretargeted Mab WC3A11 resulted in significantly higher murine tumor concentrations (approximately 7%/g) compared to monovalent haptens (approximately 1.4%/g, p < 0.001), and the same high tumor-to-background ratios (approximately 3/1). Janus was synthesized by coupling two molecules of BABE together with a 1,4 butanedithiol linker. Janus itself was rapidly excreted (T1/2b = 42 min) by the kidneys and did not concentrate in any other organs or tissues. Three-step pretargeted immunoscintigraphy (binder, chaser, tracer) with 111In- or 67Ga-Co(III) Janus produced excellent mouse tumor images in 3 hr with high tumor-to-background ratios. The use of short-lived tracers, such as 99mTc and 68Ga, with a T1/2p of hours to image antibodies that localize slowly over several days in vivo is accessible with this new technology.

Animals↗