Biomedical subjects

G A van Dongen

Publications and source records attributed to G A van Dongen.

At least 19 recordsLinked to original sources

PubMed2001

Targeting of aluminum (III) phthalocyanine tetrasulfonate by use of internalizing monoclonal antibodies: improved efficacy in photodynamic therapy.

The use of monoclonal antibodies (MAbs) directed against tumor-associated antigens for targeting of photosensitizers is an interesting option to improve the selectivity of photodynamic therapy (PDT). Hydrophilic photosensitizers are most suitable for conjugation to MAbs because of their water solubility. The photosensitizer aluminum (III) phthalocyanine tetrasulfonate [AlPc(SO3H)4] has many ideal photochemical properties; however, because of its hydrophilicity, the free form of this sensitizer does not readily reach the critical intracellular target and, therefore, is ineffective in PDT. On the basis of our previous studies, we hypothesized that AlPc(SO3H)4 might be suitable for PDT when coupled to internalizing tumor-selective MAbs. In this study, a reproducible procedure is presented for coupling of AlPc(SO3H)4 to MAbs via the tetra-glycine derivative AlPc(SO2Ngly)4. Conjugation was performed to chimeric MAb (cMAb) U36 and murine MAbs (mMAb) E48 and 425 using a labile ester. Conjugates showed preservation of integrity and immunoreactivity and full stability in serum in vitro. At molar ratios >4, the solubility of the conjugates decreased. Data on the in vitro efficacy of PDT showed that in the chosen experimental setup the internalizing AlPc(SO2Ngly)4-mMAb 425 conjugate was about 7500 times more toxic to A431 cells than the free sensitizer (IC50s, 0.12 nM versus 900 nM). The AlPc(SO2Ngly)4-mMAb 425 conjugate was also more toxic than meta-tetrahydroxyphenylchlorin-mMAb 425 conjugates and free meta-tetrahydroxyphenylchlorin that had been tested previously (M. B. Vrouenraets et al., Cancer Res., 59: 1505-1513, 1999) in the same system (IC50s, 7.3 nm and 2.0 nM, respectively). Biodistribution analysis of AlPc(SO2Ngly)4-125I-labeled cMAb U36 conjugates with different sensitizer:MAb ratios in squamous cell carcinoma-bearing nude mice revealed selective accumulation in the tumor, although to a lesser extent than for the unconjugated 125I-labeled cMAb U36, whereas tumor:blood ratios were similar. These findings indicate that AlPc(SO3H)4 has high potential for use in PDT when coupled to internalizing tumor-selective MAbs.

Animals↗

PubMed2001

Radioimmunotherapy in patients with head and neck squamous cell carcinoma: initial experience.

BACKGROUND: Despite improvements in locoregional treatment of stages III/IV squamous cell carcinoma of the head and neck (HNSCC), local and distant failure rates remain high. An effective adjuvant therapy is required for these patients. Among novel approaches is radioimmunotherapy, in which monoclonal antibodies (MAbs) are used for selective delivery of radiation to tumor cells. METHODS: The suitability of 186Re-labeled chimeric MAb U36 (186Re-cMAb U36) for radioimmunotherapy was evaluated in a phase I study, with radiation dose escalating steps of 11, 27, and 41 mCi/m2. Tumor targeting was monitored with a gamma camera, and the maximum tolerated dose was established in 13 patients with recurrent or metastatic disease. RESULTS: Administrations were well tolerated, and excellent targeting of tumor lesions was seen. Myelotoxicity was the only toxicity observed, resulting in dose-limiting toxicity in two patients treated with 41 mCi/m2. The MTD was established at 27 mCi/m2. A marked reduction in tumor size was observed in two patients, another showed stable disease for 6 months. CONCLUSIONS: Radioimmunotherapy with 186Re-cMAb U36 seems to be well tolerated, with bone marrow being the dose-limiting organ. The observation of antitumor effects is encouraging for further development of radioimmunotherapy for HNSCC.

Aged↗

PubMed2001

Clonally related but phenotypically divergent human cancer cell lines derived from a single follicular thyroid cancer recurrence (TT2609).

Starting from different regional samples taken from a heterogeneous follicular thyroid cancer recurrence in a male patient, a series of cell cultures was initiated. Three stable cancer cell lines were successfully established (TT2609-A02, TT2609-B02, and TT2609-C02) and kept in continuous culture for more than 3 years. The lines are each characterized by a unique set of biological parameters such as morphology, ploidy state, cell proliferation rate, ultrastructure, thyroid marker expression, p53 expression, karyogram, agar clonogenic capacity and tumorigenicity as xenografts in nude mice. These characterization studies point to a marked heterogeneity at the level of the clinical tumor recurrence. Karyotype analysis of the cell lines showed a pattern of aberrations indicating that the lines are clonally related and that the A02 and C02 lines are subsequently derived from the more "original" tumor cell type B02 after a tetraploidization event. It is concluded that the obtained cell lines represent an in vitro/in vivo model for human follicular thyroid cancer. The availability of a series of cell lines for human follicular thyroid cancer, mimicking the biological heterogeneity observed in patient tumors, enables both detailed fundamental investigation of thyroid cancer cell biology and the experimental exploration of new treatment approaches.

Animals↗

PubMed2001

Optimal quality (131)I-monoclonal antibodies on high-dose labeling in a large reaction volume and temporarily coating the antibody with IODO-GEN.

UNLABELLED: A novel, facile procedure for efficient coupling of high doses of (131)I to monoclonal antibodies (MAbs) was developed with minimal chemical and radiation damage. METHODS: To diminish the radiation and chemical burden during labeling, iodination was performed in a large reaction volume and by temporarily coating the MAb with a minimal amount of IODO-GEN. The MAb was coated by injection of IODO-GEN (dissolved in acetonitrile [MeCN]) into the aqueous MAb solution, and the coating was subsequently removed by addition of ascorbic acid. For chemoprotection before, during, and after PD-10 purification of the (131)I-MAbs, ascorbic acid and human serum albumin were used. The effects of autoradiolysis in the starting (131)I solution were countered by treatment with NaOH and ascorbic acid. For this so-called IODO-GEN-coated MAb method, the sensitive chimeric MAb MOv18 (c-MOv18) and the more robust murine MAbs K928 and E48 were used. The high-dose (131)I-labeled MAbs were characterized for radiochemical purity and MAb integrity by thin-layer chromatography, high-performance liquid chromatography, and sodium dodecyl sulfate polyacrylamide gel electrophoresis followed by phosphor imager quantification. The high-dose (131)I-labeled MAbs were also characterized for immunoreactivity. The radiopharmacokinetics and biodistribution of (131)I-c-MOv18 were analyzed in human tumor-bearing nude mice. For comparison, (131)I-c-MOv18 batches were made using the conventional chloramine-T or IODO-GEN-coated vial method. RESULTS: Conventional high-dose labeling of 5 mg c-MOv18 with 4.4 GBq (131)I resulted in a labeling yield of 60%, a radiochemical purity of 90%, an immunoreactive fraction of 25% (72% being the maximum in the assay used), and the presence of aggregation and degradation products. Using similar amounts of (131)I and MAb in the IODO-GEN-coated MAb method, 85%-89% overall radiochemical yield, at least 99.7% radiochemical purity, and full preservation of MAb integrity and immunoreactivity were achieved. For this labeling, 5 mg MAb were coated with 35 microg IODO-GEN during 3 min in a reaction volume of 6 mL. Also, biodistribution was optimal, and tumor accumulation was superior to that of coinjected (125)I-c-MOv18 labeled according to the conventional IODO-GEN-coated vial method. CONCLUSION: A new, facile, high-dose (131)I-labeling method was developed for production of (131)I-labeled MAbs with optimal quality for use in clinical radioimmunotherapy.

Animals↗

PubMed2001

Evaluation of limited blood sampling in a preceding 99mTc-labeled diagnostic study to predict the pharmacokinetics and myelotoxicity of 186Re-cMAb U36 radioimmunotherapy.

UNLABELLED: 186Re-labeled chimeric monoclonal antibody U36 (cMAb U36) was recently evaluated in a phase I dose escalation study in head and neck cancer patients. All 13 patients received 99mTc-labeled cMAb U36 before 186Re-cMAb U36 radioimmunotherapy. The aim of this study was to evaluate the suitability of multiple or limited blood sampling to predict clearance, red marrow absorbed dose, and myelotoxicity of 186Re-cMAb U36. METHODS: Population pharmacokinetics of 186Re-cMAb U36 were analyzed with a nonparametric expectation algorithm (NPEM 2) and used for Bayesian analysis of individual patient data to predict cMAb U36 clearance. RESULTS: 186Re-cMAb U36 clearance was most accurately predicted (r = 0.91, P < 0.001) with limited sampling for sample points 4 and 72 h after administration of 186Re-cMAb U36. These predictions were less accurate with 99mTc-cMAb U36 (r = 0.51, P = 0.078 for multiple sampling; r = 0.47, P = 0.104 for sampling at 4 and 21 h after administration). Thrombocytopenia was found to be correlated with the red marrow absorbed dose and was equally well predicted by limited blood sampling after administration of 99mTc-cMAb U36 (r = 0.81, P < 0.01) or 186Re-cMAb U36 (r = 0.79, P < 0.01). CONCLUSION: Limited sampling seems useful to predict pharmacokinetics and myelotoxicity of 186Re-cMAb U36.

Aged↗

PubMed2000

Targeting of a hydrophilic photosensitizer by use of internalizing monoclonal antibodies: A new possibility for use in photodynamic therapy.

Coupling of photosensitizers to tumor-selective monoclonal antibodies (MAbs) is an attractive option for improving the selectivity of photodynamic therapy (PDT). For this purpose, hydrophilic sensitizers would be most suitable because of their solubility in water. However, such sensitizers are known to be ineffective in PDT, probably because they cannot readily pass the cell membrane and reach the critical intracellular target. We used the model compound TrisMPyP-PhiCO(2)H, a hydrophilic porphyrin derivative, to test the hypothesis that hydrophilic photosensitizers might become of therapeutic value when directed into the tumor cell by use of internalizing MAbs. TrisMPyP-PhiCO(2)H was conjugated using a labile ester. Conjugates showed no impairment of integrity on SDS-PAGE, full stability in serum in vitro, and optimal immunoreactivity when the sensitizer:MAb ratio was </=3. At higher molar ratios, the solubility of the conjugates decreased. In vitro internalization experiments showed that TrisMPyP-PhiCONH-(125)I-cMAb U36 and TrisMPyPPhiCONH-(125)I-mMAb 425 conjugates were internalized by A431 cells, in contrast to TrisMPyP-PhiCONH-(125)I-mMAb E48 conjugates. Data on the in vitro efficacy of PDT with MAb-conjugated TrisMPyP-PhiCO(2)H showed that the internalizing cMAb U36 and mMAb 425 conjugates were phototoxic to A431 cells, while the non-internalizing E48 conjugate and the unconjugated sensitizer were not. Biodistribution data of conjugates with sensitizer:(125)I-cMAb U36 ratios varying from 1:1 to 3:1 in tumor-bearing nude mice revealed selective accumulation in the tumor. Conjugates with higher molar ratios were cleared more rapidly from the blood than the unconjugated (125)I-cMAb U36, resulting in lower tumor uptake but similar tumor-to-blood ratios. Our data suggest that hydrophilic photosensitizers might have therapeutic value when targeted to tumors by internalizing MAbs.

Animals↗

PubMed2000

Safety and biodistribution of 99mTechnetium-labeled anti-CD44v6 monoclonal antibody BIWA 1 in head and neck cancer patients.

The CD44 protein family consists of isoforms, encoded by standard exons and up to nine alternatively spliced variant exons (v2-v10), which are expressed in a tissue-specific way. Expression of v6-containing variants (CD44v6) has been related to aggressive behavior of various tumor types and was shown to be particularly high in squamous cell carcinoma (SCC). Therefore, CD44v6 might be a suitable target for radioimmunoscintigraphy (RIS) and therapy. The present study evaluates the novel high-affinity murine anti-CD44v6 monoclonal antibody (MAb) BIWA 1 for its safety and targeting potential in patients with SCC of the head and neck (HNSCC). Twelve HNSCC patients, who had planned to undergo resection of the primary tumor and neck dissection, were included. Preoperatively, 2, 12, or 52 mg of 99nTc-labeled MAb BIWA 1 was administered. RIS results obtained 21 h after injection were compared with palpation, computed tomography, and magnetic resonance imaging, with histopathology as the gold standard. Moreover, biodistribution of BIWA 1 was evaluated by radioactivity measurement in blood and bone marrow and in biopsies from the surgical specimen obtained 40 h after injection. The distribution of BIWA 1 in tumor biopsies was analyzed by immunohistochemistry. BIWA 1 integrity in the blood was assessed by high-performance liquid chromatography and related to soluble CD44v6 levels in serum samples. No drug-related adverse events were observed. Human antimouse antibody responses were observed in 11 patients. The diagnostic efficacy of RIS appeared to be comparable for the three BIWA 1 dose levels and for the four diagnostic methods. Besides activity uptake in tumor tissue, minimal accumulation of activity was observed in mouth, lungs, spleen, kidney, bone marrow, and scrotal area. Analysis of tissue biopsies revealed high uptake in tumors, with a mean value of 14.2+/-8.4% of the injected dose/kg tumor tissue and a mean tumor:blood ratio of 2.0+/-1.4 at 40 h after injection. Differences among the three dose groups were not statistically significant, although a trend toward lower uptake in the highest dose group was noted. Distribution of BIWA 1 throughout the tumor was heterogeneous for all dose groups, which might be related to the high affinity of the MAb. The mean biological half-life in blood (34.5+/-6.1 h) was not dose dependent. Extensive complex formation of BIWA 1 was observed in the 2-mg group, most probably with soluble CD44v6 present in the blood, and complex formation relatively diminished upon increase of the MAb dose. BIWA 1 is a promising MAb for targeting HNSCC because it can be safely administered to HNSCC patients, while it shows high and selective tumor uptake. However, BIWA 1 is immunogenic, and therefore a chimerized or humanized derivative of BIWA 1 with intermediate affinity will be used in future clinical trials.

Adult↗

PubMed2000

Phase I therapy study of 186Re-labeled chimeric monoclonal antibody U36 in patients with squamous cell carcinoma of the head and neck.

UNLABELLED: A phase I therapy study was conducted to determine the safety, maximum tolerated dose (MTD), pharmacokinetics, dosimetry, immunogenicity, and therapeutic potential of 186Re-labeled anti-CD44v6 chimeric monoclonal antibody (cMAb) U36 in patients with squamous cell carcinoma of the head and neck (HNSCC). The potential of a diagnostic study with 99mTc-cMAb U36 to predict the biodistribution of 186Re-cMAb U36 was evaluated. METHODS: Thirteen patients with recurrent or metastatic HNSCC were given 750 MBq 99mTc-cMAb U36 (2 mg) followed 1 wk later by a single dose of 186Re-cMAb U36 (12 or 52 mg) in radiation dose-escalating steps of 0.4, 1.0, and 1.5 GBq/m2. After each administration, planar and SPECT images were obtained, and the pharmacokinetics and development of human antimurine as well as anti-cMAb responses were determined. Radiation absorbed doses to tumor, red marrow, and organs were calculated. RESULTS: Administration was well tolerated, and excellent targeting of tumor lesions was seen in all patients. Dose-limiting myelotoxicity (thrombocytopenia being most prominent) was the only toxicity observed, resulting in grade 4 myelotoxicity in 2 patients treated with 1.5 GBq/m2. The MTD was established at 1.0 GBq/m2, at which a transient grade 3 thrombocytopenia was seen in 1 patient. One patient showed stable disease for 6 mo after treatment at the MTD. The 2 patients with dose-limiting myelotoxicity showed a marked reduction in tumor size. The reduction was of short duration and, therefore, not considered an objective response. Tumor absorbed doses at MTD ranged from 3.0 to 18.1 Gy. Red marrow doses ranged from 20 to 112 cGy (mean, 51 +/- 16 cGy/GBq) and correlated with platelet nadir (r = 0.8; P < 0.01). Pharmacokinetics varied between patients treated at the same dose level and were accurately predicted by the diagnostic procedure. Five patients experienced a human anti-cMAb response, 1 of which was a human antimouse antibody response. CONCLUSION: This study shows that 186Re-cMAb U36 can be safely administered, with dose-limiting myelotoxicity at 41 mCi/m2. The use of cMAb U36 instead of its murine counterpart did not decrease the induction of human antibody responses. The availability of a 99mTc-labeled diagnostic study that can predict the pharmacokinetics of 186Re-cMAb U36 offers the possibility of using such a study for selection of a safe radioimmunotherapy dose.

Aged↗

PubMed1999

Characterization of CD44v6 isoforms in head-and-neck squamous-cell carcinoma.

CD44 splice variants, especially those containing the v6 domain, are assumed to play a critical role in the malignant progression of many human tumors. This concept was based on (i) the aberrant expression of CD44v6 in malignant cells, often encoded by alternatively spliced transcripts, and (ii) the absence of CD44v6 expression in corresponding normal tissues. Remarkably, data on CD44v6 expression in squamous cells do not support this hypothesis: the v6 domain is highly expressed in normal squamous tissues and down-regulation has been described in the majority of squamous-cell carcinomas of the head and neck (HNSCC). In this study, we have compared the expression of v6 in normal oral mucosa and HNSCC in a qualitative and quantitative way. Immuno-histochemistry was performed with 3 different anti-v6 antibodies (U36, U39 and VFF18) on a large panel of HNSCC cell lines and tumors. The v6-encoding splice variants were characterized by screening a cDNA library of a human HNSCC cell line and by RT-PCR on HNSCC cell lines, microdissected normal mucosa and primary as well as metastatic HNSCC tissue. The results revealed that there is no, or only marginal, down-regulation of CD44v6 in HNSCC. About 97% of the primary HNSCC tumors exhibited a high and homogeneous staining pattern (U36, 270/277; U39, 268/277 tumors with more than 50% positive cells). Furthermore, the v6-containing CD44 splice variants present in HNSCC primary tumors and metastases were identical to those expressed in normal mucosa. Our data indicate that v6-containing CD44 splice variants do not play a role in the malignant progression of HNSCC.

Alternative Splicing↗

PubMed1999

Development of meta-tetrahydroxyphenylchlorin-monoclonal antibody conjugates for photoimmunotherapy.

A limitation of photodynamic therapy is the lack of tumor selectivity of the photosensitizer. To overcome this problem, a protocol was developed for coupling of meta-tetrahydroxyphenylchlorin (mTHPC), one of the most promising photosensitizers, to tumor-selective monoclonal antibodies (MAbs). mTHPC was radiolabeled with 131I to facilitate the assessment of the in vitro and in vivo behavior. After the modification to 131I-mTHPC-(CH2COOH)4, thus increasing the water solubility and creating a functional moiety suitable for coupling, conjugation was performed using a labile ester. Insoluble aggregates were not formed when mTHPC-MAb conjugates with a molar ratio of up to 4 were prepared. These conjugates showed a minimal impairment of the integrity on SDS-PAGE, full stability in serum in vitro, and an optimal immunoreactivity. To test the in vivo behavior of the mTHPC-MAb conjugates, the head and neck squamous cell carcinoma-selective chimeric MAb U36 was used in head and neck squamous cell carcinoma-bearing nude mice. Biodistribution data showed that the tumor selectivity of cMAb U36-conjugated mTHPC was increased in comparison with free mTHPC, despite the fact that conjugates with a higher mTHPC:MAb ratio were more rapidly cleared from the blood. Preliminary results on the in vitro efficacy of photodynamic therapy with MAb-conjugated mTHPC showed that mTHPC coupled to the internalizing murine MAb 425 exhibited more phototoxicity than when coupled to the noninternalizing chimeric MAb U36.

Animals↗

PubMed1999

Sensitive detection of squamous cells in bone marrow and blood of head and neck cancer patients by E48 reverse transcriptase-polymerase chain reaction.

In previous studies, we described the selective reactivity of monoclonal antibody E48 with normal squamous and transitional epithelia and their malignant counterparts and the capacity of monoclonal antibody E48 for selective tumor targeting in head and neck cancer patients. Cloning of the E48 encoding cDNA and elucidation of the gene structure enabled the selection of an intron-spanning primer set for the detection of circulating tumor cells in blood and bone marrow of head and neck cancer patients. Extensive optimizations led to a reproducible reverse transcriptase-PCR assay with an internal standard for RNA quality control and an external standard for sensitivity control. In reconstruction experiments, we were able to reach a reproducible sensitivity of one single tumor cell per 7 ml of blood (2 x 10(7) nucleated cells). When applying this method to patient material, we were able to detect positive signal in 35% of the bone marrow samples (0 of 2 stage II, 0 of 4 stage III, 4 of 11 stage IV, and 4 of 6 recurrences) and 10% of the blood samples (2 of 21) of patients with squamous cell carcinoma of the head and neck. The specificity of the method was demonstrated on 29 blood and bone marrow samples of noncancer controls, which were all negative. Our study shows the feasibility of E48 reverse transcriptase-PCR for the detection of squamous cells in nonsquamous tissues.

Adult↗

PubMed1998

Perspectives of combined radioimmunotherapy and anti-EGFR antibody therapy for the treatment of residual head and neck cancer.

Rhenium-186 based radioimmunotherapy (RIT) may have potential for the treatment of minimal residual disease in patients with squamous cell carcinoma of head and neck (HNSCC). In an effort to enhance the efficacy of RIT, we evaluated the combination of RIT and anti-epidermal growth factor receptor (EGFR) therapy in nude mice bearing established HNSCC s.c. xenografts. For this purpose we used the EGFR-blocking monoclonal antibody (MAb) 425. Treatment of HNSCC-bearing mice with the combination of a single administration of 200 microCi 186Re-labeled MAb U36 as well as 1.1 mg unlabeled MAb 425 showed an enhanced efficacy in comparison to the single treatments. When 500 microCi 186Re-labeled MAb U36 were administered, all tumors eventually regressed completely. The combination of this RIT treatment with multiple injections of MAb 425 significantly increased the rate of tumor regression. Although RIT with 186Re-labeled MAbs appears to be very efficient on HNSCC xenografts, the combination with anti-EGFR MAb 425 may enhance the efficacy.

Animals↗

PubMed1998

The impact of tumour volume and other characteristics on uptake of radiolabelled monoclonal antibodies in tumour tissue of head and neck cancer patients.

Radioimmunotherapy (RIT) seems to be a realistic option for eradication of minimal residual squamous cell carcinoma of the head and neck (HNSCC), although uptake levels of radiolabelled monoclonal antibodies (MAbs) in tumour tissue vary strongly. The aim of this study was to obtain greater insight into the factors influencing the accumulation of MAbs in HNSCC. Twenty-seven HNSCC patients were injected with radiolabelled MAb E48 or U36 and underwent surgery 2 days after injection. Radioactivity was measured in tumour biopsies taken from the surgical specimen. Uptake levels were correlated with various patient, tumour and MAb characteristics, including age, sex, site, TNM stage, volume as assesssed by computed tomography or magnetic resonance imaging, degree of differentiation, antigen expression of the tumour, the particular MAb that had been injected and the MAb dose. A stepwise regression multivariate analysis showed that tumour volume is the most significant prognostic factor (P=0. 01) for MAb uptake. In conclusion, a significantly higher MAb uptake is found in small tumours as compared to larger tumours. Therefore, RIT may be particularly effective in head and neck cancer patients when used in an adjuvant setting.

Aged↗

PubMed1998

[186Re]-labeled mouse and chimeric monoclonal antibody 323/A3: a comparison of the efficacy in experimental human ovarian cancer.

We have investigated whether [186Re]-labeled chimeric monoclonal antibody 323/A3 (MAb c-323/A3) is as effective as [186Re]-labeled mouse 323/A3 (m-323/A3) in the growth inhibition of human ovarian cancer xenografts OVCAR-3 and FMa. [186Re] was conjugated to MAbs with the use of the chelate S-benzoylmercaptoacetyltriglycine (S-benzoyl-MAG3). The maximum number of metal-MAG3 groups that could be conjugated to one MAb molecule accepting a minimal initial increase of the blood clearance (15%) was 8.5 and 2.9 for c-323/A3 and m-323/A3, respectively. With these molar ratios the immunoreactivity of both MAbs was maintained. An inverse relationship was observed between the protein dose of c-323/A3 and its blood clearance. Both [186Re]-c-323/A3 and [186Re]-m-323/A3 were comparable in the inhibition of the tumor growth when higher protein doses were used. Together with the expected lower immunogenicity, our results imply that c-323/A3 is preferable for use in [186Re]-radioimmunotherapy in ovarian cancer patients.

Animals↗

PubMed1998

Synthesis and evaluation of 99mTc/99Tc-MAG3-biotin conjugates for antibody pretargeting strategies.

Four 99mTc-MAG3-biotin conjugates were synthesized to determine their potential use in antibody pretargeting strategies for radioimmunoscintigraphy (RIS). To use these 99mTc-MAG3-biotin conjugates as model compounds for 186Re-MAG3-biotin conjugates for radioimmunotherapy (RIT), nanomolar amounts of 99Tc were added as carrier to 99mTc. The biotin derivatives used for the preparation of the conjugates-biocytin, biotin hydrazide, biotinyl-piperazine, and biotinyl-diaminosuccinic acid-differed at the site that is regarded to be susceptible to hydrolysis by biotinidase present in human plasma. All four conjugates were produced with high radiochemical purity, were stable in PBS, and demonstrated full binding capacity to streptavidin. The 99mTc/99Tc-MAG3-labeled biotinyl-piperazine and biotinyl-diaminosuccinic acid conjugates were stable in mouse as well as human plasma, whereas the corresponding biocytin and biotin hydrazide conjugates were rapidly degraded. The biodistribution in nude mice at 30 min after injection was similar for all conjugates, and a rapid blood clearance and high intestinal excretion were both observed. It is concluded that the metabolic routing of a conjugate containing biotin and MAG3 is dominated by these two moieties. For this reason, MAG3-biotin conjugates do not seem suited for pretargeted RIT, for which quantitative and fast renal excretion is a prerequisite to minimize radiation toxicity. However, in a pretargeted RIS approach the 99mTc-MAG3-biotin conjugates might have potential.

Animals↗

PubMed1997

High dose rhenium-186-labeling of monoclonal antibodies for clinical application: pitfalls and solutions.

BACKGROUND: Rhenium-186 (186Re) has ideal properties for adjuvant radioimmunotherapy (RIT). However, the lack of suitable methods for high dose 186Re labeling of monoclonal antibodies (MAbs) has hampered the use of 186Re in clinical RIT. After development of a chemically identical multistep procedure for the production of 186Re-MAG3-MAb and 99mTc/99Tc-MAG3-MAb conjugates for use as a matched pair, the authors now report on further progress to make this labeling method broadly applicable for high dose 186Re labeling. METHODS: The number of metal-MAG3 groups that can be coupled to a MAb without alteration of the biodistribution was investigated by radioimmunoscintigraphy (RIS) in patients with head and neck squamous cell carcinoma (HNSCC). For labeling with 500 mCi [186Re]ReO4-, an efficient chemoprotection was introduced to suppress the damaging effects of radiation during conjugation and conjugate purification. Furthermore, the authors developed strategies that make the procedure easy and safe to perform at any medical center. RESULTS: MAbs showed a minor variation in biodistribution in HNSCC patients when the number of metal-chelate groups per MAb varied between < 1 and 4.3. High dose 186Re-MAb conjugates (150-250 mCi) with a Re-MAG3:MAb ratio of 3.4 were obtained with a radiochemical purity of > 95% and minimal aggregate formation (< or = 6%). Furthermore, a semiautomated labeling device and a convenient 100 mCi labeling kit in the form of a dried 186Re-MAG3-TFP ester were developed. RIT studies in HNSCC-bearing nude mice showed that high dose 186Re-labeled MAb U36 is more effective than iodine-131-labeled MAb U36. CONCLUSIONS: High dose 186Re-MAb conjugates were prepared that exhibit an optimal stability, immunoreactivity, and pharmacokinetic behavior. The availability of a kit procedure for coupling 186Re to MAbs might open the possibility of a broad application of 186Re in RIT.

Animals↗

PubMed1997

Comparison of the biodistribution and the efficacy of monoclonal antibody 323/A3 labeled with either 131I or 186Re in human ovarian cancer xenografts.

PURPOSE: The radionuclide 186Re has favorable physical characteristics for use in radioimmunotherapy, including the emission of beta-particles of a high-energy and a low-abundance of gamma-emission. The gamma-emission, in particular, is ideal for tumor imaging and poses less hazards to the patient and the medical personnel when compared with the gamma-emission of the widely used radionuclide 131I. In the present study, we determined whether 186Re-labeled monoclonal antibody 323/A3 may be better suited for the treatment of ovarian cancer than 131I-323/A3. METHODS AND MATERIALS: We compared the biodistribution and the efficacy of 186Re- and 131I-labeled 323/A3 in nude mice bearing s.c. the human ovarian cancer xenografts FMa, OVCAR-3 and Ov.Pe. 186Re was conjugated to 323/A3 with the use of the S-benzoylmercaptoacetyltriglycine (S-benzoyl-MAG3) chelate. RESULTS: A molar ratio of Re-MAG3:323/A3 of 3:1 did not affect the integrity and the pharmacokinetic behaviour of the MAb. The tumor uptake and the retention of 186Re- and 131I-labeled 323/A3 were comparable, but the cumulative absorbed radiation dose in the tumor delivered by 186Re-323/A3 was 1.3-fold higher than that of 131I-323/A3. When mice were treated with equivalent radionuclide doses, the tumor growth inhibition induced by 186Re-323/A3 was similar or slightly better when compared with the efficacy of 131I-323/A3. When mice were treated with radionuclide doses that were adjusted to obtain equal cumulative absorbed radiation doses in the tumor for both conjugates, 131I-323/A3 was slightly more effective in the inhibition of the growth of FMa and OVCAR-3 xenografts. CONCLUSIONS: The favorable physical characteristics of 186Re as well as its efficacy when conjugated to a MAb indicate 186Re as an attractive radionuclide in radioimmunotherapy of ovarian cancer patients.

Animals↗

PubMed1997

Rapid elimination of mouse/human chimeric monoclonal antibodies in nude mice.

At our laboratory we are currently evaluating the suitability of mouse/human chimeric monoclonal antibodies (cmAb) for use in radioimmunotherapy of patients with head and neck squamous cell carcinoma (HNSCC). We have developed cmAb containing the human constant IgG1 domain and the variable domains of murine mAb (mmAb) E48 and U36 respectively. We considered the tumour-bearing nude mouse to be a well-validated model for a first testing of the targeting capabilities of these cmAb in comparison with the mmAb. Therefore, 3 microg cmAb E48 (labelled with (125)I) and 3 microg mmAb E48 (labelled with (131)I) were simultaneously injected into HNSCC-bearing nude mice and, at various assay times, mAb uptake in blood and other tissues was assessed. Remarkably, while in roughly 50% of the animals the biodistribution of the conjugates was similar, in the other animals cmAb E48 showed a much higher blood clearance than mmAb E48. This resulted in a lower tumour uptake of cmAb E48 in comparison with mmAb E48. To determine whether this phenomenon was related to mAb E48 or to the animal model, other cmAb-mmAb combinations were evaluated in the same way: cmAbs SF-25, 17-1A and U36 (all IgG1) were tested and all showed a rapid elimination in about 50% of the animals. Besides a decrease in blood concentration, an increase of cmAb levels in liver and spleen was observed within 24 h after injection. Isotype-specific enzyme-linked immunosorbent assays showed that mice that demonstrated a rapid elimination of cmAb from the blood had much lower endogenous IgGI, IgG2b and IgG3 titres than mice showing normal clearance. IgG2a levels were low in all mice. Biodistribution experiments with 3 microg chimeric 17-1A isoforms showed high blood clearance in a proportion of the mice for IgG1, IgG3 and IgG4, but not for IgG2. Increase of the cmAb dose to 100 microg resulted in a similar cmAb and mmAb biodistribution in all mice. Moreover, the biodistribution of the F(ab')2 fragment of an IgG1 cmAb was similar for all mice in contrast to that of coinjected whole IgG. On the basis of these results it can be hypothesized that, in mice with low endogenous IgG titres, cmAb with specific isotypes are rapidly removed from the blood (and ultimately from the body) by mediation of Fc-binding receptors. Apparently, in mice with high endogenous IgG titres or in mice receiving a high cmAb dose, these receptors are saturated. Furthermore, the rapid elimination of cmAb from nude mice, which may occur after injection at a low dose, is a phenomenon related to the nude mouse model.

Animals↗