Radiolabelled Monoclonal Antibodies Research Articles

Biokinetics of the Monoclonal Antibodies MOv 18, OV 185 and OV 197 Labelled with 125I According to the m-MeATE Method or the Iodogen Method in Nude Mice with Ovarian Cancer Xenografts

The biodistribution of the radiolabelled monoclonal antibodies MOv18, OV185 and OV197 in nude mice with subcutaneous tumours of the human ovarian cancer cell line OVCAR3 was investigated. The early uptake of MOv18 (1-24 h) and the uptake in relation to tumour size were also studied. The antibodies were labelled with 125I according to the Iodogen method or the m-MeATE method, the latter also being suitable for labelling with 211

Radioimmunotherapy of DU-145 Tumours in Nude Mice: A Pilot Study with E4, A Novel Monoclonal Antibody Against Prostate Cancer

The anti-tumour effect of the 131I-labelled antiprostate monoclonal antibody (MAb) E4 was studied in an experimental model with 41 nude mice, subcutaneously xenografted with a human prostate cancer cell line (DU-145). The mice were divided into four study groups, i.e. one receiving single and another repeated injections of the radiolabelled MAb. A third group was injected with non-labelled MAb, and the fourth served as an untreated control group. The tumour volumes increased similarly in all groups during the 27-day observation period. The tumour tissue was morphologically disintegrated in the group that received repeated radioimmunotherapy (RIT). The tumours from this group contained large fluid-filled cystic parts and demonstrated pronounced cellular and subcellular polymorphism in the remaining viable tumour tissue. The untreated control tumours and single therapy tumours remained solid. The proportion of the total tumour volume that consisted of viable tumour cells, as determined by morphometric techniques, was significantly lower in the 131I-E4-treated groups. The use of 131I-labelled E4 MAb has thus demonstrated a promising therapeutic potential.

Radiolabelled antimicrobial peptides for imaging of infections: a review.

At present, it is difficult to distinguish between bacterial infections and sterile inflammatory processes using radiopharmaceuticals. This is so for a variety of reasons, including binding to bacteria with low affinity (e.g. infection) and binding to a specific micro-organism (e.g. radiolabelled monoclonal antibodies or F(ab)2 fragments thereof against micro-organisms). In this review, we propose that radiolabelled antimicrobial peptides should be the first choice in the development of new radiopharmaceuticals for imaging of bacterial infections. Antimicrobial peptides are a recently discovered component of the innate defence system of plants, animals and humans. These peptides, which now number more than 100, with proven microbicidal activity against a variety of micro-organisms, share certain properties, such as their small size and cationic charge. The latter allows them to bind preferentially to a broad spectrum of micro-organisms. We have recently demonstrated that radiolabelled human defensins allow the rapid visualization of bacterial infections in mice. Furthermore, binding of this antimicrobial peptide to bacteria is the major factor contributing to the accumulation of this tracer in bacterial infections. Based on these considerations, we believe that radiolabelled antimicrobial peptides will be an important asset in the imaging of infections in patients.

Resection Guided by Antibodies (REGAJ) a Reappraisal after 10 Years

Between 1986 and 1990 50 patients with ovarian cancer were submitted to a procedure which we called REGAJ-resection guided by antibodies. Using the radiolabelled murine monoclonal antibody OC 125, microscopic ovarian tumors producing CA 125 were detected during second-look surgery by a hand-held probe. Retrospective analysis after 10 years revealed that this procedure resulted in the cure of 4 of 11 patients, who would otherwise have been considered tumor free during second-look surgery and not further treated. Previous problems associated with the method can now be solved so that the clinical value of the procedure should be reconsidered.

Biodistribution and localization of radiolabeled NR-LU-10 fab fragment in human breast cancer xenografts

Radioimmunodetection, which takes advantage of tumor-specific or tumor-associated radiolabeled monoclonal antibodies or other biologic molecules to diagnose the extent of disease in cancer patients, has been of limited use in studies to date in patients with breast cancer. The difficulty is in finding an antibody that is both sensitive and specific enough to localize in breast tumors. This study undertook immunohistochemical and in vivo evaluation of tumor localization and biodistribution of NR-LU-10 Fab (antibody fragment) in breast tumors to determine its ability to bind selectively to malignant tissue. NR-LU-10 Fab recognizes a pancarcinoma glycoprotein antigen found on tumors of epithelial cell origin. NR-LU-10 Fab reacted with 6/6 (100%) breast cancer cell lines and 14/16 (87.5%) breast tumors with varying degrees of immunostaining intensities. Athymic mice bearing ZR-75-1 breast cancer xenografts were injected with 125I-labeled NR-LU-10 Fab (12 μg/5 μCi) and sacrificed at fixed time intervals. These studies demonstrated the highest tumor uptake of labeled Fab at 12 h postinjection (4.58 ± 1.59% of injected dose/gram [% ID/g] of tissue); this gradually decreased to 0.13 ± 0.05% ID/g of tissue by 72 h postinjection of the radiolabeled Fab. Biolocalization to normal tissues was as predicted for a Fab fragment; i.e., initially high in clearance organs (kidney), followed by rapid clearance over the 72-h test period. NR-LU-10 Fab displays adequate breast tumor localization with minimal biolocalization to normal tissues, thus supporting its potential use in radioimmunoscintigraphy and the RIGS® system (radioimmunoguided surgery).

Orthotopic xenografts of human melanoma and colonic and ovarian carcinoma in sheep to evaluate radioimmunotherapy.

Extrapolation to humans from experimental radioimmunotherapy in nude mouse xenograft models is confounded by large relative tumour size and small volume of distribution in mice allowing tumour uptake of radiolabelled antibodies unattainable in patients. Our large animal model of human tumours in cyclosporin-immunosuppressed sheep demonstrated tumour uptake of targeted radiolabelled monoclonal antibodies comparable with uptakes reported in clinical trials. Sheep immunosuppression with daily intravenous cyclosporin augmented by oral ketoconazole maintained trough blood levels of cyclosporin within the range 1000-1500 ng ml(-1). Human tumour cells were transplanted orthotopically by inoculation of 10(7) cells: SKMEL melanoma subcutaneously; LS174T and HT29 colon carcinoma into bowel, peritoneum and liver; and JAM ovarian carcinoma into ovary and peritoneum. Tumour xenografts grew at all sites within 3 weeks of inoculation, preserving characteristic morphology without evidence of necrosis or host rejection. Lymphatic metastasis was demonstrated in regional nodes draining xenografts of melanoma and ovarian carcinoma. Colonic LS1 74T xenografts produced mucin and carcinoembryonic antigen (CEA). The anti-CEA IgG1 monoclonal antibody A5B7 was radiolabelled with iodine-131 and administered intravenously to sheep. Peak uptake at 5 days in orthotopic human tumour transplants in gut was 0.027% DI g(-1) (percentage of injected dose per gram) and 0.034% DI g(-1) in hepatic metastases with tumour to blood ratios of 2-2.5. Non-specific tumour uptake in melanoma was 0.003% DI g(-1). Uptake of radiolabelled monoclonal antibody in human tumours in our large animal model is comparable with that observed in patients and may be more realistic than nude mice xenografts for prediction of clinical efficacy of radioimmunotherapy.

Dosimetric comparison of bolus and continuous injections of CC49 monoclonal antibody in a colon cancer xenograft model

Improved understanding of dose and effective dose calculations may contribute to the optimization of fractionated radioimmunotherapy.Comparison three-dimensional tumor dosimetry was performed on athymic nude mice bearing established LS174T human colon carcinoma xenografts. Mice were given bolus intraperitoneal injections of 300 microCi 131I-labeled CC49 monoclonal antibody once (Day 0) or three times (Days 0, 3, and 7) or continuous intraperitoneal infusion with miniosmotic pumps over 7 days. Serial section autoradiography was used to reconstruct tumor activity density distributions for Days 3, 4, 7, 10, and 11 (single injection); Days 3, 4, 7, 8, and 11 (3 injections); and Days 4, 7, 10, and 13 (pump). At least three tumors were reconstructed at each time point. Uptakes in blood and tumor were measured up to 14 days (single injection), 11 days (3 injections), or 16 days (pump) after injection.Average dose values calculated from total activity uptake data only (assuming no energy loss external to the tumor) yielded 102 Gy (single injection), 158 Gy (three injections), and 47 Gy (pump). Average doses using three-dimensional dose calculations were 88 Gy, 139 Gy, and 40 Gy, respectively. The nonuniformity of dose deposition affects treatment outcome, because cell loss is an exponential function of dose. Using the linear quadratic model with fractional cell survival to define an effective dose, D(eff) were calculated to be 20 Gy, 23 Gy, and 14 Gy, respectively. Cell proliferation affects outcome for variable dose-rate treatments. With cell proliferation parameters set to reproduce single-fraction 60Co recurrence results, D(eff) (for local control endpoint) were 8.9 Gy, 12.8 Gy, and 3.9 Gy, respectively. Three bolus injections compared with a single bolus injection were relatively less efficient in tumor uptake. However, three bolus injections resulted in a more uniform dose rate over a longer period, resulting in a 50% improvement in D(eff). The slower dose delivery for pump infusion resulted in a significantly lower D(eff), although dose-rate distributions were more uniform compared with the single bolus injection.Improvement in dose-rate nonuniformities was observed for fractionated and continuous radiolabeled monoclonal antibody injections. Fractionated injections produced superior dosimetric results compared with single bolus or continuous injections.

Nuclear medicine imaging in lung cancer

The evaluation of the anatomical dissemination of lung cancer has a pivotal role in the choice of the most appropriate treatment modality. The techniques of nuclear medicine are founded on the use of different radiopharmaceuticals capable of exploiting the specific characteristics of malignant tissues. They may recognise diverse cell densities, growth rates, metabolic pathways, antigenic and surface receptor expressions. In the past, the use of Co-57-bleomycin and, then, of (67)Gallium has encountered a mixed acceptance among nuclear medicine specialists, with favourable reports claiming their utility, and others with more sceptical opinions. It is generally admitted that both Co-57-bleomycin and (67)Gallium scintigraphies are quite sensitive and rather accurate. Their use, however, is almost abandoned in favour of more innovative and encouraging approaches, including non-specific radio-tracers ((201)Thallium and Tc-99m-sestamibi), substances useful in particular clinical applications (the somatostatin analogues I-123-tyr(3) and the In-111 octreotide for neuronendocrine tumours), radio-labelled monoclonal antibodies, and the recently introduced positron emission tomography. Promising results with each of these techniques need to be further substantiated, before their entering into clinical practice. However, the abundance of choices offered by nuclear medicine might reasonably bring forward the ideal noninvasive test. We review the many scintigraphic methods investigated so far and their clinical significance.

Distribution of radiolabelled anti-CA125 monoclonal antibody OC125-F(ab′)2-fragment following resection guided by antibodies (REGAJ) in ovarian cancer patients

Ovarian cancer is a highly malignant tumor of mainly postmenopausal women. The long-term prognosis of this malignancy is largely determined by micrometastasis present at the time of second-look surgery. In general, patients face a poor outcome. New radio-immunoscintigraphic methods to target tumor tissue specifically via antigen-antibody binding were developed. However, few studies so far investigated the pattern of in vivo distribution of radiolabelled mAbs and/ or the specificity of antigen-antibody interaction. In this study we examined the immunological interaction and distribution of 131l-OC125-F(ab')2-fragment, an anti-CA-125 mAb, in patients with CA-125 positive ovarian malignancies. Sixteen patients with primarily CA-125 positive gynecological tumors underwent REGAJ surgery. Biopsies of tumor tissue and not tumor infiltrated tissue, serum, and ascites were sampled during or prior to REGAJ surgery, respectively. After preparation of tissue cytosols, samples were assessed for CA-125 and radioactive uptake. By radiochromatography immunological analysis for presence of the target antigen CA-125, the mAb 131l-OC125-F(ab')2-fragment, and immune complexes was performed on different specimen. CA-125 concentrations were higher in serum samples, ascites, and malignant tissue biopsies of malignoma patients compared to those without signs of malignant disease. CA-125 was higher in the tissue cytosol than in the cell membrane fraction. Gel filtration revealed CA-125 with moieties of 75,000 to > 600,000 d. Accumulation of radioactivity was more frequently associated with the presence of unbound 131l-OC125-F(ab')2-fragment or high molecular weight immune complexes. Radioactive uptake, however, was not confined to tissue of high CA-125 expression. Moreover, both immune complex as well as 131l-OC125-F(ab')2-fragment could be isolated from cytosols of tissue not infiltrated by tumor cells as well. Our study demonstrates that the majority of CA-125 is located intracellularly and thus inaccessible to 131l-OC125-F(ab')2-fragment per se. The uptake of 131l-OC125-F(ab')2-fragment into the cytosol of tumor-free and malignant tissue samples prompts us to speculate that certain mechanisms for antigen-specific and nonspecific cellular trafficking of mAbs do exist. We present a model to explain our observations.

A local hyperthermia treatment which enhances antibody uptake in a glioma xenograft model does not affect tumour interstitial fluid pressure

Solid tumours have an elevated interstitial fluid pressure (IFF) due to the lack of normal lymphatics, increased permeability of tumour vasculature and an expanding cell population within a potentially limited space. This elevated IFF has been proposed to be an important barrier to the delivery of drugs and marcromolecules. We have demonstrated that local hyperthermia (4h, 41.8°C) is capable of significantly enhancing the uptake of radiolabelled monoclonal antibodies (mAbs) in D-54 MG glioma xenografts grown subcutaneously in athymic mice. To determine if this increased uptake was attributable to alterations in the tumour IFF, pressure measurements using the wick-in-needle technique were made in tumours after hyperthermia treatment. These pressure measurements were taken at various time points from 4 to 90 h following the initiation of the hyperthermia and compared with pressures taken concurrently in untreated tumours. In addition, pressures were measured following a 2h, 41.8°C hyperthermia treatment, a protocol which does not result in elevated uptake of radiolabeled mAbs. No significant differences were seen at any time point in IFF measured in the tumours receiving either hyperthermia treatment when compared with untreated tumours. Thus, we conclude that the mechanism by which this hyperthermia regimen enhances mAb uptake in this human glioma xenograft model is not due to alternations in tumour IFF.

Relationship between ZCE 025 blood clearance and liver accumulation in patients with colorectal cancer

Clinical interest in radiolabelled monoclonal antibodies (mAbs) derives from their ability to target tissues and deliver radioisotopes thereto. Diagnostic and therapeutic trials suggest that the potential of mAbs would be optimized with better understanding of factors which influence blood clearance. This study sought to advance that understanding. Nine colon cancer patients were injected with 20 mg111In radiolabelled ZCE 025 mAb, then laser-positioned and imaged daily for one week via nuclear medicine views. Laser-positioned CT images were also obtained, enabling image registration. Blood samples were taken at 30, 60, and 90 min and at every imaging time point. Sample activity was plotted to estimate the elimination half-life (t12 β). Liver radiation dose was calculated from the images and evaluated for total organ (mCi-h) and region of interest (μCi-h/g). Laboratory values and patients' physical parameters were also studied to determine their impact on liver uptake and mAb blood clearance. Liver radiation dose, as measured in this study, correlated with terminal half-life of the mAb in blood; patients' liver enzymes and physical parameters did not. Radioactivity in a 48-h or 72-h blood sample correlates with t12 β and liver radiation dose, suggesting that a single 72-h sample can predict both.

Anti-islet autoantibodies detected by monoclonal antibody 1A2: further studies suggesting a role in the pathogenesis of IDDM.

Insulin-dependent diabetes mellitus (IDDM) is associated with autoantibodies to several pancreatic islet antigens. We have described an assay in which autoantibodies displace a radiolabelled monoclonal anti-islet antibody. Sera from 87% of 429 children at time of diagnosis of IDDM were positive, while sera from control groups had much lower prevalences (1.3-19%). Sera from 41.9% of diabetic subjects remained positive after 20 years duration of IDDM. Sera from 23.6% of parents and 37.9% of non-diabetic siblings were positive. Twenty relatives who subsequently developed IDDM had the same prevalence of the antibodies (85%) as did the patients at time of diagnosis. These findings confirm that the autoantibodies detected by monoclonal antibody (mAb) 1A2 are common at the onset of IDDM and their presence prior to the onset of hyperglycaemia suggests that this method may be useful in screening non-diabetic populations. The high prevalence of antibodies in relatives reduces the efficacy for diabetes prediction, but suggests either that generation of these antibodies is an autosomal dominant trait, or that the antigen detected by these antibodies is cross-reactive with a common environmental antigen. Differentiation between these hypotheses will await the identification of the specific islet-cell antigen detected by mAb 1A2.

Detection of pseudomyxoma peritonei by radioimmunohistochemistry and radioimmunoscintigraphy.

Pseudomyxoma peritonei (PP) is a local slowly progressing disease with typical abdominal swelling. Treatment is uneffective and the long-term prognosis is poor. Conventional radiology provides usually only a delineation of low density area relating gelatinous masses accumulating in the peritoneal cavity. In this study, immunohistochemistry based on digital quantitative autoradiography utilizing radiolabelled monoclonal antibody B72.3 (MoAb) recognizing TAG-72 antigen on epithelial carcinomas was used for diagnosis of pseudomyxoma (7 patients). The PP patients were studied with radioiodinated I-131-labeled MoAb after intravenous (2 patients) and intraperitoneal (7 patients) injections. Radioactivities of MoAbs varied considerably in the tumors. Both intra- and extracellular staining pattern was observed by immunohistochemistry. Gamma imaging at 1, 3, 7 and 14 days after i.v. injection (2 patients) revealed targeting of all known lesions. The intraperitoneally injected MoAb (7 patients) retained long time in the peritoneal cavity, specific tumour targeting was seen up to 16 days by an antibody-SPECT, while maximum blood radioactivity was measured between 8-12 hrs. Radiolabelled B72.3 MoAb recognizing TAG-72 antigen is also present within pseudomyxoma cells. It can be used for radioimmunohistochemistry of PP. Accurate imaging of PP is possible by MoAb suggesting earlier diagnosis and more accurate location of residual disease after operations, and evaluating treatment response. Estimated tumour dose for intraperitoneal tumour (MIRD formalism) was 13 mGy/MBq. This indicates that the radioiodinated B72.3 antibody can be used for in vivo targeting and therapeutic applications of intraperitoneal pseudomyxoma.

Carcinoembryonic antigen expression, antibody localisation and immunophotodetection of human colon cancer liver metastases in nude mice: a model for radioimmunotherapy.

Colorectal cancer frequently disseminates through the portal vein into the liver. In this study, outbred Swiss nude mice were adapted to facilitate the induction of liver metastases by a pre-grafting treatment with 6 Gy total body irradiation and i.v. injection of anti-asialo GM1 antibody. One day later, cultured LS 174T human colon cancer cells were injected into the surgically exposed spleen, which was resected 3 min later. In 48 of 65 mice, a few to several hundred liver metastases were macroscopically observed at dissection 3 to 4 weeks after transplantation. Ten of 10 mice, followed-up for survival, died with multiple large confluent liver metastases. By reducing the radiation dose to 4 or 0 Gy, or omitting the anti-asialo GM1 antibody injection, only 60%, 37% or 50% of mice, respectively, had visible metastases 3 weeks after transplantation. Carcinoembryonic antigen (CEA) measured in tumour extracts was in the mean 25.6 micrograms/g in liver metastases compared with 9.2 micrograms/g in s.c. tumours. Uptake of radiolabelled anti-CEA monoclonal antibody (MAb) in the metastases 12, 24 and 48 hr after injection gave a mean value of 39% of the injected dose per gram of tissue (ID/g). In comparison, MAb uptake in s.c. and intrasplenic tumours or lung metastases gave a mean percentage ID/g of 20, 18 and 15, respectively. Laser-induced fluorescence after injection of indocyanin-MAb conjugate allowed direct visual detection of small liver metastases, including some that were not visible under normal light. Preliminary results showed that mice, pre-treated with 4 Gy irradiation and the anti-asialo GM1 injection, were tolerant to radioimmunotherapy with a total dose of 500 muCi 131I labeled anti-CEA intact MAbs given in 3 injections.

Effect of interferon-alpha-2b on the enhancement of tumour uptake of 99Tcm-labelled monoclonal antibodies.

We have previously shown that certain biological response modifiers (BRMs) can significantly increase the tumour uptake of radiolabelled monoclonal antibodies (MAbs). Among the three different BRMs (Ukrain, Pokeweed mitogen and interferon) we evaluated, interferon gave us the best results. A series of experiments were conducted here to further elicit the influence of interferon on the enhancement of tumour uptake. Interferon-alpha-2b (IFN-alpha-2b) was used as a biological response modifier, human melanoma xenografts grown in athymic nude mice as a tumour model, and anti-human melanoma specific ME31.3 as the MAb. The MAb was labelled with technetium-99m by a direct labelling method. IFN-alpha-2b was administered in different doses via intravenous, intramuscular or intratumoral routes. The results showed a significant (P < or = 0.05) increase in absolute tumour uptake of radiolabelled ME31.3 in all IFN-alpha-2b-treated groups. The maximum increase (from 2.0 +/- 1.4% in the control group to 7.2 +/- 1.2% in the experimental group, P < 0.01) was obtained when 20K international units of IFN-alpha-2b were injected intramuscularly.

Stability and Structure of Activated Macrocycles. Ligands with Biological Applications.

Single p-toluic acid pendant groups were attached to 1,4,7,10,13-pentaazacyclopentadecane (15aneN5) and 1,4,8,11-tetraazacyclotetradecane (cyclam) to prepare bifunctional reagents for radiolabeling monoclonal antibodies with (64,67)Cu. The ligands are 1,4,7,10,13-pentaazacyclopentadecane-1-(alpha-1,4-toluic acid) (PCBA) and 1,4,8,11-tetraazacyclotetradecane-1-(alpha-1,4-toluic acid) (CPTA). For the parent macrocycles and their pendant arm derivatives, the 1:1 Cu(2+) complexes dissociate only below pH 2. At pH 0.0 and 25 degrees C the CPTA-Cu complex has a half-life toward complete dissociation of 24 days. A new approach was developed for the estimation of the Cu(2+) stability constant for the kinetically robust CPTA. All other formation constants were determined at 25.0 degrees C with batch spectrophotometric techniques. Potentiometric titrations were used to determine the protonation constants of the macrocyclic ligands as well as of the metal chelates. The protonation constants, stability constants, and pM's are discussed in terms of both molecular mechanics calculations and the ligands' potential applicability as copper(II) radiopharmaceuticals.

Comparison of four bifunctional chelates for radiolabeling monoclonal antibodies with copper radioisotopes: biodistribution and metabolism.

The bifunctional chelating agents (BFCs), 6-[p-(bromoacetamido)benzyl]-1,4,8,11-tetraazacyclotetradecane-1,4 ,8, 11-tetraacetic acid (BAT), 6-[p-(isothiocyanato)benzyl]-1,4,8, 11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid (SCN-TETA), 4-[(1,4,8,11-tetraazacyclotetradec-1-yl)methyl]benzoic acid (CPTA), and 1-[(1,4, 7,10,13-pentaazacyclopentadec-1-yl)methyl]benzoic acid (PCBA), were synthesized and conjugated to the anti-colorectal monoclonal antibody (mAB), 1A3, and antibody fragments, 1A3-F(ab')2, for radiolabeling with 64,67CU and comparison in animal models. In vivo metabolism studies were carried out in liver and kidneys in order to correlate the nature of the metabolites formed to the uptake and retention of the radiolabel in each organ. Animal biodistribution studies were performed in Golden Syrian hamsters bearing the GW39 human colon cancer tumors and in normal Sprague-Dawley rats. All conjugates showed good tumor uptake in hamsters. Biodistribution in rats showed that 64CU-BAT-2IT-1A3 had the lowest liver and kidney uptake of the intact 1A3 conjugates (p < 0.03), whereas in hamsters, there were no significant differences in liver and kidney uptake between the four intact BFC-1A3 conjugates. Tumor-bearing hamsters injected with 64CU-CPTA-1A3-F(ab')2 and 64CU-PCBA-1A3-F(ab')2 had from 3 to 7 times greater uptake in the kidneys than hamsters given 64CU-labeled BAT and SCN-TETA 1A3-F(ab')2 conjugates, while rats injected with 64Cu-CPTA-1A3-F(ab')2 and 64Cu-PCBA-1A3-F(ab')2 had nearly twice the uptake. The in vivo metabolism of the mAbs 1A3 and 1A3-F(ab')2 radiolabeled with 67Cu through the SCN-TETA, CPTA, and PCBA BFCs was investigated by excising the livers and kidneys of normal rats from 1-5 days post-injection of the radiolabeled conjugates. Liver and kidney homogenates were analyzed by size exclusion chromatography and thin layer chromatography (TLC). The size exclusion chromatography data showed that all of the 67Cu-labeled 1A3-F(ab')2 conjugates were > 85% degraded in the kidneys to small molecular weight metabolites by 1 day post-injection. In contrast, in the liver at 1 day post-injection, greater than 70% of the 67Cu-labeled 1A3 conjugates were unmetabolized. By day 5, a 35 kDa peak appeared in the liver of rats injected with the 67 Cu-labeled 1A3 conjugates, possibly due to transchelation of the 67Cu to proteins. Superoxide dismutase chromatographically elutes at the same retention time as this 67Cu-labeled metabolite. The TLC data indicate that the low molecular weight metabolite (< 5 kDa) of both 67Cu-CPTA-1A3 and 67Cu-CPTA-1A3-F(ab')2 conjugates co-chromatographed with a 67Cu-CPTA-epsilon-lysine standard. Our data suggest that chelate charge and lipophilicity play a large role in kidney retention of 64/67Cu-labeled BFC-1A3-F(ab')2 conjugates, while transchelation of the copper label appears to be the major factor for liver accumulation of 64/67Cu-labeled BFC-1A3 conjugates.

980 New drugs in the treatment of malignant lymphomas

The development of new drugs has been greatly hampered in the field of lymphomas by many methodological problems, mainly related to the heavy pre-treatment of many patients. These methodological problems will be discussed. During the last years, however, a few interesting new compounds have emerged. The most exciting ones belong to the family of the purine-analogues, among them Fludarabine and 2-CdA (2-Chlorodeoxyadenosine) have been widely tested in many phase II trials. In selected series the response rate has approached 75–80% depending on the selection of the patients. Currently, phase II trials in previously untreated patients as well as randomized phase III trials are ongoing. Further developments in this area are represented by immunotoxins, ra-diolabelled monoclonal antibodies and tumour vaccines. Therapeutic possibilities as well as still existing methodological weaknesses of these approaches will be discussed.

Monoclonal antibody Po66 uptake by human lung tumours implanted in nude mice: effect of co-administration with doxorubicin.

The efficacy of radioimmunotherapy of tumours with radiolabelled monoclonal antibodies (MAbs) depends on the amount of antibody taken up by the tumour and on its intratumoral distribution. In the case of MAbs directed against intracellular antigens, increasing the permeability of the cytoplasmic membrane may augment the bioavailability of the antigen for the antibody. This raises the question whether the induction of tumour necrosis by chemotherapy can enhance the tumour uptake of radiolabelled monoclonal antibodies. In this work, the effect of doxorubicin on the biodistribution of Po66, an MAb directed against an intracellular antigen, was studied in nude mice grafted with the human non-small-cell lung carcinoma cell line SK-MES-1. After injection on day 0 of 125I-labelled Po66, tumour radioactivity increased up to days 3-5, and then remained unchanged to day 14. The combined administration of 125I-labelled Po66 with 8 mg kg-1 doxorubicin, in two doses separated by 7 days, doubled the radioactivity retained by the tumour. Histological and historadiographic analysis showed, however, that the drug induced cellular damage. In the absence of doxorubicin, the accumulation of Po66 was restricted to some necrotic areas, whereas with doxorubicin the necrosis was more extensive and the antibody more evenly distributed. These results suggest that chemotherapy and immunoradiotherapy combined would enhance tumour uptake of radioisotope and promote more homogenous distribution of the radiolabelled MAb. This would promote eradication of the remaining drug-resistant cells in tumours.

144 Distribution kinetics of radiolabelled monoclonal antibodies (MAB) in experimental tumours following interstitial and systemic administration

In this study we compared the pattern of distribution of intralesional (IL) vs intravenous (IV) I¹²⁵ labelled MAb within a transplantable tumour (HSN), the adjacent tissue (muscle) and systemic organs. Both a specific (ALN/11/53) and non-specific control (ICR2) MAb, were evaluated to determine the importance of MAb specificity for these routes of administration. The distribution of ¹²⁵I was assessed by autoradiography and gamma counting in 37 rats at 4 time points post-infusion up to 72 hours. There was a three-fold greater regional percentage uptake/g of IL vs IV MAb (Student's t-test, <i>P</i>=0.012). Specific MAb had a 14% greater uptake than non-specific MAb following IL administration (<i>P</i>=0.028). The systemic organ uptake was lower with IL than IV administration (<i>P</i>=0.047). Up to 24 hours post-infusion, there was a prominent leakage pattern of ¹²⁵I within the surrounding muscle. At later time points, although the activity was not distributed across the whole tumour, there was a sharpe demarcation at the tumour/muscle junction indicating a tendency for the radioactivity to conform to the shape of the tumour. The higher ¹²⁵I activity and favourable distribution attributes suggest a role for further evaluation of intralesional radiolabelled MAb therapy in patients with localised malignancy.