Effects of temperature on radiochemical purity and immunoreactivity of radiolabeled monoclonal antibody 1H10

OBJECTIVE: This study was undertaken to investigate the effects of temperature on preserving the radiochemical purity and immunoreactivity of 125I- and 131I-labeled monoclonal antibody (MAb) 1H10--an antibody against human cervical carcinoma cell-surface antigen. METHODS: An antibody-irrelevant human melanoma cell line, H2269, served as the control group. Iodine-125 and 131I radiolabeling of MAbs 1H10 and H2669 was performed by the chloramine-T method. All the prepared MAbs were divided into aliquots and stored at 4, -20, and -70 degrees C for 2-14 d. The radiochemical purity and immunoreactivity of the labeled antibodies in set conditions were measured by thin-layer chromatography and a modified index, respectively, after a single freeze-and-thaw cycle. RESULTS: Reduced release of free radioiodide and better preservation of immunoreactivity were observed in the radiolabeled MAbs stored at -70 degrees C than in those stored at -20 degrees C or 4 degrees C. The extent of free iodide dissociation and immunologic binding degradation of 125I-labeled MAb 1H10 appeared milder than that of 131I-labeled MAb under the same conditions. However, both 125I- and 131I-labeled MAb stored at -70 degrees C or -20 degrees C retained more than 90% radiochemical purity for at least 3d. CONCLUSION: Freezing provides an appropriate alternative for reducing radiolysis and preserving immunoreactivity of radioiodinated MAbs. MAb 1H10, labeled with either 125I or 131I and stored at temperatures of -20 degrees C or below for 3 d after labeling, appeared stable in both radiolabeling and binding studies in vitro and was still acceptable for in vivo use.     

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

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.     

c-erbB2 protein overexpression in breast cancer as a target for PET using iodine-124-labeled monoclonal antibodies.

ICR 12, one of a panel of rat monoclonal antibodies recognizing the external domain of the human c-erb B2 proto-oncogene product, (Styles, 1990) was chosen as a candidate for radiolabeling with 124I for positron emission tomography of selected patients with breast cancer. By using N-bromosuccinimide (NBS), optimal labeling conditions were established using 125I. The labeling efficiency was determined using instant thin-layer chromatography (ITLC) and gel filtration (HPLC). The antibody was then labeled with the positron emitter 124I, and a labeling efficiency of 96% and immunoreactivity of 80%-90% was obtained. The product was stable, with less than 5% of the radiolabel being eluted after six days storage in plasma at 37 degrees C. Immunolocalization studies were performed in athymic mice bearing human breast carcinoma xenografts overexpressing the c-erb B2 gene product using as controls 125I labeled isotype-matched rat antibody, and antigen-negative tumors. Good uptake of 124I-labeled ICR12 was obtained in c-erb B2 expressing tumors (up to 12% injected dose per gram at intervals up to 120 hr), with localization indices of 3.4-6.2. Tumor xenografts of 6 mm diameter were successfully imaged with high resolution at 24, 48 and 120 hr using the RMH/ICR MUP-PET camera. We suggest that 124I-labeled ICR12 is a suitable agent to image and quantify immunolocalization in patients whose tumors overexpress the c-erb B2 proto-oncogene product.     

Radioimmunolocalization of neuroblastoma xenografts with chimeric antibody chCE7.

This study was performed to evaluate the tumor targeting ability of chCE7 with a view to clinical applications in neuroblastoma imaging and therapy. A chimeric (mouse/human) monoclonal antibody (chCE7) of gamma 1/kappa isotype directed against a neuroblastoma-associated cell-surface glycoprotein is described. In vitro chCE7 binds with high affinity (KD approximately 1 x 10(-10) M) to SKN-AS human neuroblastoma cells. Binding studies with 125I-labeled chCE7 show temperature-dependent modulation of antigen binding and indicate that a proportion of the bound antibody is internalized due to rapid antigen turnover. In vivo biodistribution of radioiodinated chCE7 in nude mice bearing SKN-AS tumors shows optimal tumor uptake after 24 hr with about 30% of the injected dose per g. Optimal tumor/blood ratios (3.4:1) are reached after 4-5 days. Uptake in other organs including the reticuloendothelial system is low with tumor/organ ratios of 10 and more. Tumor uptake of chCE7 and the parent murine CE7 are found to be similar. Stability of chCE7 during and after radiolabeling is good with no loss of immunoreactivity in preparations labeled with 123I up to 100 mCi/mg and 80% immunoreactivity after labeling with 13 mCi/mg of 131I. Neuroblastoma xenografts can be imaged by radioimmunoscintigraphy with 123I- and and 131I-labeled chCE7.     

Isotype switch variant anti-idiotype monoclonal antibodies: comparative radiolabeling and in vitro binding

The influence of antibody isotype on radiolabeling efficacy and immunoreactivity has been difficult to systematically examine as antibodies of different isotypes generally vary in both constant and variable regions. The recent ability to isolate class or isotype switch monoclonal antibodies that have identical binding regions but different constant regions allows for such a comparison to be undertaken. Two isotype switch variant families of murine anti-idiotypic monoclonal antibodies were studied for radioiodination efficacy and immunoreactivity following labeling. In one of the families (S3H5), the IgG2a isotype switch variant derived from an IgG2b parent had nearly 70% greater iodine incorporation and over 50% greater immunoreactivity than the IgG2b parent. In the other family (S5A8), the IgG2a isotype switch variant had virtually identical efficacy of iodine incorporation and binding to antigen after labeling as did its IgG2b parent. Differences in relative heavy and light chain iodine incorporation were seen among isotype switch variants and their parents regardless of alterations in quantitative iodine incorporation or immunoreactivity. We conclude that in certain instances, cloning of an isotype switch variant antibody can result in an antibody offspring that has superior radiolabeling characteristics to its parent antibody. This isotype switching approach may find utility in converting highly-specific, but difficult to label antibodies, to more useful agents.     

131I radioimmunotherapy and fractionated external beam radiotherapy: comparative effectiveness in a human tumor xenograft.

This article compares the effectiveness of radiation delivered by a radiolabeled monoclonal antibody, 131I-labeled A33, that targets colorectal carcinoma, with that of 10 fractions of conventional 320 kVp x-rays. METHODS: Human colorectal cancer xenografts (SW1222) ranging between 0.14 and 0.84 g were grown in nude mice. These were treated either with escalating activities (3.7-18.5 MBq) of 131I-labeled A33 or 10 fractions of 320 kVp x-rays (fraction sizes from 1.5 to 5 Gy). Tumor dosimetry was determined from a similar group of tumor-bearing animals by serial kill, tumor resection and counting of radioactivity in a gamma counter. The relative effectiveness of the two radiation therapy treatment approaches was compared in terms of tumor regrowth delay and probability of tumor cure. RESULTS: The absorbed dose to tumor per MBq administered was estimated as 3.7 Gy (+/-1 Gy; 95% confidence interval). We observed a close to linear increase in tumor regrowth delay with escalating administered activity. Equitumor response of 1311 monoclonal antibody A33 was observed at average radiation doses to the tumor three times greater than when delivered by fractionated external beam radiotherapy. The relationship between the likelihood of tumor cure and administered activity was less predictable than that for regrowth delay. CONCLUSION: The relative effectiveness per unit dose of radiation therapy delivered by 131I-labeled A33 monoclonal antibodies was approximately one third of that produced by fractionated external beam radiotherapy, when measured by tumor regrowth delay.     

Relative therapeutic efficacy of (125)I- and (131)I-labeled monoclonal antibody A33 in a human colon cancer xenograft.

A33, a monoclonal antibody that targets colon carcinomas, was labeled with (125)I or (131)I and the relative therapeutic efficacy of the 2 radiolabeled species was compared in a human colon cancer xenograft system. METHODS: Nude mice bearing human SW1222 colon carcinoma xenografts were administered escalating activities of (125)I-A33 (9.25-148 MBq) or (131)I-A33 (0.925-18.5 MBq), (125)I- and (131)I-labeled control antibodies, unlabeled antibody, or no antibody. The effects of treatment were assessed using the endpoints of tumor growth delay and cure. RESULTS: Tumor growth delay increased with administered activity for all radiolabeled antibodies. Approximately 4.5 times more activity was required for (125)I-A33 to produce therapeutic effects that were equivalent to those of (131)I-A33. This ratio was approximately 7 for a nonspecific, noninternalizing isotype-matched, radiolabeled control antibody. Unlabeled A33 antibody had no effect on tumor growth. Approximately 10 times more activity of (125)I-A33 produced toxicity similar to that of (131)I-A33, and this ratio fell to approximately 6 for radiolabeled control antibody. CONCLUSION: Treatment with (125)I-A33 resulted in a relative therapeutic gain of approximately 2 compared with (131)I-A33 in this experimental system.     

A simple remote system for the high-level radioiodination of monoclonal antibodies

We have developed a system for the remote control radiolabeling of monoclonal antibodies (MoAb). The system is simple, inexpensive to assemble, and completely disposable. Reagents are pneumatically manipulated into vials containing Iodogen and an anion exchange resin, wherein iodination and separation of bound from free iodine occur, respectively, and sterilized (by filter) into a final vial. Radiolabeling yields of 38.9 +/- 5.5% were consistently produced with 99.3 +/- 0.2% of the isotope bound to the antibody. These results were not significantly different from those obtained in manual iodinations. However, the radiation dose to the hands of the laboratory worker in the remote mode was reduced at least ninefold. The system is presently being introduced for 300 to 400 mCi 131I iodinations of the anti-ovarian cancer MoAb OC 125 in a Phase I radioimmunotherapy trial.     

Radioimmunoscintigraphy of gynecologic tumors with 131I-labeled anti-PLAP monoclonal antibodies

Radioimmunoscintigraphy (RIS) was performed in 20 patients with gynecologic tumors, 14 ovarian, 5 cervical, and one endometrial carcinoma. One murine monoclonal antibody (mab) against placental alkaline phosphatase (H7) was used after radiolabeling with 131I. The labeling procedure yielded antibodies with specific activity varying between 60 and 73 MBq/mg mab. Each patient received 57 to 100 MBq of the preparation. RIS was performed 7 to 35 days later. Patients with ovarian adenocarcinoma had an accumulation of activity on RIS at tumor sites (79%, 11/14) verified by ultrasonography, CT, and clinical examination. A low or absent accumulation of activity was seen in patients with cervical tumors. The patient with an endometrial adenocarcinoma was seen to have an activity accumulation at RIS corresponding to tumor sites determined by ultrasound and/or CT. It is concluded that RIS using monoclonal antibodies against placental alkaline phosphatase can provide information which will supplement that gained from other investigations of patients with ovarian adenocarcinomas.     

Biodistribution and planar gamma camera imaging of (123)I- and (131)I-labeled F(ab')(2) and Fab fragments of monoclonal antibody 14C5 in nude mice bearing an A549 lung tumor

Detection of antigen 14C5, involved in substrate adhesion and highly expressed on the membrane of many carcinomas, including lung cancer, provides important diagnostic information that can influence patient management. The aim of this study was to evaluate the biodistribution and planar gamma camera imaging characteristics of radioiodinated F(ab')(2) and Fab fragments of monoclonal antibody (mAb) 14C5 in tumor-bearing mice. METHODS: F(ab')(2) and Fab 14C5 fragments were radioiodinated using the Iodo-Gen method. In vitro stability, binding specificity and affinity of (125)I-labeled 14C5 fragments were studied in A549 lung carcinoma cells. Biodistribution, blood clearance and tumor-targeting characteristics of (131)I-labeled 14C5 fragments and intact mAb 14C5 were studied in Swiss nu/nu mice bearing A549 lung carcinoma tumors. Planar gamma imaging illustrated the potential use of these (123)I-labeled 14C5 fragments for radioimmunodetection (RID). RESULTS: Saturation binding experiments showed highest affinity for (125)I-labeled F(ab')(2) fragments (K(d)=0.37+/-0.10 nmol/L) and lowest affinity for (125)I-labeled Fab fragments (K(d)=2.25+/-0.44 nmol/L). Blood clearance studies showed that the alpha half-life (t(1/2)alpha) value for Fab, F(ab')(2) and mAb 14C5 was 14.9, 21 and 118 min, respectively. The beta half-life t(1/2)beta value for Fab, F(ab')(2) and mAb 14C5 was 439, 627 and 4067 min, respectively. (131)I-Fab fragments showed highest tumor uptake 3 h after injection (2.4+/-0.8 %ID/g), (131)I-labeled F(ab')(2) showed highest tumor uptake 6 h after injection (4.7+/-0.7 %ID/g) and for (131)I-labeled mAb highest tumor uptake was observed at 24 h (10.7+/-2.3 %ID/g). In planar gamma imaging, both labeled fragments gave better tumor-to-background contrast than (123)I-mAb 14C5. CONCLUSION: Fab and F(ab')(2) fragments derived from intact mAb 14C5 have significant potential for diagnostic and therapeutic applications and may provide new tools in mAb-based radiopharmaceuticals for targeting non-small cell lung cancer.     

Comparison of the distribution and binding of monoclonal antibodies labeled with 131-iodine or 111-indium

The distribution of two monoclonal antibodies with reactivity against human leukemia/lymphoma associated antigens (BA-1 antibody) and carcinoembryonic antigen (202 antibody) when labeled with 131I or 111In was studied in normal Balb/c mice. The BA-1 antibody of the IgM subclass was labeled with 131I by the micro iodine monochloride method at a 12:1 molar ratio and with 111In by the cyclic DTPA anhydride method at a 10:1 molar ratio. In vitro, the 131I-labeled BA-1 antibody bound 35.5% to 10(7) KM-3 leukemic cells while the 111In-labeled BA-1 antibody bound 29.9% to the same number of KM-3 cells. In vivo, the 111In-labeled BA-1 antibody showed a higher accumulation in liver, spleen, and kidney than the 131I-labeled BA-1 antibody. The 202 antibody of the IgG1 subclass was labeled with 131I at a 5:1 molar ratio and with 111In at a 7:1 molar ratio. In vitro, the 131I-labeled 202 antibody bound 30.9%, 27.4%, and 30.0% to 10(7) CO-112, WIDR, and LS-174T colon cancer cells, respectively. The 111In-labeled 202 antibody bound 20.5%, 30.2%, and 33.6%, respectively to the same number of colon cancer cells. In vivo, the 131I-labeled 202 antibody showed a higher tissue to blood ratio in liver, spleen, and kidney than the 111In-labeled 202 antibody. The data indicate that the relative distribution of 131I-labeled versus 111In-labeled monoclonal antibody may depend on the immunoglobulin subclass of the antibody and the molar ratio used in labeling.     

Iodine-131-labeled MAb F(ab')2 fragments are more efficient and less toxic than intact anti-CEA antibodies in radioimmunotherapy of large human colon carcinoma grafted in nude mice

During one week, beginning 18 days after transplantation, nude mice bearing human colon carcinoma ranging from 115 to 943 mm3 (mean 335 mm3) were treated by repeated intravenous injections of either iodine-131-(131I) labeled intact antibodies or 131I-labeled corresponding F(ab')2 fragments of a pool of four monoclonal antibodies (MAbs) directed against distinct epitopes of carcinoembryonic antigen (CEA). Complete tumor remission was observed in 8 of 10 mice after therapy with F(ab')2 and 6 of the animals survived 10 mo in good health. In contrast, after treatment with intact MAbs, tumors relapsed in 7 of 8 mice after remission periods of 1 to 3.5 mo despite the fact that body weight loss and depression of peripheral white blood cells, symptoms of radiation toxicity, and the calculated radiation doses for liver, spleen, bone, and blood were increased or equal in these animals as compared to mice treated with F(ab')2.     

Imaging and treatment of B-cell lymphoma

Ten patients with non-Hodgkin's lymphoma have been evaluated as candidates for experimental radioimmunotherapy and five of those patients have been treated with a single high dose of iodine-131-(131I) labeled anti-pan B-cell antibodies. The evaluation protocol involved collecting biodistribution data by quantitation of gamma camera images and by tumor biopsy from trace labeled doses of antibody, to estimate the relative radiation dose delivered to normal organs and tumor sites. Each patient received up to three escalating mass doses (0.5 mg/kg, 2.5 mg/kg, and 10.0 mg/kg) of radioiodinated antibody for determination of the antibody amount that yielded the most favorable biodistribution for treatment. The millicuries of 131I-labeled to the optimal antibody dose for therapy was selected to deliver 1,000 rads (three patients) or 1,500 rads (two patients) to normal uninvolved organs. Because severe bone marrow toxicity was expected, all patients had their bone marrow cryopreserved prior to entry into the study. This report details the methods and results of quantitative imaging, biodistribution data collection, and absorbed radiation dose estimation in patients with lymphoma receiving high level radioimmunotherapy with 131I-labeled antibodies.     

Intratumoral distribution of radiolabeled antibody and radioimmunotherapy in experimental liver metastases model of nude mouse.

The biodistribution and intratumoral distribution of radiolabeled anticarcinoembryonic antigen (CEA) monoclonal antibody in experimental liver metastases and the therapeutic effect of 131I-labeled anti-CEA antibody on the metastases were studied. METHODS: Three weeks after an intrasplenic injection of human colon cancer cells, mice received an intravenous injection of 125I- or 111In-labeled anti-CEA antibody F33-104. The biodistribution and tumor penetration of radiolabeled antibody were examined by using quantitative autoradiography. To evaluate the therapeutic effect, 5.55, 9.25 or 11.1 MBq (150, 250 or 300 microCi) 131I-labeled F33-104 were injected into groups of mice that had micrometastases smaller than 1 mm. Control groups were injected with phosphate-buffered saline or 131I-labeled control antibody. Mice were killed 3 wk later to determine the size of liver metastases. RESULTS: 1251-labeled F33-104 showed a high accumulation in the liver metastases (percentage of injected dose per gram of metastases [%ID/g] >24%, metastasis-to-liver ratio >9.8, metastasis-to-blood ratio >2.1); however, its accumulation was heterogeneous or peripheral in the nodules more than 1 mm in diameter. When the antibody dose was increased, antibody penetration was improved, but tumor uptake of radioactivity and specificity ratios decreased. In mice with large metastases, radioactivity in the normal tissue was lower than that in mice with small metastases, resulting in higher metastasis-to-background ratios. 111In-labeled antibody showed even higher tumor uptake than 125I-labeled antibody (>51 %ID/g). Metastases formation was suppressed in a dose-dependent manner by 131I-labeled F33-104 injection (5 of 8 mice had no macroscopic tumor after an injection of 5.55 MBq (150 microCi), and all mice had no visible metastasis after an injection of 9.25 or 11.1 MBq [250 or 300 microCi]), whereas tumor progression was seen in the control groups. CONCLUSION: Liver metastases had easy accessibility to the antibody. Micrometastases of less than 0.5 mm in diameter showed homogeneous intratumoral distribution of injected antibody and were successfully treated with 131I-labeled antibody. Very high uptake and satisfactory metastasis-to-liver ratios with 111In-labeled antibody suggest that the use of a radiometal with high beta-energy, such as 90Y or 188Re, is preferable for the successful radioimmunotherapy of metastases larger than 1 mm.     

Dosimetry of 131I-labeled 81C6 monoclonal antibody administered into surgically created resection cavities in patients with malignant brain tumors.

The objective of this study was to perform the dosimetry of 131I-labeled 81C6 monoclonal antibody (MAb) in patients with recurrent malignant brain tumors, treated by direct injections of MAb into surgically created resection cavities (SCRCs). METHODS: Absorbed dose estimates were performed for nine patients. Dosimetry was performed retrospectively using probe counts (during patient isolation) and whole-body and SPECT images thereafter. Absorbed doses were calculated for the SCRC interface and for regions of interest (ROIs) 1 and 2 cm thick, measured from the margins of cavity interface. Also, mean absorbed doses were calculated for normal brain, liver, spleen, thyroid gland, stomach, bone marrow and whole body. The average residence time for the SCRC was 111 h (65-200h). RESULTS: The average absorbed dose per unit injected activity (range) to the SCRC interface and ROIs 1 and 2 cm thick from the cavity interface were 31.9 (7.8-84.2), 1.9 (0.7-3.6) and 1.0 (0.4-1.8) cGy/MBq, respectively. Average absorbed doses per unit administered activity to brain, liver, spleen, thyroid, stomach, bone marrow and whole body were 0.18, 0.03, 0.08, 0.05, 0.02, 0.02 and 0.01 cGy/MBq, respectively. The high absorbed dose delivered to the SCRC interface may have produced an increase in cavity volume independent of tumor progression. CONCLUSION: At the maximum tolerated dose of 3700 MBq 131I-labeled 81C6 MAb, the absorbed doses to the SCRC interface and ROIs of 1 and 2 cm thickness were estimated to be 1180, 71 and 39 Gy, respectively. The estimated average absorbed dose to the brain was 6.5 Gy. There was no neurological toxicity and minimal hematologic toxicity at this maximum tolerated administration level.     

Radiolabeled monoclonal antibodies specific to the extracellular domain of prostate-specific membrane antigen: preclinical studies in nude mice bearing LNCaP human prostate tumor.

Prostate-specific membrane antigen (PSMA), a transmembrane glycoprotein, is highly expressed by virtually all prostate cancers. PSMA is also expressed on the tumor vascular endothelium of virtually all solid carcinomas and sarcomas but not on normal vascular endothelium. PSMA is currently the focus of several diagnostic and therapeutic strategies. We have previously reported on the radiolabeling and in vitro binding properties of monoclonal antibodies (mAbs) (J415, J533, and J591) that recognize and bind with high affinity to the extracellular domain of PSMA (PSMA(ext)). This article reports on the in vivo behavior and tumor uptake of (131)I- and (111)In-labeled antiPSMA(ext) mAbs (J415, J533, and J591) and their potential utility for radioimmunotherapy. METHODS: In nude mice bearing PSMA-positive human LNCaP tumors, the pharmacokinetics, biodistribution, and tumor uptake of these antibodies was compared with (111)In-7E11 mAb, specific to the intracellular domain of PSMA (PSMA(int)). Autoradiographic studies were done to identify intratumoral distribution of radiolabeled mAbs. RESULTS: With (131)I-labeled antibodies, the net tumor retention of radioactivity by day 6 was significantly higher with J415 (15.4% +/- 1.1%) and 7E11 (14.5% +/- 1.7%) than with J591 (9.58% +/- 1.1%). By contrast, the tumor uptake of (111)In-1,4,7,10-tetraazacyclododecane-N,N',N", N"'-tetraacetic acid-labeled J415 and J591 gradually increased with time and was quite similar to that of 7E11. In addition, the blood clearance of (111)In-labeled J415 and J591 antibodies was relatively faster than that of radiolabeled 7E11. As a consequence, the tumor-to-blood ratios with J415 and J591 were higher than that of 7E11. The localization of radiolabeled anti-PSMA(ext) antibodies in PSMA-positive LNCaP tumors was highly specific because the tumor uptake of (131)I-labeled J415 and J591 was more than twice that of a nonspecific antibody. Furthermore, the tumor uptake of (131)I-J591 was almost 20 times higher in PSMA-positive LNCaP tumors than in PSMA-negative PC3 and DU145 tumor xenografts. Autoradiographic studies suggested that 7E11 (anti-PSMA(int)) distinctly favors localization to areas of necrosis whereas J415 and J591 (anti-PSMA(ext)) demonstrated a distinct preferential accumulation in areas of viable tumor. CONCLUSION: These results clearly demonstrate that PSMA-specific internalizing antibodies such as J415 and J591 may be the ideal mAbs for the development of novel therapeutic methods to target the delivery of beta-emitting radionuclides ((131)I, (90)Y, and (177)Lu) for the treatment of PSMA-positive tumors. In addition, because J591 and J415 mAbs are specific to PSMA(ext), thus targeting viable tumor, these immunoconjugates are better candidates for targeted radioimmunotherapy than are antibodies targeting PSMA(int).     

Phase I trial of iodine-131-chimeric B72.3 (human IgG4) in metastatic colorectal cancer.

Twelve patients with metastatic colorectal cancer participated in a Phase I trial of 131I-labeled chimeric B72.3 (human IgG4). Consecutive groups of patients received 18 mCi/m2, 27 mCi/m2 and 36 mCi/m2. No acute side effects related to antibody administration were noted. Bone marrow suppression was the only side effect; it was dose-dependent and correlated with whole-body radiation dose estimates. The lowest dose level produced no marrow suppression, whereas 27 mCi/m2 resulted in Grade 1 and 2 marrow suppression in two of three patients. The maximum tolerated dose was 36 mCi/m2 with all six patients at this dose level having at least Grade 1 and two patients with Grade 3 and 4 marrow suppression. Eight of 12 patients had radioimmune imaging of tumor sites at 5-22 days. Seven patients had an antibody response to initial infusion. On retreatment, whole-body kinetics and imaging were altered for patients with a high anti-ch-B72.3 response. Thus, chimeric B72.3 (IgG4) has limited utility as a means of delivering multiple therapeutic doses of 131I in the majority of patients; alternative strategies including second generation anti-TAG-72 monoclonal antibodies, other radioisotopes and other chimeric human isotypes will need to be pursued.     

Stabilisation of [131I]meta-iodobenzylguanidine at room temperature as organic extract in ethyl acetate/chloroform.

A method of purification of [131I]mIBG (diagnostic dose) by solvent extraction and possibility of its stabilisation by storage in ethyl acetate and chloroform at up to 40 degrees C for not less than 14 days is described. In order to further enhance the utility of this finding, the organic extract was dispensed into vials and dried. Vials containing such dried residue of [131I]mIBG stored at up to 40 degrees C was also found to be stable for at least 10 days. The final product reconstituted as aqueous solution resulted in 90% recovery with 99% radiochemical purity as assessed by previously reported methods, as well as a relatively more rapid paper chromatography method standardised by us. These interesting findings promise simpler and economic packaging besides transport of diagnostic doses of [131I]mIBG.     

Antibody guided diagnosis and therapy of brain gliomas using radiolabeled monoclonal antibodies against epidermal growth factor receptor and placental alkaline phosphatase

Twenty-seven patients with brain glioma were scanned using 123I-labeled monoclonal antibodies against epidermal growth factor receptor (EGFR1) or placental alkaline phosphatase (H17E2). Successful localization was achieved in 18 out of 27 patients. Eleven out of 27 patients were also studied using a nonspecific control antibody (11.4.1) of the same immunoglobulin subclass and observable tumor localization was also achieved in five patients. The specificity of targeting was assessed by comparing images obtained with specific and nonspecific antibodies and by examining tumor and normal tissue biopsies after dual antibody administration. Ten patients with recurrent grade III or IV glioma who showed good localization of radiolabeled antibody were treated with 40-140 mCi of 131I-labeled antibody delivered to the tumor area intravenously (n = 5) or by infusion into the internal carotid artery (n = 5). Six patients showed clinical improvement lasting from 6 mo to 3 yr. One patient continues in remission (3 yr after therapy), but the other five who responded initially relapsed 6-9 mo after therapy and died. No major toxicity was attributable to antibody-guided irradiation. Targeted irradiation by monoclonal antibody may be clinically useful and should be explored further in the treatment of brain gliomas resistant to conventional forms of treatment.     

Potentiation of radioimmunotherapy with response-selective peptide agonist of human C5a.

Physiologic barriers to the delivery of macromolecules to solid tumors are a major obstacle to the clinical success of radioimmunotherapy (RIT). Only a small fraction of the injected dose of the radiolabeled monoclonal antibody (mAb) localizes at the tumor site. This situation worsens as the tumor burden increases. It is hypothesized that improvements to RIT of adenocarcinoma can be realized by inclusion of vasoactive agents, in particular agents able to increase the vascular permeability of tumor capillaries. In these studies, a response-selective peptide agonist of human C5a, GCGYSFKPMPLaR (AP), was used to transiently increase tumor vascular permeability in an effort to improve RIT of solid tumors. METHODS: Athymic mice xenografted with human colorectal adenocarcinoma LS174T were treated intravenously with low doses (9.25 MBq) of 131I-labeled mAb B72.3 in combination with various intravenous doses of AP. The progression of the disease or the loss of >20% body weight was taken as the endpoint. Biodistribution and tumor uptake kinetics were studied in the same tumor-antibody system. RESULTS: The uptake of 125I-B72.3 in LS174T xenografts increased in a dose-dependent manner with an apparent maximal effect between 3 and 6 h after intravenous administration of AP. Augmenting the dose of 9.25 MBq 131I-B72.3 with a single administration of 0.1 mg AP delayed tumor growth nearly 2-fold; the tumor quadrupling time (T(q)) was 14.2 +/- 3.3 d for 131I-B72.3 alone versus 26.0 +/- 3.6 d for 131I-B72.3 plus 0.1 mg AP (P < 0.001). An additional dose of 0.1 mg AP 24 h after 131I-B72.3 further improved the therapeutic outcome (T(q) = 48.5 +/- 7.9 d; P < 0.001) and resulted in several cases of tumor regression. CONCLUSION: The inclusion of agonist peptides of human C5a in the RIT scheme results in improved tumor responses without any manifest side effects.