Initial clinical evaluation of iodine-125-labeled chimeric 17-1A for metastatic colon cancer.

UNLABELLED: The internalizing properties of murine antibody 17-1A in human colon cancer cells make it attractive as a carrier for radionuclides with short range emissions such as 125I. Murine 17-1A IgG2a antibody, which reacts against human gastrointestinal cancers, has been chimerized by joining its variable region with human IgG1 k constant region. A pilot clinical trial of increasing doses of 125I-chimeric 17-1A in patients with metastatic colorectal cancer has been conducted. METHODS: Patients were treated in groups of 2-4; 2 patients at Hahnemann University and 26 at the University of Alabama at Birmingham. Groups 1-5 received single administrations with 125I doses of 20, 40, 60, 80 or 100 mCi. Subsequent groups received therapeutic doses of 150, 200 or 250 mCi, with the dose subdivided into infusing of 50 or 100 mCi at 4-day intervals. All treatments were delivered in an outpatient setting using radiation precautions. Labeling at 10 mCi/mg antibody was performed on the day of treatment. RESULTS: Pharmocokinetics of circulating antibody was studied for initial patients, showing alpha T 1/2 of 17-27 hr and beta T 1/2 of 100-190 hr. Whole-body T 1/2 of radioactivity was determined by measuring urinary excretion or gamma emissions. Treatment was well tolerated without significant acute or late side effects. No significant bone marrow suppression or other dose-limiting toxicities were noted over this dose range. No objective responses were noted. CONCLUSION: These results show that high-dose outpatient radioimmunotherapy with an 125I-labeled internalizing antibody can be achieved without significant patient toxicity or radiation hazard.

Indium-111-labeled anti-EGFr-425 scintigraphy in the detection of malignant gliomas.

BACKGROUND: The monoclonal antibody anti-epidermal growth factor receptor (EGFr) antibody-425, against the epidermal growth factor receptor, has the potential to bind specifically to gliomas and not normal brain tissue. A prospective study was conducted (1986-1988) to evaluate the use of Indium-111 (111In)-labeled anti-EGFr-425 in the localization of gliomas before radioimmunotherapy with Iodine-125 (125I)-labeled anti-EGFr-425. METHODS: Twenty-eight patients with intracranial neoplasms were injected intravenously with an average dose of 2.2 mCi 111In-labeled anti-EGFr-425. Planar and single-photon emission computed tomography scans were performed after 48 and 72 hours. Control studies also were performed in two cases with 111In-labeled Co 17-1A (an antibody to colorectal cancer) and in one case with unlabeled 111In chloride. RESULTS: The immunoscintigraphic findings were generally in good agreement with computerized tomographic findings. The definitive diagnosis was established by biopsy findings: 23 gliomas (1 Grade I, 5 Grade II, 6 Grade III, and 11 Grade IV), 1 meningioma, and 4 metastatic lesions. The localization of gliomas with 111In-labeled anti-EGF-425 had a sensitivity of 0.96, a specificity of 0.60 and an accuracy of 0.90. CONCLUSION: Immunoscintigraphy with 111-In labeled anti-EGFr-425 can be useful in the management of malignant gliomas, especially before radioimmunotherapy with 125I-labeled anti-EGFr-425.

Brain uptake of thallium-201 from the cerebrospinal fluid compartment.

The present report elucidates the movement of 201Tl through the cerebrospinal fluid compartment and its subsequent uptake by normal brain. Autoradiographic studies of rat brain, after stereotaxic 201Tl injection into either the lateral or fourth ventricle, reveal that 201Tl moves freely through the cerebrospinal and extracellular fluid compartments. Subsequent to lateral ventricular injection, central thalamic and specific hypothalamic nuclei are heavily labeled. Densely labeled mesencephalic nuclei include the periaqueductal grey and oculomotor nuclear complex. Labeling of giant cells within the vestibular complex is suggestive of neuronal uptake. Major fiber tracts are devoid of label confirming that 201Tl uptake does not occur in white matter. Most labeling following fourth ventricle injection occurs within the caudal medulla and cervical spinal grey. Our findings suggest that 201Tl uptake by normal brain from the cerebrospinal fluid occurs as a function of thallium concentration and neuronal activity.

Comparisons between two monoclonal antibodies that bind to the same antigen but have differing affinities: uptake kinetics and 125I-antibody therapy efficacy in multicell spheroids.

It has been predicted that low affinity antibodies (Abs) should penetrate into tumors more readily than high affinity Abs. However, the absolute uptake and residence time of a high affinity Ab may be better. It is, therefore, not clear whether a high affinity Ab would have a therapeutic advantage. This is particularly relevant with 125I radioimmunotherapy, where targeting of every cell is important. This study compared the uptake kinetics and toxicity in multicell spheroids of two murine monoclonal Abs labeled with 125I. 17-1A was produced by immunization with a human colon cancer cell line and has an affinity of 5.15 x 10(7) M-1. 323/A3 was produced by immunization with a human breast cancer cell line and has an affinity of 1.87 x 10(9) M-1. Binding of both Abs to LS174T spheroids was similar at 4 degrees C, but binding of 17-1A was 8-10-fold less than that of 323/A3 at 37 degrees C. Despite this difference, the toxicity of 125I-17-1A in spheroids after 7 days of incubation was similar to that of 125I-323/A3. Autoradiography showed that 17-1A penetrated the spheroids much more deeply and evenly than did 323/A3. It appears that much of the radiation dose to spheroids treated with 125I-323/A3 was wasted because of the uneven Ab distribution. This study demonstrates the potential advantage of using Abs of lower affinity for 125I radioimmunotherapy, because of their more even distribution. It also suggests that a large number of binding sites per cell may be a disadvantage if more 125I is bound than is necessary to kill the cell, because this may slow Ab penetration.

Malignant astrocytomas treated with iodine-125 labeled monoclonal antibody 425 against epidermal growth factor receptor: a phase II trial.

Twenty-five patients with primary presentation of malignant astrocytoma, astrocytoma with anaplastic foci, and glioblastoma multiforme were treated with surgical resection and definitive radiation therapy followed by intravenous or intra-arterial administration of Iodine-125 labeled monoclonal antibody-425, which binds specifically to human epidermal growth factor receptor. The patients presented with primary untreated disease, positive contrast enhanced computed tomography scans of the brain, and compatible clinical symptoms. In this Phase II clinical trial, the patients had surgical debulking or biopsy followed by definitively administered external beam radiation therapy and one or multiple doses (35 to 90 mCi per infusion) of radiolabeled antibody. The total cumulative doses ranged from 40 to 224 mCi. The administrations of the radiolabeled antibody were performed in most cases 4-6 weeks following completion of the primary surgery and radiation therapy. Ten patients had astrocytoma with anaplastic foci and 15 had glioblastoma multiforme. No significant life-threatening toxicities were observed during this trial. At 1 year 60% of the patients with astrocytoma with anaplastic foci or glioblastoma multiforme are alive. The median survival for both groups was 15.6 months.

A novel heterobifunctional linker for formyl to thiol coupling.

A maleimide hydrazide has been synthesized as a heterobifunctional cross-linking agent for thiol to formyl coupling. This linker has been applied to the coupling of the monoclonal antibody 17-1A, or an Fab' derived therefrom, to polyaldehyde dextran onto which the antineoplastic agent ellipticine has been attached. High binding avidities for the unshed antigen on the SW1116 colorectal tumor cell are retained in these drug-dextran-linker-antibody conjugates.

Misonidazole conjugates of the colorectal tumor associated monoclonal antibody 17-1A.

The radiation sensitizer misonidazole has been linked to the monoclonal antibody 17-1A which recognizes a nonshed antigen of a human gastrointestinal tumor. Linkage was accomplished through a hemisuccinate of misonidazole attached by a mixed anhydride coupling and gave a conjugate whose plasma half-life (for drug cleavage) was ca. 70 h. The degree of substitution on the antibody could be precisely regulated by varying the reactant ratios. The binding avidities of the resulting conjugates to the SW1116 colorectal tumor cells decrease logarithmically with increasing drug load. Four to six misonidazoles per antibody represented the optimum drug loading on this system. Enzymatic cleavage of the conjugate-drug union took place at both the ester and the amide linkages with the former scission predominating.

Iodine125 labeled anti-epidermal growth factor receptor-425 in the treatment of malignant astrocytomas. A pilot study.

In 1986, a pilot Phase I/II project was initiated using Iodine-125 labeled anti-epidermal growth factor receptor-425 in the treatment of patients with recurrent glioblastoma multiforme of the brain. The monoclonal antibody was administered intra-arterially by the internal carotid arterial system or the vertebral arterial system depending upon the blood supply to the tumor. The treatment program was repeated at intervals for two or three times. Demonstrated was the intense localization of the monoclonal antibody in the brain tumor prior to therapy using Indium-111 labeled anti-epidermal growth factor receptor-425. This localization was demonstrated prior to any therapy as well as after failure from primary radiation therapy with or without concomitant chemotherapy. To date, 15 patients have been treated following recurrence of their glioma (1/15 metastatic adenocarcinoma) with the monoclonal antibody labeled with Iodine-125. Of the 15 patients, there has been one surgically documented complete response, two partial responders, and five patients with stable disease. The results indicate the potential activity of this radiolabeled monoclonal antibody and have prompted continued accession of patients into a Phase II study as a part of the primary treatment regimen (surgery, radiation therapy with or without chemotherapy) followed by administration of the Iodine-125 labeled anti-epidermal growth factor receptor-425.

Selective chromosomal damage and cytotoxicity of 125I-labeled monoclonal antibody 17-1a in human cancer cells.

A monoclonal antibody, 17-1a, which reacts with antigen expressed in human colon cancers was radiolabeled in high specific activity with 125I. The combination of the antibody and this radionuclide was observed to elicit specific cellular damage after being internalized into cells of the SW1116 human colon cancer cell line. The degree of internalization was quantitatively measured and found to increase over time to 49% after a 48-h incubation period. During this period, significant chromosome aberrations were observed in the SW1116 cell line due to the Auger electrons of 125I. This damage was not observed using Na125I, a nonimmunoreactive radiolabeled antibody, or cells which did not contain the requisite antigen. The number of chromosomal aberrations increased with increasing radioactive concentration of 125I-17-1a. The nuclear damage resulted in specific cellular cytotoxicity and decreased cell survival of SW1116 cells exposed to various concentrations of 125I-17-1a.

Radiosensitizer conjugation to the carcinoma 19-9 monoclonal antibody.

Misonidazole was covalently conjugated (3-68 mol drug/mol antibody) to 19-9 monoclonal antibody directed against a colorectal carcinoma tumor-associated antigen as a method for targeting radiosensitizing agents. This attachment was accomplished by the mixed anhydride method using the hemisuccinate derivative of misonidazole. Evaluation of conjugates in vitro shows a loss of antibody binding activity with increasing loading levels; however, significant binding activity is retained even at relatively high sensitizer/antibody ratios. This observation was consistent in three binding assays: a competitive radioimmunoassay; an enzyme immunoassay; and an affinity column assay. From these studies, it was concluded that the optimal loading factor for misonidazole-antibody conjugates, when it is used for immunochemotherapy lies between 8 and 15. In vitro release studies indicated that conjugates are hydrolytically stable (t1/2 = 4 days) under physiological conditions.

Damage in canine hearts following defibrillator shocks.

Experiments were performed to investigate possible differences in potential myocardial cell damage following the use of two clinically available difibrillators. One had a damped sine wave (DSW), and the other a truncated exponential waveform (TEW). The latter, therefore, had a lower peak current and voltage. After pilot studies to determine damage potential, an energy content of 10 Joules per kg was selected with three transthoracic shocks at 30 second intervals, delivered with paddles, by both defibrillators. Thirteen dogs were shocked with DSW (10 sacrificed at 24 hours and three at 72 hours). Eleven dogs were shocked with TEW (eight sacrificed at 24 hours and three at 72 hours). Three were used as untreated controls. Observations included ECG monitoring, technetium99m pyrophosphate (Tc-PyP) uptake, and gross and microscopic observations including electron microscopy. This report will concentrate on the morphologic changes. Eleven of 13 dogs shocked with DSW developed ventricular tachycardia, transient heart block, Tc-PyP uptake and myocardial cell death with acute necrosis, and aberrant contraction patterns associated with cell death. Progression of the lesions occurred from 24 to 72 hours. Only two (one at 24 hours and one at 72 hours) of the dogs shocked with TEW showed microscopic foci of necrosis. One was a chance finding (24 hours); the other was associated with an overlying "yellow streak". In neither case were arrhythmias or Tc-PyP uptake observed. The results indicate that in dogs at equivalent energy levels, TEW caused significantly less myocardial damage than DSW.

Design, preparation, and biodistribution of a technetium-99m triaminedithiol complex to assess regional cerebral blood flow.

A new ligand (N-piperidinylethyl-DADT, 5) has been prepared which forms two complexes with 99mTc when stannous chloride is used as a reducing agent for [99mTc] pertechnetate. Biodistribution studies of one of the complexes in mice showed that 2.2% of the injected dose of the tracer was in the brain at 5 min postintravenous injection with 0.53% of the dose remaining in the brain at 30 min postinjection. Brain-to-blood ratios at these times were 5.3 and 3.0, respectively. Biodistribution studies of the other complex showed similar behavior with a slightly lower initial uptake by and faster clearance from the brain. Imaging studies of the more promising of the two complexes were conducted in a monkey and a baboon. In both cases, rapid uptake of the tracer in the brain was observed and clear brain images were obtained. Time-activity curves showed peak uptake in the brain at approximately 5 to 7 min postintravenous injection followed by a plateau of about 11 min. The half-lives for clearance of the tracer from the brains of the monkey and baboon were found to be 63 and 58 min, respectively. These results suggest that this tracer may be useful for brain imaging in humans.

Delineation of a transplanted malignant melanoma with indium-111-labeled porphyrin.

An indium complex of a cationic, water-soluble, synthetic porphyrin, tetra-4-N-methylpyridyl porphyrin tosylate, has been prepared and evaluated for its selective localization in tumors in Syrian Golden hamsters with flank transplanted malignant melanoma (Fortner MMI). Tumor-to-blood ratios of the compound increase from 42:1 at 6 hr postinjection, at which time activity levels in the tumor exceed those in all other tissues, to 162.1 at 72 hr. Images in one hamster at three time periods clearly delineated the tumor from nontarget background and also demarcated the viable and necrotic zones within the tumor itself.