Cytotoxic activity, tumor accumulation, and tissue distribution of ruthenium-103-labeled bleomycin.

Bleomycin (BLM) was labeled with gamma-emitting 103Ru. Yields of 103Ru-labeled BLM as high as 50.6% were attained. 103Ru-labeled BLM was stable in vitro and the 103ru label was not displaced by large excesses of Cu (II) and Co (II) or Fe (III). Chromatography of the urine following 103Ru-labeled BLM injection indicated no in vivo decomposition. Pharmacokinetic studies in healthy inbred SD and tumor-bearing inbred BUF rats demonstrated tumor accumulations, tissue distributions, and clearance nearly identical with those reported for 3H-labeled BLM. Cytotoxicity studies on a WI-L2 human B-cell line showed that BLM labeled with nonradioactive Ru retained 100% of the activity demonstrated by native BLM. Thus BLM may be labeled with isotopes of Ru to form stable complexes by a simple, rapid reaction without loss of its chemotherapeutic properties or variations in its in vivo distribution. BLM labeled with the proper Ru isotope should prove useful as a gamma-emitting tracer for BLM or a beta-emitting compound capable of providing combination chemotherapy and radiotherapy of tumors.

Stability, characterization, and kinetics of 111In-labeled monoclonal antitumor antibodies in normal animals and nude mouse-human tumor models.

Monoclonal antibodies (MoAbs) against carcinoembryonic antigen were successfully radiolabeled with 111In, and the radiopharmaceutical was characterized in vitro and in normal and tumor-bearing mice. The 111In-MoAb proved to be stable in vitro and in vivo under normal conditions, although instability could be induced in vitro with large quantities of iron-free transferrin. Animal distribution studies with 111In-MoAb demonstrated tumor localization superior to 67Ga and pharmacokinetics that were highly similar to those of endogenously labeled 75Se-MoAb. The 111In-MoAb followed first-order kinetics and fit a two-compartmental model when studied in nude mice bearing human colon tumors known to express carcinoembryonic antigen. Significant quantities of radiolabel appeared in tissues other than tumor, with liver and skin having the highest concentrations. Sufficient tumor/background ratios were formed for scanning purposes. The data indicate that 111In-MoAb may prove to be effective as a radiopharmaceutical for tumor imaging.

Athymic mouse model of a human T-cell tumor.

Because of the large number of different immunoconjugates which can be produced from monoclonal antibody-directed anti-cancer therapy, it would be useful to have in vivo tumor models to compare such preparations. Although historically human leukemias-lymphomas have been difficult to establish in athymic mice we have succeeded in establishing human T-cell tumors from primary MOLT-4 cultures in 290 of 353 animals and have successfully transferred tumors in 42 of 45 animals during ten serial passages. The potential utility of this model for testing immunoconjugates of murine monoclonal antibody T101 have been confirmed by: (a) in all 148 tumors sampled including all passaged tumors the human T-cell antigen, T65, was expressed in a manner identical to that of cultured cells; (b) 111In-T101 was concentrated preferentially in the tumor; and (c) T101 injected by both the i.p. and i.v. routes bound to tumor and induced antigenic modulation to the same extent as that observed previously in vitro and in human studies.

Optimal handling of dimercaptosuccinic acid for quantitative renal scanning.

Methods of optimizing quantitative renal imaging with Tc-99m dimercaptosuccinic acid (DSMA) were investigated. Rats were injected with DMSA (one kit per rat) and sacrificed at 0.5, 2.0, and 24 hr after injection. Fifty percent of the injected dose localized in the kidneys at 0.5, 2, and 24 hr after injection while background activity peaked at 0.5 hr and then declined to give substantially higher kidney-to-background ratios at 24 hr. Delayed scanning should increase the accuracy of clinical studies in patients with low kidney-to-background ratios at 1-2 hr. After injection of DMSA, 1 ml of air was introduced into the reaction vials and incubated 20 min. Kidney uptake decreased from 50 to 40% and liver uptake increased from 7.5 to 17%. If multiple doses must be drawn from a single vial, air should not be introduced, and the doses should be drawn together and administered immediately to minimize radiopharmaceutical deterioration.

A rapid method for the labeling of albumin microspheres with In-113 and In-111: concise communication.

A rapid and simple method for preparing microspheres labeled with In-113m or In-111 is described. The procedure requires 10 min and gives labeling yields approaching 100%. Biodistribution studies in rats, mice, and dogs show the product to be biologically stable, with approximately 90% of the injected dose localized in the lung at 1 hr postinjection. The convenient production of In-113m or In-111 albumin microspheres provides an alternate radiopharmaceutical to complement or substitute for Tc-99m microspheres in lung perfusion imaging and other circulation studies.

Distribution of 67-Ga following intravenous administration: effects of disodium edetate therapy.

Following intravenous administration of 67Ga-citrate to normal and abscessed rats, the colon content of 67Ga decreased with increasing oral doses of Na2EDTA. The effects observed, however, were not thought to be of potential clinical value.

99mTc-thiomalic acid complex: a nonstannous chelate for renal scanning.

Thiomalic acid (monomercaptosuccinic acid) has been labeled with 99mTc without the use of an intermediary reducing agent. Tissue distribution studies in rats following the injection of 99mTc-tagged thiomalic acid (99mTc-TMA) showed 40-48% of the injected dose in the kidneys. Renal incorporation of this compound was influenced by various parameters such a pH, quantity of thiomalic acid, heating time, and the preparation-injection interval. Scintigrams of a midline kidney slice showed that the 99mTc activity concentrated mainly in the renal cortex. As a proposed renal-imaging agent 99mTc-TMA compared favorably with 99mTc-Sn-dimercaptosuccinate and 99mTc-penicillamine regarding the precent incorporation into the kidney and was superior in this respect to 99mTc-Sn-glucoheptonate and 99mTc-Sn-diethylenetriamine pentaacetic acid. The 99mTc-TMA was also shown to be highly stable through 24 hr. The reagent can be made available in kit form and is easily combined with 99mTc in two steps. Finally, the absence of stannous ion in the 99mTc-TMA complex should avoid the problem of interference with other procedures involving pertechnetate 99mTco4- as the imaging agent.

Tumor model studies of 131I-tetracycline and other compounds.

Iodine-131-tetracycline (131I-TET) was prepared by allowing tetracycline hydrochloride to react with radioiodide in acidic methanol (labeling efficiency greater than 85%). This preparation was found to be stable at--4 degrees C for at least 72 hr. Some minimal in vivo breakdown did occur. The 131I-TET, 67Ga, and several 99mTc compounds were studied in a rat hepatoma model. The incorporation of the radiopharmaceuticals into the tumor occurred rapidly, with peak levels at 0.5 and 24 hr after injection for 131I-TET and 67Ga, respectively. The clearnace of the radiopharmaceutical from nonviable tumor was slower than for viable tumor, and by 72 hr after injection the greatest concentration of radioactivity appeared in the nonviable fraction. All normal tissues showed faster clearance than did tumor tissue, regardless of viability. Decreasing the quantity of 131I-TET injected increased the percent of uptake in the nonviable tumor tissue but had no effect on the viable tumor uptake. Of the 99mTc compounds studied, the phosphates developed the highest tumor-to-background ratios. Unfortunately these ratios were not as high as those achieved for 67Ga or 131I-TET.

Tissue distribution studies with radioactive manganese: a potential agent for myocardial imaging.

Manganese, a trace metal, is known to localize in mitochondria. Because mitochondria are abundant in heart muscle, the possible utility of radioactive manganese as a myocardial imaging agent was examined in 25 rats and six dogs. Myocardial uptake of Mn-54 in rats was found to exceed that of thallium-201; myocardium-to-blood ratios averaged 306:1 versus 48:1 for Tl-201. In the dog, uptake of Mn-54 by ischemic myocardium was reduced by 17-75% compared with normal myocardium. Thus, radioactive manganese appears promising as an intravenous myocardial imaging agent, and might be useful in studying the function of myocardial mitochondria by external imaging.

In vivo kinetics of radiolabeled monoclonal anti-CEA antibodies in animal models.

Studies were performed to determine the effect of the radiolabel and circulating carcinoembryonic antigen (CEA) on the pharmacodynamics of monoclonal anti-CEA antibodies (MoAbs). The studies were performed in normal BALB/c mice and in nude mice bearing human colon tumors. Three different tumors were used, each of which produced CEA levels characteristic of that particular tumor's secretory rate. The CEJ-326 MoAb labeled with either 111In or 125I was used in all studies. Circulating CEA induced the removal of 125I and 111In MoAbs from the vascular compartment. Liver concentrations of 111In increased and 125I levels decreased as the CEA secretory rate of the tumor rose. This indicates that circulating CEA complexes form in the vascular compartment which, in an animal model, are removed by the liver and spleen. This results in decreased tumor uptake of the labeled MoAb. The iodinated MoAb complexes are dehalogenated while the 111In is retained by the liver. This dehalogenation may account for the relatively low liver activity observed in radioimmunoimaging with intact radioiodinated anti-CEA MoAbs, provided the CEA complexes are similarly removed from the vascular compartment by the human liver.

Distribution of radiolabeled human and mouse monoclonal IgM antibodies in murine models.

The distribution and kinetics of six human and one murine monoclonal IgM antibodies (MoAb) were studied in BALB/c mice. Labeling was with 111In, 75Se, and 125I. The monomers and pentamers of certain MoAbs were studied. Human distribution studies were also performed. The serum containing [111In]MoAb was obtained from one of the patients 24 hr after administration and injected into mice which were then killed and assayed for 111In distribution. In general, the [75Se] and [111In]MoAbs had distribution and kinetic patterns that were similar while the 125I-labeled MoAbs dehalogenated after 4 hr. Monomers and pentamers had highly similar distributions suggesting that the distribution of IgMs may be based on factors other than molecular size. The murine IgM showed a somewhat different distribution in mice than did human IgMs. Serum from the patient containing [111In]MoAb had a distribution in mice similar to that of the patient with high liver and gastrointestinal uptake. The human imaging indicates that it is possible to target tumor with human IgM MoAbs, but significant problems remain in regard to their clinical use.

Tumor size: effect on monoclonal antibody uptake in tumor models.

Studies were performed to determine the effect of tumor size on the incorporation of radiolabeled monoclonal antitumor antibodies (MoAbs) into human tumors growing in nude mice. The colon tumors ranged in size from 0.03-1.6 g, the melanoma from 0.1 to 6.7 g, and the lymphoma from 0.06 to 10.2 g. Indium-111 was primarily used as the radiolabel, however, both 125I and 111In were used as tracers for the MoAb in one experiment. The per g radiopharmaceutical uptake by tumors was inversely proportional to tumor size when tumor specific MoAb was administered. This finding was independent of the radiolabel and was demonstrable when the mice bore two tumors of differing size. When the MoAb was not specific for the tumor, the data were less well defined and a statistically significant correlation with size did not occur. These data are strong evidence for a decrease in per g uptake of labeled tumor specific antibodies as tumors increase in size.

The effect of certain variables on the tumor and tissue distribution of tracers: VI. False-carrier effect, Part III, Fe.

Previous work has shown that Fe3+, when administered in the proper dose and time sequence, increases the tumor uptake of gallium-67 (67Ga) while decreasing its uptake by normal tissues. The purpose of this series of experiments was to examine further the postulate that the false carrier effect is mediated at the cellular as well as the vascular level, determine the lowest concentration of ionic Fe3+ that will induce near maximum tumor/background ratios (T/Bkg), determine the best technique for its administration, and decide whether Benadryl and dexamethasone could be used to offset side effects of the Fe3+ without altering tumor and tissue kinetics. Fe3+ altered tissue levels of 67Ga prior to changes in the blood. The threshold for initiation of the false-carrier effect varied to some extent from one organ to another. Tumor uptake of 67Ga was either enhanced or unaltered at 4 hours after injection; 0.3 mg Fe3+/kg administered 0.5 hour before and 2 hours after the 67Ga enhanced 4-hour T/Bkg by a factor of about ten. Twenty-four-hour ratios were improved (to a lesser extent than 4-hour), but decreased concentrations of 67Ga occurred in the tumor. Dexamethasone and Benadryl did not alter the outcome of the experiment. This technique should be useful for imaging with gallium-68 and the PET camera.

The effect of certain variables on the tumor and tissue distribution of tracers. II. Carrier effect: rapidity of onset and concentrations necessary for initiation and maximum response.

The smallest quantity of carrier Ga and Mn necessary to initiate and maximize a carrier effect was studied in the Morris 7777 rat hepatoma model. The quantity needed for a maximum response did not appear to adversely effect the rats. Not all tissues were equally affected at the same plasma concentrations. If carrier Ga was administered 2 hours following 67Ga injection and the rats sacrificed 30 minutes later, a dramatic change occurred in background activity, which was more pronounced in healthy than malignant tissues. Early viable and nonviable tumor/background ratios were improved by this technique. The data suggest that the use of carrier Ga and Mn might improve early lesion/background ratios in patients. This could be of use if tumor imaging were undertaken with 68Ga or 52mMn with positron detector systems.

Distributions of 137Cs, 201Tl, 203Hg, 203Pb and 57Co in a rat hepatoma model. Comparison with 67Ga.

The distribution of carrier-free 203Pb-acetate, 203HgCl2, 57 CoCl2, 137CsCl and 201TlCl was investigated in rats bearing thigh-implanted Morris 7777 hepatomas. Viable and nonviable tumor tissue was collected in order to determine the relative affinities of the radiopharmaceuticals for these tissues. The animals were sacrificed at 4, 24, 48, 72 and 96 hrs following intravenous injection. Washout of the radioisotope from the viable tumor tissue was rapid, the maximum concentration being reached on or before 4 hrs following injection. In contrast, residual activity within the nonviable tumor tissue decreased much more slowly and in some cases even increased with time. Viable tumor-to-muscle and nonviable tumor-to-muscle ratios for 203Pb, 203Hg and 57Co were comparable to the analogous ratios reported for 67Ga. However, none of these isotopes approached 67Ga as a potential tumor imaging agent because the large ratios were the result of low muscle uptake rather than high tumor uptake. Blood clearance of 67Ga was faster than any of the five cations, while viable and nonviable tumor affinity for 67Ga was greater than for any of the radiopharmaceuticals studied.

The effect of certain variables on the tumor and tissue distribution of tracers. IV. False carriers: ferric citrate.

The intravenous administration of Fe+3 -citrate (1.6 mg/kg body weight) was demonstrated to alter the concentration of carrier-free 67Ga and 54Mn in malignant and healthy tissues of the rat, Morris 7777 hepatoma model. When the Fe+3 was injected 2 hours before, simultaneously with, or 2 hours after 67Ga (and the rats sacrificed 4 hours after injection), the 67Ga in most normal tissues decreased, and the viable tumor concentrations increased by 135, 24, and 47%, respectively. Twenty-four hours after a simultaneous administration of Fe+3 and 67Ga, egress of 67Ga from the tumor was much less than from the healthy tissues. These changes resulted in significant improvements in viable tumor to background ratios, especially at 4 hours. These changes induced in the distribution of the two tracers by Fe+3 indicate that some kinetic characteristics are shared. This is discussed in the light of their response to carrier Ga and Mn. The use of Fe+3 shows promise as a means of improving tumor/background ratios for 68Ga and 52mMn, two short-lived positron emitters that can be used with positron scanners. Gallium-67 imaging may also be improved by these techniques. The Fe+3 increases excretion of 67Ga from the animal, and this could result in a lower radiation dose to a patient.

The effect of certain variables on the tumor and tissue distribution of tracers V: false carrier effect, II, Fe.

The effect of Fe3+ citrate on carrier-free 67Ga and 59Fe kinetics was studied in a Buffalo rat-Morris 7777 hepatoma model. Two mg Fe3+ citrate/Kg were administered intravenously in a variety of time sequences, prior to and following the tracers, and the rats were killed at 4 or 24 hours. 67Ga concentrations could be increased in tumor and decreased in most normal tissues. Administering Fe3+ both one half hour before and 2 hours after the tracers produced 67Ga values equivalent to 72-hour carrier-free values after 4 hours, while simultaneously decreasing gut secretion and increasing urinary excretion. The 59Fe and 67Ga kinetics suggested that events at both vascular and cellular levels were responsible for these changes. This study demonstrated the potential utility of Fe3+ citrate for improving both conventional 67Ga and positron (68Ga) imaging of tumors.

The effect of certain variables on the tumor and tissue distribution of tracers. Part 1; Carrier.

Buffalo rats bearing thigh-implanted strain-7777 Morris hepatomas were used as a model for studying the effect of carrier material on the body distribution, tumor uptake, excretion, and tumor-to-background ratios of 67Ga and 54Mn. An effort was also made to observe the changes in 67Ga and 54Mn concentrations induced by carrier in viable tumor and skeletal muscle, relative to their interstitial fluid space. This value is referred to as the Tissue Distribution Index. Carrier manipulation resulted in striking changes in the distribution of the two ions from the carrier-free state. The data also indicated a difference in the pharmacodynamics of 67Ga and 54Mn in malignant and healthy tissues which could be of importance to nuclear medicine and oncology.

Detection of occult tumor using indium 111-labeled anticarcinoembryonic antigen antibodies.

Even with the advancement of radiologic techniques, metastatic cancers can still be difficult to detect. In this study, 48 patients suspected of having occult metastases were studied by radioimmunodetection following the administration of 92.5 to 181.3 MBq of indium 111-labeled monoclonal anticarcinoembryonic antigen antibody. All but seven patients were thought to have metastatic colorectal carcinoma. In the majority of cases, physical examinations and computed tomographic scans had failed to detect a lesion. At least one lesion that was later proved to exist was detected in 34 of the 50 studies performed on these patients. Seven of eight patients with normal radioimmunodetection scans remain free of disease. One hundred one sites were detected overall; 60 were considered true-positive sites and 27 false-positive sites. Fourteen sites remained in question. Nineteen false-negative sites occurred. Radioimmunoimaging appears valuable for the detection of occult cancer where standard, noninterventional techniques have failed to detect the suspected disease.

Quantitation of skeletal-muscle necrosis in a model compartment syndrome.

UNLABELLED: Skeletal-muscle necrosis was evaluated in previously pressurized canine compartments using technetium-99m stannous pyrophosphate and classic histological criteria. Intracompartmental necrosis was quantitated in the anterolateral muscle compartment of each dog by uptake of 99mTc stannous pyrophosphate using the contralateral anterolateral compartment as an internal control. Representative specimens of muscle were sampled in experimental and control legs of each dog and were analyzed by qualitative histological techniques. Muscle necrosis was assessed in compartments forty-eight hours after pressurization to levels of ten to 120 millimeters of mercury for eight hours in thirty-seven dogs. In another dog, neither anterolateral compartment was pressurized so that both compartments acted as control muscle. The results in these experiments identify a threshold pressure level (thirty millimeters of mercury) and duration (eight hours) at which significant muscle necrosis occurs at normal blood pressure. Our findings imply that a quantitative relationship exists between incorporation of 99mTc stannous pyrophosphate and the level of intracompartmental pressure. This uptake technique, however, is not suitable for diagnosing compartment syndrome in patients with a threatened compartment syndrome. We suggest that intracompartmental pressure measurements by the wick-catheter technique, in conjunction with clinical findings, offer the best means for diagnosing compartment syndrome. CLINICAL RELEVANCE: Significant muscle necrosis associated with an impending compartment syndrome occurs at a threshold intracompartmental pressure of thirty millimeters of mercury after eight hours. Since time variables are often unknown in suspected compartment syndromes, fasciotomy is recommended when intracompartmental pressure exceeds thirty millimeters of mercury in a patient with normal blood pressure. The use of this threshold pressure level as an indication for fasciotomy requires a device for measuring intracompartmental pressure such as the wick catheter.