Biological distribution of 99mTc-labeled YIGSR and IKVAV laminin peptides in rodents: 99mTc-IKVAV peptide localizes to the lung.

Two laminin-derived peptides containing either YIGSR or IKVAV (single amino acid code) sequences were radiolabeled with 99mTc and their biological distribution evaluated in rodents. Both 99mTc-peptides cleared rapidly from the circulation though the kidney, and to a lesser extent, through the liver. 99mTc-YIGSR peptide did not accumulate in any organ examined in normal, tumored, and emphysemic mice. The 99mTc-IKVAV peptide localized within 10 min to the lung of normal animals, resulting in lung-to-blood ratios of approximately 23:1. The 99mTc-IKVAV peptide localized to lung after submicron filtration and after intraperitoneal injection, suggesting that particulates do not major role in localization. Pre-incubation of 99mTc-IKVAV peptide in whole blood decreased lung localization, suggesting that margination of radiolabeled cells does not play a major role in the lung localization. When 99mTc-IKVAV was injected into mice with tumored lungs (melanoma), the lung uptake was markedly increased (up to 20% injected dose higher than control lungs) at all time points examined (10, 30, and 120 min). When 99mTc-IKVAV was injected into mice with genetic emphysema, the lung uptake was markedly decreased at all time points. The localization of the 99mTc-IKVAV-containing peptide to the lung is consistent with a receptor-based mechanism.

Assessment of peripheral vascular disease in patients with diabetes. Two case studies.

This report proposes that perfusion scanning in combination with arteriography be included in the diagnostic work-up of the diabetic patient who, because of peripheral vascular complications, is a candidate for surgery. Two cases are reported which illustrate the extremes of the findings: abnormal arteriogram-normal scan indicating large-vessel disease without significant small-vessel involvement. It is suggested that these patients are candidates for vascular reconstruction. The other extreme is the normal arteriogram-abnormal scan indicating small-vessels disease without significant large-vessel involvement. It is apparent that these patients are not candidates for vascular reconstruction.

Quality control test for immunoreactivity of radiolabeled antibody.

A rapid method for the measurement of the immunoreactive fraction of a radiolabeled monoclonal antibody or antibody fragment has been developed. This may be used as a quality control test prior to patient administration of the radiolabeled antibody preparation. The test employs solid phase antigens and the assay is conducted under conditions of antigen excess. Assay parameters have been evaluated and a standardized procedure has been developed. The assay has been compared to a standard extrapolation method and found to give approximately the same result. The test has been used on four different radiolabeled antibodies currently in clinical trials in patients with colorectal cancer. Mean immunoreactive fractions for these radiolabeled antibodies ranged from 35 to 65% and the variability of the immunoreactive fraction ranged from 140 to 240% for different antibodies. We conclude that the quality, defined as the immunoreactive fraction, of radiolabeled antibodies is both low and highly variable, indicating the need for a quality control test of these radiopharmaceuticals in the clinic prior to patient administration.

Affinity thin-layer chromatography test of the immunoreactive fraction of radiolabeled antibodies.

A simple, rapid and self-contained system for assaying the immunoreactive fraction of radiolabeled antibodies was developed using affinity thin-layer chromatography (ATLC). ATLC combines use of solid-phase-bound antigen and conventional TLC. The technique is an improvement over existing means of measuring immunoreactive fraction (bead-type or cell-type assays) in that it has neither wash steps nor centrifugation steps, yet provides results essentially identical to those obtained with the more time-consuming assays. ATLC is accomplished using chromatography strips that are coated with antigen material in a discrete region near the origin. The antigen-coated strips are then blocked in serum, air-dried and stored. For use, radiolabeled antibody is spotted at the origin, and the strip is developed using a buffered solvent. Immunoreactive antibody binds to the antigen at or near the origin, while radioactivity not associated with immunoreactive antibody migrates with the solvent front. Antigen-negative strips (serum-blocked only) are used to measure "nonspecific" binding. The ATLC development time is about 16 min, and the results can be obtained in about 30 min. The assay described in this report uses antigens from colon tumor and is suitable for use with B72.3 and other colon cancer-reactive antibodies.

Exercise lowers thyroid radioiodine uptake: concise communication.

The effect of exercise upon the uptake of radioiodine by the thyroid was examined in both rats and humans. Rats that exercised intermittently on a mechanical wheel for a period of 20 days had significantly lower uptake values (p < 0.0001) than sedentary controls. Human volunteers that ran at least ten miles/week had a lower mean 24-hr uptake value (8.0 +/- 2.8%) than nonexercising subjects (14.3 +/- 5.1%, p < 0.01). Other thyroid function studies (thyroxine, triiodothyronine, triiodothyronine resin uptake, thyroid-stimulating hormone) did not differ significantly between the exercising and nonexercising groups. These studies suggest that exercise significantly alters thyroid iodine economy.

Technetium-99m labeling of murine monoclonal antibody fragments.

F(ab')2 fragments of several murine monoclonal antibodies have been labeled with 99mTc by a direct, pretinning method. The fragments were incubated with stannous ions overnight to split disulfide groups--a process which converts dimeric F(ab')2 to monomeric fragments. The pretinned fragments were then either directly labeled with 99mTc, frozen for subsequent labeling, or lyophilized to make kits for 99mTc-labeling at some later date. The 99mTc-labeled fragments were shown to be stable against transchelation when challenged with ethylenediaminetetraacetic acid, retained immunoreactivity, and were capable of binding to human tumor xenografts in nude mice.

Radioimmunoimaging of experimental thrombi in dogs using technetium-99m-labeled monoclonal antibody fragments reactive with human platelets.

Monoclonal antibody 50H.19, which reacts with human platelets, was converted to fragments, pretinned, and made into kits for subsequent radiolabeling with 99mTc. The antibody, which cross-reacts with dog platelets, was used to evaluate in vitro binding to blood clots and in vivo in experimental thrombi in dogs. After radiolabeling, 97.4 +/- 6.4% of the 99mTc was antibody-associated. The preparations retained immunoreactivity, as determined by: binding studies using whole blood and determining the ratio of cell-to-plasma radioactivity (ratios of 57.6-61.2) and binding of the antibody to clots (clot/serum ratios were 57.2-74.6%). Approximately 50% of the radioactivity was cleared from the blood in 3-6 min and 18-24% was excreted in urine within 3 hr. Experimental thrombi in dogs could be visualized consistently within 2-3 hr postinjection in peripheral veins and arteries, pulmonary arteries, and the right ventricle. In addition, damage to blood vessel intima without visible thrombi could also be detected. This method has the following advantages: short and simple pre-imaging preparation, and rapid visualization of thrombi with no need for blood-pool subtraction or delayed imaging.

Effects of a 99mTc-labeled murine immunoglobulin M antibody to CD15 antigens on human granulocyte membranes in healthy volunteers.

UNLABELLED: An injectible, 99mTc-labeled, murine immunoglobulin M antibody to stage-specific embryonic antigen-1 has been developed that can localize infections by binding to CD15 glycoproteins expressed on the cell membranes of human granulocytes in vivo after systemic administration. The purpose of this study was to measure its clinical effects on healthy people. METHODS: Multiple blood samples were aspirated before and after the intravenous administration of about 125 microg antibody labeled with approximately 370 MBq (10.0 mCi) 99mTc in 10 healthy human volunteers. Complete blood cell counts were performed at each time point. Whole-body scans were acquired contemporaneously with a dual-head gamma camera. The fraction of the administered dose at each time point was quantified in 18 regions of interest. Statistical analyses included paired t tests. RESULTS: Administration was associated with a transient decrease in the concentration of red and white blood cells in the whole blood. The effect always began within 3 min of administration. Its nadir was always reached 15-20 min after administration. There was full recovery with mild overcompensation in about an hour. The hematocrit dropped by a mean of 3.8% (P<0.002), whereas the total white blood cell count fell 44.0%+/-3.1% (P<0.001). The effect was most pronounced on the number of circulating granulocytes, which fell from 5.7+/-2.1 to 3.2+/-1.3x10(3)/microL blood. The drop paralleled a decrease in the percentage of whole blood radioactivity bound to the white blood cell membranes, which peaked at 50.4%+/-7.6% at 3 min after injection and then fell to 26.1%+/-9.3% over the next 30+/-13.4 min before recovering to 40.7%+/-8.2% at 2 h. Image analysis showed that the effect was temporally associated with an increase in the amount of radioactivity within the liver and the spleen. Recovery was associated with a decrease in hepatosplenic radioactivity. No evidence of cell destruction or agglutination could be detected. CONCLUSION: This study confirmed that administration of this radiolabeled antibody is associated with a transient decrease in the number of circulating granulocytes. However, there also seems to be a secondary hemodilutionlike effect on all blood components that has not been reported previously. The effect appears to be clinically silent and very short-lived.

Clinical radiopharmacy: principles and practices.

On the average, radiopharmacists spend about 17.2% of their time in clinical activities if their practice setting is in an institution, and about 8.5% of their time if their practice setting is in a centralized nuclear pharmacy. A recent survey of radiopharmacists was conducted to determine: (1) the percentage of time they spend engaged in selected activities, and (2) the specific clinical activities in which they are involved. A few radiopharmacists spend as much as 50% of their time in clinical activities, but most spend only 5% to 20% of their time. Some of the clinical activities involve direct interactions with patients, such as explaining the reasons for administering the radioactive material or actually administering the dose. Other clinical activities are indirect, such as reviewing charts before or after studies and making recommendations to other health care professionals. About half of the pharmacists surveyed see a need for increasing their clinical activities. The need to maximize the time involved in providing pharmaceutical care is discussed and several patient-care activities/responsibilities are proposed.

COTA (colon-ovarian tumor antigen). An immunohistochemical study.

A goat anti-serum was prepared against mucinous ovarian cyst fluid and absorbed with normal colon and a variety of normal tissues until the only residual immunoreactivity was directed against colon cancer and ovarian tumor mucin. The set of antigenic determinants defined by this anti-serum has been called COTA, standing for colon-ovarian-tumor-antigen. This highly absorbed anti-serum (anti-COTA) was used for immunohistochemical staining of 42 different tissues in parallel with staining with a goat anti-CEA, which was also highly absorbed. The results suggest that COTA is a highly sensitive and specific antigen for colon carcinoma and may have potential for the early detection of malignant changes predictive of cancer of the colon.

Perfusion of ischemic ulcers of the extremity: a prognostic indicator of healing.

Forty patients with an ischemic ulcer of the lower extremity had peripheral vascular perfusion studies, performed with intra-arterial injections of aggregated technetium Tc 99m serum albumin microspheres (15mu to 30mu in diameter), in an attempt to develop an objective prognostic criterion for healing. The association between ulcer healing and the presence or absence of diabetes mellitus, palpable peripheral pulses, and patent trifurcation vessels on the arteriogram was reviewed and no association was noted. When, however, there was a relative hyperemia of the ulcer bed in comparison to the adjacent tissue of at least 3.5:1, as determined by counting the amount of radioactivity per unit area, 86% of patients went on to heal their ulcers. In those without this degree of hyperemia, only 11% were healed with conservative nonsurgical management. The results have shown that relative hyperemia of the ulcer bed is a clinically useful prognostic indicator in the patient with ischemic ulcer disease.

Quantitation of radiolabeled antibody binding to cells by thin-layer chromatography.

Thin layer chromatography (TLC) was used to monitor binding of radiolabeled antibodies to cells. Labeled antibodies were reacted with cells and aliquots chromatographed on serum-blocked, ITLC strips. The cell-antibody complexes remain at the origin and unbound antibody migrates with the solvent front. The antibody binding was estimated from the ratio of radioactivity at the origin compared to the total applied. Separations are completed in about 10 min. This method does not use centrifugation or wash steps, and provides an inexpensive and self-contained system to evaluate radioligand binding. Cell binding assay results using this method are approximately the same as those obtained using bead- or cell-type assays.

Abscess scintigraphy with 99mTc-human immunoglobulin (IgG) using a one-step labeling method.

It was shown earlier that non-specific human gamma globulin (IgG) labeled with 111In can be used as an agent for abscess localization. We describe experimental results with 99mTc-IgG in animals bearing abscesses and tumors using a one-step labeling method with 99mTc. We studied this compound in several animal models: mice bearing turpentine abscesses and subcutaneously transplanted sarcomas, in rats with turpentine or E. coli abscesses and intracerebrally implanted gliomas and in rabbits with E. coli or turpentine abscesses. Blood clearance was studied in dogs. It was found that the absolute concentration of 111In-IgG in abscess and tumor was higher than that of 99mTc-IgG. However, the abscess-to-tumor ratio was higher for 99mTc-IgG. The 99mTc-IgG images were of high quality and abscesses could be detected as early as 30 min post-injection (p.i.). It appears that 99mTc-IgG has many potential advantages over 111In-IgG because of better physical properties of 99mTc, simpler preparation, lower cost and greater availability and the possibility of using higher 99mTc doses.

Clinical aspects of local and regional tumor therapy with 188Re-RC-160.

Somatostatin-receptors have been found to be overexpressed in a variety of neuro-endocrine and epithelial cancers. While the introduction of a long-acting somatostatin-analogue, octreotide, exerted mainly anti-cancer activity in neuro-endocrine tumors, no convincing results have been demonstrated in other cancers. RC-160, another somatostatin-analogue has been selected because of its high receptor affinity and its anti-cancer activity. 188Re is a generator produced radionuclide with favourable gamma and beta-emission, allowing diagnostic and therapeutic application. The results of in vivo biodistribution and therapeutic outcome following systemic, intralesional and intracavitary application in animal studies employing 188Re-RC-160 are summarized. Safety considerations, dosimetry estimates and applicable indications are outlined. The clinical impacts of this radiopharmaceutical in cancer management are discussed.

Pre-clinical experience with Re-188-RC-160, a radiolabeled somatostatin analog for use in peptide-targeted radiotherapy.

The clinical potential of radiolabeled peptides such as octreotide and VIP has been widely established for tumor localization. Radiotherapy based on the tumor binding potential of the peptides and the radiotoxic effects of beta- or a-emitting radionuclides is an extension of such applications. Rhenium-188 (T1/2 16.9 hr, beta-max 2.1 MeV) coupled to the analogue RC-160 has been used to establish the feasibility of treating tumors with radiolabeled peptides, and our experience with this approach is summarized. In three different experimental tumor models (human prostate, mammary gland, and small cell lung carcinomas) in nude mice, treatment resulted in significant reduction or elimination of tumor burden. Two routes of administration were used: intra-lesional injection (prostate carcinoma) and intra-cavity injection (mammary and SCLC). Re-188-labeled negative control peptides bound to tumor cells to a low extent and did not exhibit therapeutic benefit. RC-160 by itself did not result in therapeutic benefit. Tumors which did not bind Re-188-RC-160 did not evidence a therapeutic benefit. Uncoupled Re-188 (control) was rapidly excreted via the urinary bladder and did not accumulate in either tumors or normal tissues even following direct injection. Instant radiolabeling kits containing 200 micrograms of RC-160 were labeled with < 3000 MBq of Re-188 in 30 minutes with no need for subsequent purification. These studies establish the conceptual feasibility of targeted radiotherapy based on the local or regional administration of radiolabeled peptides.

Comparison of expanded PTFE and vein grafts in lower extremity arterial reconstructions.

Polytetrafluoroethylene (PTFE) bypasses were used in a series of arterial reconstructions to the popliteal artery (45) and to arteries below that level (11). These were performed in high-risk situations in patients who lacked a suitable saphenous vein. Vein bypasses were performed in a comparable series of high-risk situations in patients having a suitable autologous saphenous vein (45 to the level of the popliteal artery and 11 to an artery below that level). PTFE patency rates at 4-14 months were 43 to 45 (96%) for the femoro-popliteal reconstructions (with a limb salvage rate of 39 to 45 or 87%) and 5 of 11 (45%) for the distal bypasses. Saphenous vein bypass patency rates at 8-14 months were 39 of 45 (87%) for the femoropopliteal reconstructions (with a limb salvage rate of 36 of 45 or 80%) and 5 of 11 (45%) for the distal bypasses. These results justify continued use of PTFE grafts in patients without saphenous veins who require lower extremity arterial reconstructions for limb salvage. The exact place of PTFE grafts in arterial reconstructive surgery of the lower extremity definition based on longer periods of observation.

Duplex ultrasonographic insonation and visualization of intracerebral arteries.

Until now, clinical, noninvasive interrogation of intracranial vessels has consisted only of insonation via transcranial Doppler. Such devices have utilized a 2.0 MHz, continuous wave probe with Doppler spectral waveform display. Clincial aapplication of these techniques has required precise location of cranial "windows" and has been hampered by the extreme anatomic variability of both cranial bony structures and intracerebral arteries. The lack of simultaneous intracranial arterial visualization has limited the clinical pplicability of transcranial Doppler technology. Recently, the authors have utilized a 2.25 MHz curved phased array probe with a pulsed Doppler to image and insonate simultaneously the intracerebral arteries. Colorflow imaging of both near- and far-field regions is the necessary first step for vessel localization and identification. Once this is accomplished, the image of each artery in turn is amplified and gray scale tuning is employed to permit direct visualization of the arterial walls and lumen. Pulsed Doppler waveform analysis is perfomred simultaneously and along the entire visible artery length. In this manner the arteries of both right and left hemispheres are examined in detail. We have found the duplex technique to be superior to the use of Doppler alone in the examination on intracerebral vessels. The ability to visualize and insonate simultaneously eliminates the uncertainty caused by anatomic variation. These advantages, long applied to the evaluation of peripheral vessels, are now available for use in the diagnosis of intracranial arterial disease.

Comparison of duplex ultrasonography and ascending contrast venography in the diagnosis of venous thrombosis.

The application of duplex ultrasonography to the diagnosis of venous thrombosis requires validation by comparison of the duplex findings with the results of ascending contrast venography. In this study, 2534 veins were examined by both methods with contrast venography results serving as the standard for comparison. In this setting, duplex ultrasonography proved to be 100% sensitive and 99% specific for venous thrombosis. Duplex ultrasonography is as reliable as venography in the diagnosis of venous thrombosis and has no associated risks or known complication. In addition, duplex ultrasonography provides information regarding pathologic anatomy that is comparable to the detail provided by high-quality venography. The authors conclude that duplex ultrasonography should be the diagnostic method of choice for evaluating patients with suspected venous thrombosis.

Production of monoclonal antibody SP-21 to colon-ovarian tumor antigen, COTA.

An IgG monoclonal antibody, SP-21, directed against colon-ovarian tumor antigen, COTA, is reported. The antibody had no reactivity with CEA, normal colonic mucin, CSAp, ABO blood group antigens, or with normal human lung, liver, spleen, kidney, plasma and saliva in studies using the enzyme-linked immunoassay method (ELISA). Immunoperoxidase staining of colon, lung, kidney, and prostate cancer tissues and benign and inflammatory colon disease tissues revealed a specificity identical to that of the polyclonal (goat) anti-COTA antibodies.

Targeting peptides for pleural cavity tumor radiotherapy: specificity and dosimetry of Re-188-RC-160.

Re-188-RC-160, a radiolabeled somatostatin analogue, is being explored for its potential as a local/regionally administered radiotherapeutic agent targeting somatostatin receptor-positive tumors. In studies in vitro and in vivo, Re-188-RC-160 was found to bind to somatostatin receptor-positive cells (NCI-H69, human small cell lung carcinoma), but not to binding-negative cells (Raji, Burkitt's lymphoma). The comparative binding of Re-188-labeled RC-160 [cyclic NH2-(D)-Phe-Cys-Try-(D)-Trp-Lys-Val-Cys-Trp-NH2] or CTOP [cyclic NH2-(D)-Phe-Cys-Try-(D)-Trp-Orn-Thr-Pen-Thr-ol], a mu-opioid receptor antagonist used a negative control compound, was also determined in vitro and in vivo using NCI-H69 cells as targets. Re-188-RC-160 demonstrated higher net binding in vitro and in vivo compared to Re-188-RC-CTOP, and in vitro its binding could be reduced by excess unlabeled RC-160 whereas binding of Re-188-CTOP could not be reduced. Using another somatostatin receptor-positive human tumor line, ZR-75-1, a substantial amount of the cell-bound Re-188-RC-160 was found to be internalized. Does estimates for Re-188-RC-160 in animals containing NCI-H69 tumors indicated that with three serial injections therapeutic doses greater than 60 Gy can be achieved.