Phase I trial on sms-D70 somatostatin analogue in advanced prostate and renal cell cancer.

Plasma concentrations and tolerability of a novel somatostatin analogue sms-D70 were studied in patients with metastatic hormone-resistant prostate cancer (HRPC) or metastatic renal cell cancer. To overcome the limitations of the octapeptides having affinity only to somatostatin receptor subtypes 2 and 5, HRPC expressing mainly somatostatin receptors 1 and 4, a somatostatin derivative based on the natural somatostatin having affinity to all five somatostatin receptor subtypes, was developed. The in vivo stability of this dextran-conjugated derivative, somatostatin-D70, was confirmed previously in animal studies, and the nanomolar "panaffinity" has been shown in in vitro receptor binding studies on cell lines transfected with the somatostatin receptor genes. Sms-D70 was given with subcutaneous injection once a week at dose levels of 5, 10, 20, 35, and 50 mg. For pharmacokinetic studies, sms-D70 was labeled with 131I. Fourteen patients were treated, of whom 10 had prostate and 4 renal cell cancer. The kinetic data revealed high stability with a long half-life in the blood. The drug was well tolerated, and no grade 4 (WHO) toxicity was observed. The maximal tolerated dose could not be established due to the lack of dose-limiting toxicities. Objective PSA responses were not recorded in these heavily treated patients, but subjective stabilization of pain was observed and urinary symptoms were alleviated in four patients. Three patients with metastatic HRPC received 5-10-mg intravenous injections of sms-D70 once weekly for 4-14 months on a compassionate use basis. In all cases, serum PSA values decreased more than 50% from the pretreatment level, but these results are difficult to interpret due to concomitant treatments given to these patients. In conclusion, sms-D70 was well tolerated in the treatment of metastatic prostate and renal cell cancer, but no responses were found in these heavily treated patients.

Technetium-99m labelling of glycosylated somatostatin-14.

This study presents a technetium-99m labelling method based on organometallic chemistry. It describes the simple mixing of a 99mTc(I)-carbonyl compound [99mTc(OH2)3(CO)3]+ with a histidine-tagged somatostatin-dextran (SMS-Dx-His) conjugate. Somatostatin and histidine was coupled to periodate activated dextran. The linkage was stabilised by reductive amination. The conjugate was then radiolabelled with 99mTc by using the 99mTc(CO)3 core. The labelling efficiency was 65-80% and the radiochemical purity > 95%. In the in vitro cysteine challenge, the result showed that 25% of the radiolabel was released after 1 h incubation at 37 degrees C (cysteine-conjugate at 1000:1 molar ratio). The radiolabelled SMS-Dx-His showed similar HPLC profile as the unlabelled conjugate. This labelling method, employing non reducing conditions, is useful for the labelling of peptides containing disulphide bonds. It should be possible to be used also for labelling with rhenium-188 for therapeutic applications.

Biodistribution, blood half-life, and receptor binding of a somatostatin-dextran conjugate.

Derivatives of somatostatin (sms) are attracting increasing interest as part of the treatment of several cancer diseases expressing sms receptors (srs). Radiolabeled sms analogs can additionally be used for systemic radiotherapy and for diagnostic investigations. Glycosylated sms-14 (sms-dextran70) was characterized regarding in vitro srs binding, biodistribution, and blood half-life in mice. Rat brain cortex membranes (expressing srs 2) were used for the srs binding study. Tyr3-Octreotide was used as positive control. The binding data were analyzed by competition curve analysis. In the biodistribution study, the Bolton-Hunter reagent was used for the radioiodination of sms-dextran70. Organs and blood were collected at different time-points and the percentage of the injected dose per gram of tissue (%ID/g) was calculated. The conjugate was administered subcutaneously (sc). The sms-dextran70 showed high srs binding affinity (i.e., in the same nanomolar range as the reference ligand Tyr3-octreotide (IC50 approximately 2.5 nM). The blood half-life was approx 27 h after reaching maximum blood concentration 24 h postinjection. Because of the molecular weight of the conjugate (i.e., approx 75,000) being above the kidney threshold for dextran (i.e., 50,000), the digestion and excretion is assumed to be mainly through the hepatobiliary system. Increased uptake was seen in the adrenals, which are receptor-positive organs. Some accumulation was seen in the stomach, indicating certain deiodination of the conjugate label. The sms-dextran70 showed promising properties and its clinical relevance is currently being evaluated in clinical phase I-II studies.

Intravesical administration of EGF- dextran conjugates in patients with superficial bladder cancer.

OBJECTIVES: Overexpression of the epidermal growth factor receptor (EGFR) has been reported in bladder cancer and is a potential target for therapy with radionuclides. In this study, we investigated the binding of EGF-dextran-(99m)Tc to the EGFR. The aim of this study was to determine if intravesically administered EGF-dextran conjungate selectively accumulated in the tumor tissue and to correlate the uptake to tumor characteristics. METHODS: Eight patients received the conjugate intravesically for about 30 min followed by bladder irrigation and then transurethral resection. Radioactivity of the biopsy specimens from normal urothelium and tumor areas was measured in a gamma counter. RESULTS: Five patients received EGF-dextran-(99m)Tc, three received dextran-(99m)Tc and one received only (99m)Tc. The 5 patients who received the complete conjugate had a mean ratio of radioactivity between tumor and normal urothelium of 664:1 (range: 2.4-1,710). The dextran-(99m)Tc showed a slightly increased ratio and (99m)Tc did not bind at all. CONCLUSION: The results are encouraging and further studies are warranted to investigate if EGF-dextran could be effective as intravesical therapy, either conjugated with cystostatic drugs or labeled with suitable radionuclides.

Radiolabeling of dextran with rhenium-188.

This study describes a method for the radiolabeling of dextran with rhenium-188 (188Re). In nuclear oncology 188Re is very useful for therapeutic applications. Its nuclear characteristics allow radiotherapy and in situ monitoring of tumor uptake as well as dosimetry calculations. Consequently new compounds with this radiolabel are of general interest. Dextran was oxidized with sodium periodate yielding reactive aldehyde groups and subsequently reacted with cysteine. The linkage was stabilized by reducing the Schiff bases with sodium cyanoborohydride. The conjugate was then radiolabeled with 188Re by using 188Re-gluconate as the transchelator, labeling the free thiols. Synthesis and radiolabeling were done in the absence of oxygen. The labeling efficiency was 60-70% and the radiochemical purity > 95%. The in vitro stability study, using "cysteine challenge" demonstrated that 50% of the radiolabel was transcomplexed to the 100 mM cysteine solution (after 1 h incubation at 37 degrees C). However, at physiologic conditions and presence of an antioxidant good stability was achieved. The 188Re labeled dextran presented in this study provides a template with therapeutic and diagnostic potential in nuclear oncology, either alone for local treatment or as a backbone in a tumor specific conjugate for systemic treatment.

Ion exchange tumor targeting: a new approach.

Connective tissues are distinguished by the types, concentrations, and organizations of material in the extracellular matrix. Many physiological functions are determined largely by the nature and organization of the extracellular components. The components are characterized by their content and distribution of charged, mostly anionic groups. The distinct roles played by the charges are sometimes modeled by analogy to the transport theory of ion exchange resins. The intent of this study was to investigate whether the properties of the tumor matrix could be used for selective, charge-dependent accumulation of charge-modified dextran. Ten patients with diagnosed superficial urinary bladder carcinoma were included in the study. They received intravesical instillations of technetium-99m-labeled charge-modified dextran derivatives (approximately 0.1-1 mg; approximately 50 MBq in saline; 30-min incubation). After treatment and resection, samples were taken from normal and diseased tissue. The result clearly demonstrated a charge-dependent difference in the quotient of radioactive uptake in tumor tissue: normal tissue. Instillations of cationic dextran yielded a high quotient, up to 3000. Normal tissue had background activity. Anionic dextran yielded a low quotient, 1.8-2, with increased background (i.e. uptake in normal tissue). Neutral dextran gave a quotient of up to 90. No radioactivity could be detected in blood. The tumors in this study apparently displayed cation-exchanging properties. We will continue this investigation and determine whether this is a general property of bladder carcinomas and whether other carcinomas display ion exchange properties. If this is the case, the finding could have important implications for the local treatment of several cancers.

The potential of radiolabeled EGF-dextran conjugates in the treatment of urinary bladder carcinoma.

BACKGROUND: Muscle-invasive urothelial carcinoma of the urinary bladder has a poor prognosis in spite of available therapies. These tumors frequently overexpress the epidermal growth factor receptor (EGFr), a possible target for therapeutic conjugates. The aim of this study was to construct an EGF-carbohydrate conjugate that would be potentially useful for both local (i.e., intravesical) and systemic radiotherapy. METHODS: EGF was coupled to periodate activated dextran by reductive amination. Receptor binding tests in vitro were conducted, using the human urothelial carcinoma cell line RT4 and the human malignant glioma cell line U-343-MG as a positive control. In the in vivo experiments, nude mice with subcutaneously grown xenografts of the RT4 tumor were injected intravenously with technetium-99m-labeled EGF conjugates. Samples of organs, blood, and tumors were collected after 24 hours. The radioactive uptake was calculated as a percentage of the dose per gram of tissue. RESULTS: The specific binding in the in vitro experiment was 90-95%. The binding was similar in both cell lines. There was a positive uptake in the tumors in the in vivo experiment, with tumor-to-blood ratios from 2:1 to 6:1. The uptake level in the kidneys was similar to that in the tumors (i.e., approximately 0.05% dose per gram of tissue). The other organs had a lower uptake compared with the tumor uptake. CONCLUSIONS: The results indicate that radiolabeled EGF-dextran has the potential to become a tool for local treatment of recurrent bladder carcinoma. With appropriate modifications, it should be possible to use this conjugate for systemic radiotherapy as well.

99mTc-dextran-antibody conjugates. Labelling procedures.

Dextran forms stable chelates with 99mTc, a radionuclide with ideal properties for planar scintigraphic and tomographic imaging. This study investigates some of the factors of importance to the formation of 99mTc-dextran. The complex was used for the technetium labelling of a monoclonal antibody. Two radiolabelling methods were studied: direct dextran labelling with the reductant dissolved in HCI and labelling via a weak 'transfer' chelator (tartaric acid) with the reductant dissolved in ethanol. Different conditions during the labelling reaction were studied. Finally, dextran was coupled to a monoclonal anticytokeratin antibody and the conjugate was subsequently radiolabelled with 99mTc. Gel filtration (GFR) and thin layer chromatography (TLC) were compared as methods for estimation of the labelling efficiency. When using 10-500 microM of ligand, 5-100 microM SnCl2 with 10-500 MBq of technetium at pH 7 incubated for 10-15 min, the radiolabelling seemed optimal (70-75% labelling efficiency). It was found that 100 microM tartaric acid used as a weak intermediate chelator with SnCl2 dissolved in ethanol improved the reproducibility of the labelling. The labelling efficiency was not affected by either the presence of oxygen or the addition of an oxygen scavenger during the labelling incubation. In general, TLC showed higher labelling efficiencies than GFR, indicating inadequate separation of the different moieties.