Microdosing in early lead discovery.

Microdosing provides a tool to enhance drug development by initiating human studies prior to Phase I studies. The purpose is to assist in the go versus no-go decision-making process and to eliminate early ineffective compounds from the drug pipeline. Selection of multiple potential leads can be performed at the clinical stage instead of in preclinical studies. The microdosing approach can be easily used for a molecularly targeted potential drug compound with a known mechanism of action. It provides useful data regarding accessibility and biodistribution that can be used in many estimations benefiting the development of the molecule. In addition, steady state and genetic investigations are becoming possible. Microdosing has a sparing effect on timelines and costs, however, the real importance is not yet known because, although it is known to be widely performed, only a few original reports have been published.

Endoglin is a suitable target for efficient imaging of solid tumors: in vivo evidence in a canine mammary carcinoma model.

Increasing evidence suggests that endoglin (CD105) is a new powerful marker of neovascularization in solid malignancies; thus, using breast cancer as a model, we investigated whether targeting of CD105 by monoclonal antibody (mAb) MAEND3 can be used for in vivo imaging of solid tumors. Immunohistochemistry and flow cytometry identified differential expression of CD105 on breast cancer and endothelial cells; in fact, neoplastic cells were weakly and rarely stained by mAb MAEND3, which in contrast, strongly and invariably stained blood vessel endothelia within the breast adenocarcinomas investigated and cultured endothelial cells. Moreover, in contrast to CD31, which currently represents the reference marker to assess angiogenetic activity, CD105 expression was highest in semiconfluent and actively proliferating endothelial cells, and it progressively decreased as cells reached tight confluency and low [3H]thymidine uptake. i.v. administration of 18 MBq of 125I-labeled mAb MAEND3 efficiently imaged spontaneous mammary adenocarcinomas in two dogs; the uptake of radiolabeled mAb was rapid and intense because tumor: background ratios of 8.2:1 and 9.3:1 were reached 8 h after mAb administration, in the absence of immediate and/or long-term clinical side effects. Altogether, our present data suggest that targeting of CD105 on tumor-associated blood vessels may represent a new strategy for in vivo imaging of solid malignancies, regardless of their histological origin.

High response rate with a lower dose of paclitaxel in combination with cisplatin in heavily pretreated patients with advanced breast carcinoma.

BACKGROUND: Paclitaxel has been found to be efficacious in the treatment of breast carcinoma either when administered alone or in combination with other anticancer agents. Synergistic interaction between paclitaxel and cisplatin has been demonstrated in vitro. METHODS: Thirty-two patients with breast carcinoma that was resistant to anthracyclines and to several other antineoplastic agents were selected to receive 80 mg/m(2) of paclitaxel on Day 1 and 80 mg/m(2) of cisplatin on Day 2 with a 3-week interval between the courses. RESULTS: High response rates were observed, with 3 complete responses (9.4%) and 13 partial responses (40.6%) reported. Furthermore, the disease remained stable in 7 patients (21.9%) and progressed in only 9 patients (28.1%). CONCLUSIONS: The results show that high response rates can be achieved with the combination of paclitaxel and cisplatin, even in heavily pretreated breast carcinoma patients. The combination of paclitaxel plus cisplatin was found to be highly efficacious and well tolerated.

The use of TL detectors in dosimetry of systemic radiation therapy.

A method for determining absorbed doses to organs in systemic radiation therapy (SRT) is evaluated. The method, based on thermoluminescent (TL) dosimeters placed on the patient's skin, was validated and justified through a phantom study showing that the difference between measured (TL dosimeters in the phantom) and derived (TL method) values is within 10%. Six radioimmunotherapy (RIT) patients with widespread intraperitoneal pseudomyxoma were also studied. In dose evaluations, special emphasis was on kidneys. In addition to the TL method, the absorbed doses to kidneys were calculated using MIRD formalism and a point dose kernel technique. We conclude that in SRT the described TL method can be used to estimate the absorbed doses to those critical organs near the body surface within 50% (1 SD).

Imaging of olfactory neuroblastoma--an analysis of 17 cases.

A total of 17 histologically confirmed olfactory neuroblastomas treated at Helsinki University Central Hospital between 1962 and 1996 were reviewed retrospectively. The tumors displayed a variety of imaging characteristics and aggressiveness. Imaging evolved from plain X-rays at the beginning of the study period to CT and MRI during the latter part of the study. CT provided the best information about the tumor and its local invasion especially into surrounding bony structures. MRI allowed an estimate of tumor spread into surrounding soft-tissue areas, such as the anterior cranial fossa and the retromaxillary space. However, signal intensity characteristics were not specific for olfactory neuroblastomas. Bone scintigraphy and MIBG scan allowed us to detect distant metastases. Olfactory neuroblastoma is an aggressive malignancy and the prognosis is poor in most cases, as shown by the short survival rates (average 45.3 months) in our study group. The tumor can be detected, delineated and its characteristics suspected by modern radiology. Definite diagnosis is based on histopathology. This study proposes general imaging strategies for detecting this disease.

Gene therapy using antisense oligodeoxynucleotides labeled with Auger-emitting radionuclides.

Antisense oligomers may be used as a vehicle for carrying a radiation source into a specific location inside a tumor cell. The effects of radioactive-labeled oligodeoxynucleotides (ODNs) may have both direct antisense inhibition and radiation. Thus far, the use of radioactive ODNs has been limited mostly to clinical biokinetic studies. Therapeutic possibilities remain unknown if the basic question of the optimal source of radiation is unanswered. We have shown previously that oligonucleotide therapy can be effective theoretically with the internally labeled ODN phosphorothioates 32P, 33P, and 35S. Here, we expand the selection of radionuclides; we calculated in vivo subcellular tissue distribution for ODN phosphorothioates using the decay characteristics of several beta- and Auger-emitting radionuclides: 32P, 35S, 51Cr, 67Ga, 111In, (1114m)In, 123I, 125I, 131I, and 201Tl. The absorbed nuclear doses of these radiolabeled oligonucleotides were estimated in different cellular dimensions using the subcellular biodistribution data for two oligonucleotides (ISIS 2105 and ISIS 2922). Our results indicate that Auger-emitter isotopes do not give higher absorbed cell nuclear doses than the isotopes suitable for internal labeling of ODN phosphorothioates. However, the biological difference is difficult to estimate. The best isotope for subcellular targeting was 35S, which gives the smallest variation of nuclear dose in the different cell dimensions we studied (nuclear diameter, 6-16 microm; cellular diameter, 12-20 microm). Therefore, we conclude that in oligonucleotide radiotherapy, nuclear targets should be treated with short-range beta-emitters (35S or 33P) that are suitable for the internal labeling of oligonucleotides unless the relative biological effectiveness of Auger-emitters could be remarkably improved. Dual labeling with 32P and 35S may provide therapeutic benefits when treating smaller and larger targets simultaneously. Further in vivo development, especially with 33P and 35S labels for ODNs, is strongly indicated.

The effect of three dimensional activity distribution on the dose planning of radioimmunotherapy for patients with advanced intraperitoneal pseudomyxoma.

BACKGROUND: Six patients with histologically proven peritoneal carcinomatous pseudomyxomas were treated with radioimmunotherapy. METHODS: All the patients received a tracer dose of iodine-131 (131I) labeled B72.3 anti-TAG-72 monoclonal antibody (MoAb) to test the in vivo affinity. After informed consent was obtained the therapeutic dose (>3.7 gigabecquerels [GBq], 100 mCi) of the 131I labeled B72.3 anti-TAG-72 MoAb was infused within 60 minutes intraperitoneally using 2 catheters on both sides of the abdomen. The patients were imaged with single photon emission computed tomography (SPECT) at 3, 10, and 24 days after the therapeutic infusion. Treatment-planning software has been developed in which functional information obtained from SPECT is integrated with anatomic information obtained from computed tomography (CT). The activity distribution from SPECT images is converted to absorbed dose distributions using a point source kernel convolution dose calculation. The absorbed dose calculation requires a radionuclide specific dose kernel. The activity map is divided into equally sized source voxels from which the distribution is calculated for the target voxels that cover the patient volume. The resulting three dimensional (3D) absorbed dose distribution is viewed as isodose contours superimposed on the CT images or as 3D isodose surfaces. RESULTS: The measured activity distribution shows that the cumulated activity and biologic half-life vary in the patient's body. The developed planning system provides a method for calculating patient specific absorbed dose distributions. CONCLUSIONS: The variation of biologic clearance indicates that a 3D dose calculation method incorporating measured activity distributions is needed to quantify absorbed dose distribution.

Imaging of Hand-Schüller-Christian syndrome by a monoclonal antibody.

PURPOSE: Multifocal eosinophilic granuloma (Hand-Schüller-Christian syndrome) consists of localized lesions of histiocytosis X originating from proliferating Langerhans' cells involving male children and young adults. MATERIALS: The authors present the first case of multifocal eosinophilic granuloma that was imaged by Tc-99m monoclonal antibody BW250/183 recognizing NCA-95 antigen on granulocytes. A 30-year-old man had a diagnosis of histiocytosis X for 16 years. RESULTS: All active known lesions were visualized. The previous radiotherapy fields were also accurately demonstrated as areas with no antibody accumulation. Bone marrow biopsy taken 30 hours after injection showed no infiltrations, but mild hypoplasia suggesting chronic disease was demonstrated. Retrospectively, the authors performed digital autoradiography from tissue samples and found specific targeting with the same BW250/183 antibody. CONCLUSION: Their results indicate that antibody targeting is possible in multifocal eosinophilic granuloma, probably even allowing radioimmunotherapy.

Increased thymidylate synthase gene expression in metastatic melanoma.

Thymidylate synthase (TS) provides the only de novo source of thymidylate for DNA synthesis and is a key target for cancer chemotherapeutic agents. We investigated the TS gene expression by semiquantitative reverse-transcriptase polymerase chain reaction in metastatic melanoma and compared the results with those from control tissues. The relative TS/beta-actin level ratios were 0.5, 0.9, 0.3, 0.4, and 0.5 (mean 0.5) in skin, lymph node, thyroid, muscle, and spleen, respectively. In metastatic melanoma samples, the ratios varied from 0.9 to 2.7 (mean 2.0). The differences of expression levels between these two groups of samples were statistically highly significant (p = 0.0000713). A similar statistical significance (p = 0.0002) was observed between patients achieving a complete response and patients who had progressive disease despite immunochemotherapy. There was no clear relationship between a high TS/ beta-actin ratio and the S phase fraction, as all melanomas had a high S phase fraction.

Effects of lithium gammalinolenate on the perfusion of liver and pancreatic tissues in pancreatic cancer.

Because of its poor prognosis, new modalities to treat pancreatic cancer are highly welcome. Gammalinolenate (GLA) has been shown to possess antitumor activity on various human cancer cell lines in vitro and some evidence has been found of its modulative activity on tubulin active agents, such as vinca alkaloids. GLA treatment is thought to change the penetration and distribution of chemotherapeutic agents in pancreatic tumor tissue. The in vivo effects of GLA are widely unknown. This is the first study on the modulation effects of both oral or intravenous GLA on blood perfusion in vivo. We analysed tissue perfusion prior to treatment and on the 10th day of GLA treatment in patients with pancreatic cancer. Dynamic gamma imaging was performed for 20 minutes after Tc-99m-MIBI injection, and the whole body was scanned after the dynamic study and at 4 hours. Half-lives in liver, left kidney, spleen, pancreas and tumor were recorded using a developed macro program for background corrected geometric mean data from irregular region of interests. Half-lives in the liver did not change due to oral GLA treatment, but they decreased dramatically in two of three patients after i.v. GLA treatment. Additionally, individual changes were observed in pancreatic half-lives, as in four out of five cases the half-life increased and in one case it decreased. No major changes were observed in kidney and spleen half-lives. GLA treatment had no effects on the blood brain barrier. This technique demonstrates perfusion in salivary glands, thyroid, lungs, heart, spleen, kidneys, muscles, spine and bladder, but no changes in perfusion could be detected due to GLA treatment. However, qualitatively enhanced blood flow through the pancreatic tumor was observed. In all patients irrespective of the route of administration of GLA, the organ-to-background ratios in liver decreased. The effect is, however, smallest after oral dosing. The pancreas-to-background ratio was increased in 3/5 patients, these patients exhibited stabilized disease. In a patient with large liver metastases the pancreas-to-background ratio decreased, and she showed a rapid disease progression during GLA therapy. The change in the pancreatic uptake was inversely proportional to the change in CA 19-9 concentration. Our results indicate the that GLA treatment dramatically changes tissue perfusion, especially in liver and pancreatic tumors, even at low doses, and these changes may predict response to GLA therapy.

Somatostatin receptor imaging of olfactory neuroblastoma.

Neural-crest tumours, including neuroblastomas, express somatostatin receptors. This can be shown by radionuclide labelling of octreotide, a somatostatin analogue. Studies on imaging with this substance have dealt with childhood neuroblastomas. Olfactory neuroblastoma (aesthesioneuroblastoma) is a rare tumour in which somatostatin receptor content has not been analysed, nor have radionuclide methods for diagnostic purposes been described. We report a case of olfactory neuroblastoma, in which scanning with 111In-labelled octreotide was performed. A strong uptake was seen at the base of the skull. This was confirmed as a recurrent tumour by magnetic resonance (MR) imaging. Uptake was also observed in the neck and chest, indicating extensive spread of the disease. Somatostatin receptor expression has been shown to correlate with prognosis in childhood neuroblastoma. The accuracy of labelled octreotide in the diagnosis of olfactory neuroblastoma indicates that it might be useful in radionuclide therapy of patients with advanced disease, when no other treatment modalities are available.

Modulation of antibody kinetics by the cell membrane active agent tween 80 in vivo.

The modulation of antibody uptake by the tumour has been crucial in many radioantibody applications for delivering optimal dose for therapy. Our approach was to modulate the monoclonal antibody (MAb) uptake by using a surface detergent (Tween 80). Our Mab was raised against the tyrosine kinase receptor recombinant protein tie. Mice bearing Lewis lung carcinoma xenografts were studied after injecting I-125 labeled IgG1 subclass monoclonal antibody 3c4c7g6 tie protein. The biodistribution was studied at 4, 24, 48, 72, 96 and 120 hours after intravenous injection. Tween 80 was administered intratumourally, 0.04% of tumour volume. Without Tween 80 the antibody half-lives in tumour were 90 hours, in blood 39 hours, in liver 22 hours, and in kidney 52 hours, whereas using intratumoural Tween 80 half-lives were in tumour 66 hours, in blood 26 hours, in liver 27.5 hours and in kidney 27.5 hours. Although the Tween manipulation did not increase uptakes by organs, it did enhance clearance rate from the blood. This data indicates that antibody dose can be optimized by surface detergent, enhancing clearance without any burden to critical organs. This might be crucial in adjusting delivered radioimmunotherapy dose by changing the mean residence time of an antibody.

Detection of pseudomyxoma peritonei by radioimmunohistochemistry and radioimmunoscintigraphy.

Pseudomyxoma peritonei (PP) is a local slowly progressing disease with typical abdominal swelling. Treatment is uneffective and the long-term prognosis is poor. Conventional radiology provides usually only a delineation of low density area relating gelatinous masses accumulating in the peritoneal cavity. In this study, immunohistochemistry based on digital quantitative autoradiography utilizing radiolabelled monoclonal antibody B72.3 (MoAb) recognizing TAG-72 antigen on epithelial carcinomas was used for diagnosis of pseudomyxoma (7 patients). The PP patients were studied with radioiodinated I-131-labeled MoAb after intravenous (2 patients) and intraperitoneal (7 patients) injections. Radioactivities of MoAbs varied considerably in the tumors. Both intra- and extracellular staining pattern was observed by immunohistochemistry. Gamma imaging at 1, 3, 7 and 14 days after i.v. injection (2 patients) revealed targeting of all known lesions. The intraperitoneally injected MoAb (7 patients) retained long time in the peritoneal cavity, specific tumour targeting was seen up to 16 days by an antibody-SPECT, while maximum blood radioactivity was measured between 8-12 hrs. Radiolabelled B72.3 MoAb recognizing TAG-72 antigen is also present within pseudomyxoma cells. It can be used for radioimmunohistochemistry of PP. Accurate imaging of PP is possible by MoAb suggesting earlier diagnosis and more accurate location of residual disease after operations, and evaluating treatment response. Estimated tumour dose for intraperitoneal tumour (MIRD formalism) was 13 mGy/MBq. This indicates that the radioiodinated B72.3 antibody can be used for in vivo targeting and therapeutic applications of intraperitoneal pseudomyxoma.

Oligoradionuclidetherapy using radiolabelled antisense oligodeoxynucleotide phosphorothioates.

Radiolabelled antisense oligodeoxynucleotides have been used for in vivo biokinetic studies in AIDS and cancer patients. The therapeutic possibilities are still unknown and the major question in therapeutic use of radio-oligonucleotide is the optimal source of radiation. We studied the pharmacokinetics and in vivo tissue distribution for oligodeoxynucleotide phosphorothioates by using the data from three different radionuclides: sulphur-35 (t1/2 = 87.4 days, maximum beta-energy = 167 keV), phosphorus-33 (t1/2 = 24.4 days, maximum beta-energy = 250 keV) and phosphorus-32 (t1/2 = 14.3 days, maximum beta-energy = 2270 keV). The absorbed doses of 32P-, 33P- and 35S-labelled oligonucleotides were estimated using the published biodistribution data for several oligonucleotides in two animal models for both tumour xenografts and AIDS. The local energy absorption of 33P turned out to be higher than that of 32P if the mass was smaller than approximately 300 micrograms, and the local absorption of 35S was higher than that of 32P when the mass was <80 micrograms. In a mouse tumour xenograft model an i.v. injected activity seemed to achieve sufficient radiation doses in the tumour: in a 1 g tumour 4.9 Gy for 32P, 5.1 Gy for 33P and 5.5 Gy for 35S were calculated when the kidney dose was kept as 5 Gy. In the same model in smaller tumours the doses were for a 1 mg tumour 0.73 Gy (32P), 5.1 Gy (33P) and 5.5 Gy (35S), and for a 1 microgram tumour 0.08 Gy (32P), 3.1 Gy (33P) and 3.9 Gy (35S). Thus, 33P and 35S have more beneficial radiotherapeutic characteristics than 32P. Relative advantage factors (33P and 35S versus 32P) for kidney and liver doses using these nuclides varied from 0.997 to 1.001 for a 1 g tumour and there was no difference in the radiation dose to normal organs. Therefore, we conclude that in oligonucleotide radiotherapy tumours >1 g should be treated with 32P, whereas smaller tumours should be treated with 33P or 35S. There is no significant difference between 33P and 35S, and either radionuclide could be selected according to labelling properties.

Treatment of olfactory neuroblastoma. A report of 11 cases.

Eleven olfactory neuroblastomas treated at Helsinki University Central Hospital between 1970 and 1991 were reviewed retrospectively. The distribution of the patients was according to Morita's staging (modified Kadish's classification) as follows: one stage A, one stage B and nine stage C. Tumor resections were performed in all cases, and five were considered radical. All patients received radiotherapy: total doses ranged from 42 to 70 Gy. At least a short treatment response was achievable in all cases. Chemotherapy was given to two patients: one remission was obtained by methotrexate with leucovorin rescue and doxorubicin for residual disease after radiotherapy. Distant metastases were observed in three cases, two in the lungs and one intraperitoneally. After a median follow-up of 63 (range, 6-140) months, five patients are alive and well, two patients have died with no evidence of disease; two patients who received only 42 and 50 Gy in 5 and 6 weeks have died of local recurrences and two of distant metastases. None of the patients with advanced (stage B or C) disease who received radical radiotherapy ( > or = 60 Gy given in 6 to 9 weeks) developed local recurrence. The increasing incidence of distant metastasis justifies an intensification of initial treatment, especially in state C disease.

Indium-111 bleomycin complex for radiochemotherapy of head and neck cancer--dosimetric and biokinetic aspects.

Bleomycin (BLM) is used for the treatment of head and neck cancer. In order to improve the effectiveness of this chemotherapeutic drug, BLM was combined with indium-111. A complex of these agents (111In-BLMC), formed at low pH, was injected intravenously into ten head and neck cancer patients in escalating activities of 75, 175 and 375 MBq. The internally delivered dose to the tumours varied from 0.20 to 2.73 mGy at 75 MBq, from 0.33 to 2.51 mGy at 175 MBq, and from 0.87 to 31.3 mGy at the 375 MBq activity level. Uptake of radioactivity was 0.45+/-0.24x10(-3)% ID/g in primary tumours and 0. 52+/-0.20x10(-3)% ID/g in metastases (at 48 h). Tumour volumes varied from 0.51 to 49.0 cm3. The radioactivity half-lives in the tumours were 30+/-7 h. The activity distribution and penetration into tumour tissue were not affected by increasing the injected activity. There was a positive correlation between BLMC uptake and Ki-67/Mib activity as well as number of mitoses in tumour tissue. These data indicate that 111In-BLMC has potential as a radiochemotherapeutic agent in head and neck cancer and that adjuvant Auger-electron therapy is possible using 114mIn-labelled BLMC.