Blood Levels Of Radioactivity Research Articles

11C]Paraoxon: Radiosynthesis, Biodistribution and In Vivo Positron Emission Tomography Imaging in Rat.

Synthesis of the acetylcholinesterase inhibitor paraoxon (POX) as a carbon-11 positron emission tomography tracer ([11C]POX) and profiling in live rats is reported. Naïve rats intravenously injected with [11C]POX showed a rapid decrease in parent tracer to ∼1%, with an increase in radiolabeled serum proteins to 87% and red blood cells (RBCs) to 9%. Protein and RBC leveled over 60 minutes, reflecting covalent modification of proteins by [11C]POX. Ex vivo biodistribution and imaging profiles in naïve rats had the highest radioactivity levels in lung followed by heart and kidney, and brain and liver the lowest. Brain radioactivity levels were low but observed immediately after injection and persisted over the 60-minute experiment. This showed for the first time that even low POX exposures (∼200 ng tracer) can rapidly enter brain. Rats given an LD50 dose of nonradioactive paraoxon at the LD50 20 or 60 minutes prior to [11C]POX tracer revealed that protein pools were blocked. Blood radioactivity at 20 minutes was markedly lower than naïve levels due to rapid protein modification by nonradioactive POX; however, by 60 minutes the blood radioactivity returned to near naïve levels. Live rat tissue imaging-derived radioactivity values were 10%-37% of naïve levels in nonradioactive POX pretreated rats at 20 minutes, but by 60 minutes the area under the curve (AUC) values had recovered to 25%-80% of naïve. The live rat imaging supported blockade by nonradioactive POX pretreatment at 20 minutes and recovery of proteins by 60 minutes. SIGNIFICANCE STATEMENT: Paraoxon (POX) is an organophosphorus (OP) compound and a powerful prototype and substitute for OP chemical warfare agents (CWAs) such as sarin, VX, etc. To study the distribution and penetration of POX into the central nervous system (CNS) and other tissues, a positron emission tomography (PET) tracer analog, carbon-11-labeled paraoxon ([11C]POX), was prepared. Blood and tissue radioactivity levels in live rats demonstrated immediate penetration into the CNS and persistent radioactivity levels in tissues indicative of covalent target modification.

Population pharmacokinetic dosimetry model using imaging data to assess variability in pharmacokinetics of 177 Lu-PSMA-617 in prostate cancer patients.

Studies to evaluate and optimize [177 Lu]Lu-PSMA treatment focus primarily on individual patient data. A population pharmacokinetic (PK) dosimetry model was developed to explore the potential of using imaging data as input for population PK models and to characterize variability in organ and tumor uptake of [177 Lu]Lu-PSMA-617 in patients with low volume metastatic prostate cancer. Simulations were performed to identify the effect of dose adjustments on absorbed doses in salivary glands and tumors. A six-compartment population PK model was developed, consisting of blood, salivary gland, kidneys, liver, tumor, and a lumped compartment representing other tissue (compartment 1-6, respectively), based on data from 10 patients who received [177 Lu]Lu-PSMA-617 (2 cycles, ~ 3 and ~ 6GBq). Data consisted of radioactivity levels (decay corrected) in blood and tissues (9 blood samples and 5 single photon emission computed tomography/computed tomography scans). Observations in all compartments were adequately captured by individual model predictions. Uptake into salivary glands was saturable with an estimated maximum binding capacity (Bmax ) of 40.4MBq (relative standard error 12.3%) with interindividual variability (IIV) of 59.3% (percent coefficient of variation [CV%]). IIV on other PK parameters was relatively minor. Tumor volume was included as a structural effect on the tumor uptake rate constant (k15 ), where a two-fold increase in tumor volume resulted in a 1.63-fold increase in k15 . In addition, interoccasion variability on k15 improved the model fit (43.5% [CV%]). Simulations showed a reduced absorbed dose per unit administered activity for salivary glands after increasing radioactivity dosing from 3 to 6GBq (0.685 Gy/GBq vs. 0.421 Gy/GBq, respectively). All in all, population PK modeling could help to improve future radioligand therapy research.

Molecular Imaging of HER2 Expression in Breast Cancer patients Using the [99mTc] Tc-Labeled Small Peptide

PurposeThe accurate determination of human epidermal growth factor receptor 2 (HER2) status can predict response to treatment with HER2-targeted therapy for HER2-positive breast cancer patients. [99mTc]Tc-HYNIC-(Ser)3-LTVPWY ([99mTc]Tc-HYNIC-LY) is a small synthetic peptide molecule targeting of the HER2 receptor. This clinical study evaluated the pharmacokinetic, dosimetry, and efficacy of [99mTc]Tc-HYNIC-LY for determining the HER2 status in primary breast cancer patients. Materials and MethodsIn total, 24 women with suspected primary breast cancer received an intravenous injection of approximately 20 µg (∼740 MBq) of [99mTc]Tc-HYNIC-LY. In the first 3 patients, blood levels of radioactivity were analyzed for pharmacokinetic evaluation and planar gamma camera imaging was conducted at 30 min and 1, 2, 4, and 24 hour after injection for dosimetry assessment. In the last 21 patients, planar imaging was performed at the baseline, as well as 1, 2, 3, and 4 hour, followed by single-photon emission computed tomography (SPECT) imaging after 4 hour to evaluate the tumor-targeting potential in primary lesions. ResultsInjection of [99mTc]Tc-HYNIC-LY was safe and well tolerated. Fast blood clearance provided high-contrast HER2 imaging within 1 to 4 hour. The highest absorbed radiation dose was found for kidneys (6.78E-03 ± 2.62E-04 mSv/MBq), followed by the heart (3.73E-03 ± 1.98E-04 mSv/MBq). The [99mTc]Tc-HYNIC-LY peptide was able to detect HER2 status in primary tumors at an acceptable level. ConclusionThe findings of this study indicated that [99mTc]Tc-HYNIC-LY SPECT is safe and feasible for the identification of HER2-positive lesions in primary breast cancer patients, and may provide an accurate and non-invasive modality for guiding HER2 targeted therapy.

Synthesis and evaluation of N-isopropyl-p-[11C]methylamphetamine as a novel cerebral blood flow tracer for positron emission tomography

IntroductionIncreases in fasting plasma glucose (PG) levels lead to a decrease in 2-deoxy-2-[18F]fluoro-d-glucose ([18F]FDG) uptake in the normal brain, especially in the precuneus, resulting in an Alzheimer’s disease (AD)-like uptake pattern. Therefore, patients with higher PG levels, such as those with diabetes, can be erroneously diagnosed with AD when positron emission tomography (PET) imaging is done using [18F]FDG, due to reduced uptake of [18F]FDG in the precuneus. To help avoid an erroneous diagnosis of AD due to differences in glucose metabolism, evaluating cerebral blood flow (CBF) in the brain is useful. However, current techniques such as single photon emission computed tomography (SPECT) and [15O]H2O PET have limitations regarding early diagnosis of AD because the images they produce are of low resolution. Here, we developed a novel CBF PET tracer that may be more useful than [18F]FDG for diagnosis of AD.MethodsWe synthesized and evaluated N-isopropyl-p-[11C]methylamphetamine ([11C]4) as a carbon-11-labeled analogue of the standard CBF SPECT tracer N-isopropyl-p-[123I]iodoamphetamine. Fundamental biological evaluations such as biodistribution, peripheral metabolism in mice, and brain kinetics of [11C]4 in non-human primates with PET with successive measurement of [15O]H2O were performed.Results[11C]4 was synthesized by methylation of the corresponding tributyltin precursor (2) with [11C]MeI in a palladium-promoted Stille cross-coupling reaction. The brain uptake of [11C]4 in mice peaked at 5–15 min after injection and then promptly decreased. Most radioactivity in the brain was detected in the unchanged form, although in the periphery, [11C]4 was rapidly metabolized to hydrophilic components. Acetazolamide (AZM) treatment significantly increased the brain uptake of [11C]4 without affecting the blood levels of radioactivity in mice. Preliminary kinetics analysis showed that the K1 of [11C]4 reflected regional CBF in a vehicle-treated monkey, but that the K1 did not reflect CBF in higher flow regions after AZM loading.Conclusion[11C]4 is a potential novel CBF PET tracer. Further validation studies are needed before [11C]4 can be used in humans.

Impact of multiple negative charges on blood clearance and biodistribution characteristics of 99mTc-labeled dimeric cyclic RGD peptides.

This study sought to evaluate the impact of multiple negative charges on blood clearance kinetics and biodistribution properties of 99mTc-labeled RGD peptide dimers. Bioconjugates HYNIC-P6G-RGD2 and HYNIC-P6D-RGD2 were prepared by reacting P6G-RGD2 and P6D-RGD2, respectively, with excess HYNIC-OSu in the presence of diisopropylethylamine. Their IC50 values were determined to be 31 ± 5 and 41 ± 6 nM, respectively, against 125I-echistatin bound to U87MG glioma cells in a whole-cell displacement assay. Complexes [99mTc(HYNIC-P6G-RGD2)(tricine)(TPPTS)] (99mTc-P6G-RGD2) and [99mTc(HYNIC-P6D-RGD2)(tricine)(TPPTS)] (99mTc-P6D-RGD2) were prepared in high radiochemical purity (RCP > 95%) and specific activity (37–110 GBq/μmol). They were evaluated in athymic nude mice bearing U87MG glioma xenografts for their biodistribution. The most significant difference between 99mTc-P6D-RGD2 and 99mTc-P6G-RGD2 was their blood radioactivity levels and tumor uptake. The initial blood radioactivity level for 99mTc-P6D-RGD2 (4.71 ± 1.00%ID/g) was ∼5× higher than that of 99mTc-P6G-RGD2 (0.88 ± 0.05%ID/g), but this difference disappeared at 60 min p.i. 99mTc-P6D-RGD2 had much lower tumor uptake (2.20–3.11%ID/g) than 99mTc-P6G-RGD2 (7.82–9.27%ID/g) over a 2 h period. Since HYNIC-P6D-RGD2 and HYNIC-P6G-RGD2 shared a similar integrin αvβ3 binding affinity (41 ± 6 nM versus 31 ± 5 nM), the difference in their blood activity and tumor uptake is most likely related to the nine negative charges and high protein binding of 99mTc-P6D-RGD2. Despite its low uptake in U87MG tumors, the tumor uptake of 99mTc-P6D-RGD2 was integrin αvβ3-specific. SPECT/CT studies were performed using 99mTc-P6G-RGD2 in athymic nude mice bearing U87MG glioma and MDA-MB-231 breast cancer xenografts. The SPECT/CT data demonstrated the tumor-targeting capability of 99mTc-P6G-RGD2, and its tumor uptake depends on the integrin αvβ3 expression levels on tumor cells and neovasculature. It was concluded that the multiple negative charges have a significant impact on the blood clearance kinetics and tumor uptake of 99mTc-labeled dimeric cyclic RGD peptides.

First-in-Human Molecular Imaging of HER2 Expression in Breast Cancer Metastases Using the 111In-ABY-025 Affibody Molecule

The expression status of human epidermal growth factor receptor type 2 (HER2) predicts the response of HER2-targeted therapy in breast cancer. ABY-025 is a small reengineered Affibody molecule targeting a unique epitope of the HER2 receptor, not occupied by current therapeutic agents. This study evaluated the distribution, safety, dosimetry, and efficacy of (111)In-ABY-025 for determining the HER2 status in metastatic breast cancer. Seven patients with metastatic breast cancer and HER2-positive (n = 5) or -negative (n = 2) primary tumors received an intravenous injection of approximately 100 μg (∼ 140 MBq) of (111)In-ABY-025. Planar γ-camera imaging was performed after 30 min, followed by SPECT/CT after 4, 24, and 48 h. Blood levels of radioactivity, antibodies, shed serum HER2, and toxicity markers were evaluated. Lesional HER2 status was verified by biopsies. The metastases were located by (18)F-FDG PET/CT 5 d before (111)In-ABY-025 imaging. Injection of (111)In-ABY-025 yielded a mean effective dose of 0.15 mSv/MBq and was safe, well tolerated, and without drug-related adverse events. Fast blood clearance allowed high-contrast HER2 images within 4-24 h. No anti-ABY-025 antibodies were observed. When metastatic uptake at 24 h was normalized to uptake at 4 h, the ratio increased in HER2-positive metastases and decreased in negative ones (P < 0.05), with no overlap and confirmation by biopsies. In 1 patient, with HER2-positive primary tumor, (111)In-ABY-025 imaging correctly suggested a HER2-negative status of the metastases. The highest normal-tissue uptake was in the kidneys, followed by the liver and spleen. (111)In-ABY-025 appears safe for use in humans and is a promising noninvasive tool for discriminating HER2 status in metastatic breast cancer, regardless of ongoing HER2-targeted antibody treatment.

Abstract P4-01-17: Molecular imaging and quantification of HER-2 status in metastatic breast cancer by using 68Ga-ABY-025/PET-CT

Abstract Background: Molecular imaging, in particular, positron emission tomography (PET) provides unique possibility to non-invasively quantify the biomarkers. A small peptide, ABY-025, is targeting a unique epitope of the HER2-receptor and can be labeled with 111In or 68Ga. It is non-competitive with current antibody-targeted epitopes and is approximately 25 times smaller than an antibody and posses superior pharmacokinetics as imaging agent. We have earlier reported promising results of lesion visualization and determination of HER2 status by 111In-ABY-025/SPECT-CT in seven patients with metastatic breast cancer (MBC). As PET-CT provides superior possibilities for quantification and allows for imaging with shorter time frames, we now investigate 68Ga-ABY-025 in HER2-positive and HER2-negative MBC patients in a prospective study. The influence of the administered amount of ABY-025 on the discrimination between HER2-positive and HER2-negative metastases as well as detection rate and image contrast is investigated. The HER2 status changes will be followed at disease progression. We report here promising data on the first three patients. Patients and methods: The study considers twenty patients in two cohorts. Of the first 10 patients, 4 should have HER2-negative MBC and 6 patients HER2-positive disease. Metastases were localized by 18F-FDG/PET-CT prior HER2 imaging. Each patient (two HER2-positive and one HER-negative) underwent two subsequent examinations. First, 68Ga-ABY-025 PET/CT (ABY-025: 75±14 μg) was performed with dynamic acquisition during 0-45 minutes and static images after 1, 2 and 4 hours. The procedure was repeated after one week with a higher dose of 68Ga-ABY-025 (ABY-025: 445±10 μg) to identify the optimal dosage for the discrimination between HER2-positive and HER2-negative metastases. Blood levels of radioactivity, anti-ABY-025-antibodies, shed serum HER2 and toxicity markers were evaluated. Biopsies were taken to verify the HER2 status of the lesions. The second cohort of 10 patients (all with HER2-positive disease), will have their primary PET-CT with the optimal dose of 68Ga-ABY-025 and will, as the HER-positive from the first cohort, have a follow-up 68Ga-ABY-025 PET/CT after disease progression or, at latest, after 9 months. Results: The imaging agent was well tolerated by the patients without adverse events. The rapid blood clearance and normal tissue wash out allowed high contrast HER2 images within a few hours. The HER2 status was confirmed by histological analysis of biopsies. In one patient with HER2-negative primary tumor, imaging suggested a HER2-positive liver metastasis. The change of HER2 expression was verified with analysis of the corresponding biopsy. This finding resulted in the change of the treatment management. Preliminary validation suggests the preference of high dose ABY-025 in terms of image contrast and detection. The study is ongoing and more patients will be reported Conclusion: Preliminary results indicate strong potential of 68Ga-ABY-025 PET/CT for discriminating HER2-positive versus HER2-negative metastatic breast cancer, regardless of ongoing HER2-targeted antibody treatment. The method introduces rapid whole-body receptor mapping of primary tumor and metastases in a single examination. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-01-17.

Biodistribution of Nanostructured Lipid Carriers (NLCs) after intravenous administration to rats: Influence of technological factors

Nanoparticles for medical applications are frequently administered via parenteral administration. In this study, the tissue distribution of three lipid formulations based on Nanostructured Lipid Carriers (NLCs) after intravenous administration to rats was evaluated. NLCs were prepared by a high pressure homogenization method and varied in terms of particle size, surface charge, and surfactant content. The 99mTc radiolabeled NLCs were intravenously administered to rats, and radioactivity levels in blood and tissues were measured. Cmax, AUC0–24, and MRT0–24 were obtained from the radioactivity level versus time profiles. The radiolabeled nanocarriers exhibited a long circulation time since radioactivity was detected in blood even 24h post-injection. No differences on the MRT values in blood among the NLCs were observed, in spite of the different particle size and surface charge. The highest radioactivity levels were measured in the kidney, followed by the bone marrow, the liver, and the spleen. In the kidney, there was a higher accumulation of the positive nanoparticles, and in the liver, uptake of negative nanoparticles was higher than positive ones. NLCs with the largest particle size showed a higher uptake in the lung and lower accumulation in liver and bone marrow, in comparison with the smaller ones.

Afatinib pharmacokinetics and metabolism after oral administration to healthy male volunteers

To investigate the pharmacokinetics, metabolism and tolerability of afatinib (BIBW 2992), an oral irreversible ErbB family blocker, in healthy male volunteers. In this open-label, single-center study, 8 healthy male volunteers received a single oral dose of 15 mg [(14)C]-radiolabeled afatinib (equivalent to 22.2 mg of the dimaleinate salt) as a solution. Blood, urine and fecal samples were collected for at least 96 hours (h) after dosing. Plasma and urine concentrations of afatinib were analyzed using high-performance liquid chromatography-tandem mass spectrometry. [(14)C]-radioactivity levels in plasma, whole blood, urine and feces were measured by liquid scintillation counting methods. Metabolite patterns were assessed by high-performance liquid chromatography. [(14)C]-radioactivity was mainly excreted via feces (85.4%). Overall recovery of [(14)C]-radioactivity was 89.5%, indicative of a complete mass balance. Afatinib was slowly absorbed, with maximum plasma concentrations achieved at a median of 6 h after dosing, declining thereafter in a biexponential manner. The geometric mean terminal half-life of afatinib was 33.9 h in plasma and longer for [(14)C]-radioactivity in plasma and whole blood. Apparent total body clearance for afatinib was high (geometric mean 1,530 mL/min). The high volume of distribution (4,500 L) in plasma may indicate a high tissue distribution. Afatinib was metabolized to only a minor extent, with the main metabolite afatinib covalently bound to plasma proteins. Oxidative metabolism mediated via cytochrome P-450 was of negligible importance for the elimination of afatinib. Afatinib was well tolerated. Afatinib displayed a complete mass balance with the main route of excretion via feces. Afatinib undergoes minimal metabolism.

MicroSPECT/CT Imaging of Human Leukemia Engraftment In NOD-Scid Mice Using [111In]-Labeled 7G3 Anti-CD123 Antibodies

AbstractAbstract 968Radioimmunotherapy (RIT) of acute myelogenous leukemia (AML) is a promising strategy that could improve the long-term outcome of patients. A limitation of current therapies is their inability to eradicate the leukemic stem cell (LSC), which is responsible for causing and propagating the disease and the failure to achieve long-term survival. Monoclonal antibody (mAb) 7G3 recognizes CD123 (IL-3R α) in the absence of CD131 (IL-3R β) a LSC characteristic but not of normal hematopoietic stem cells. Our laboratory is developing 111In-labeled 7G3 modified with nuclear localization sequence (NLS) peptides for Auger electron RIT of AML aimed at the LSC. These electrons have only nanometer-micrometer range but are lethal when released inside the cell nucleus. Our objective was to examine the ability of 111In-NLS-7G3 to localize at sites of AML engraftment in NOD-scid mice taking advantage of the concurrent gamma emissions of 111In for microSPECT/CT imaging. These properties were examined in NOD-scid mice engrafted with CD123+/CD131- AML cell lines which avoids the challenges of engrafting primary AML specimens including their variability in engraftment efficiency and heterogeneity in CD123 expression. Alternatively, mice bearing s.c. CD123-transfected Raji (Raji-CD123) tumors were studied. 7G3 was reacted with sulfo-SMCC to introduce maleimide groups for conjugation to NLS peptides (CGYGPKKKRKVGG) then directly derivatized with DTPA for complexing 111In. The pharmacokinetics (PK) of elimination from the blood of 111In-NLS-7G3 intact IgG2a and F(ab')2 fragments were studied in NOD-scid and Balb/c mice. The effect of pre-administration of a 10-fold excess of unlabeled BM4 IgG2a or IgG1 irrelevant mAbs on the PK was assessed. Imaging and biodistribution studies were performed at 48 to 72 h p.i. of 111In-NLS-7G3 IgG2a and F(ab')2 in AML-engrafted mice. Sites of engraftment visualized on the images were analyzed for AML by flow cytometry. PK studies revealed an unusually rapid elimination of 111In-NLS-7G3 IgG2a but not F(ab')2 in NOD-scid compared to Balb/c mice with blood radioactivity levels at 18 h 4-fold lower (3.9 ± 0.9 vs. 17.0 ± 4.4% i.d./ml, respectively). This rapid elimination prevented uptake into Raji-CD123 xenografts and did not allow their visualization by microSPECT/CT. The elimination of 111In-NLS-7G3 F(ab')2 was not significantly different in NOD-scid or Balb/c mice (0.60 ± 0.17 vs. 0.87 ± 0.06 %i.d./ml, respectively at 72 h p.i.). Pre-administration of unlabeled BM4 improved the retention of 111In-NLS-7G3 in the blood by 3.2-fold and allowed for high uptake (14.7 ± 2.8 % i.d./g) and imaging of tumors. Xenografts also had high uptake (7.2 ± 3.8 % i.d./g) and imaged with 111In-NLS-7G3 F(ab')2. AML-3, -4 and -5 cells engrafted into bone marrow (BM) and NOD-scid mice assessed by the proportion of CD45+ cells (12%, 0.5% and 63%, respectively). CD45+ cells in the spleen for AML-5 engraftment was 83%. Uptake of 111In-NLS-7G3 F(ab')2 in the spleen and BM of AML-3 engrafted mice were 5-fold and 1.5-fold higher than 111In-NLS-BM4 control F(ab')2. The spleen was visualized by microSPECT/CT in engrafted mice receiving 111In-NLS-7G3 F(ab')2 but not with 111In-NLS-BM4 F(ab')2. In AML-3 engrafted mice imaged with 111In-NLS-7G3 IgG2a, multiple intense foci of radioactivity were noted in the spine, pelvis and femur, representing leukemia in the marrow. Extramedullary (EM) disease was also imaged in the brain and lymph nodes. Uptake in the femur was 2-fold higher in engrafted mice compared to non-engrafted mice. In AML-5 engrafted mice, outgrowths of AML at the femoral heads was imaged with 111In-NLS-7G3 IgG2a following pre-administration of an excess of unlabeled BM4. No visualization of these AML growths was noted for mice receiving 111In-NLS-BM4. We conclude that microSPECT/CT allowed sensitive detection of the specific uptake of 111In-NLS-7G3 and its F(ab')2 fragments in AML engrafted mice. PK and Raji-CD123 xenograft microSPECT/CT studies identified a tumor delivery barrier. A robust AML engraftment model was established with cells displaying the CD123+/CD131- phenotype. These results are promising for preclinical evaluation of 111In-NLS-7G3 for Auger electron RIT of AML and suggest that imaging is a useful tool to assess engraftment in NOD-scid mice as well as for evaluating response to 111In-NLS-7G3. Preliminary results of studies to examine 111In-NLS-7G3 for RIT of AML will also be presented. Disclosures:No relevant conflicts of interest to declare.

Enhanced Target-Specific Accumulation of Radiolabeled Antibodies by Conjugating Arginine-Rich Peptides as Anchoring Molecules

We have devised and estimated a new strategy to prolong the residence time of radiolabeled antibodies in tumor in which an octaarginine peptide (R₈) was used as an anchoring molecule to fix antibodies against CD20 (NuB2; IgG2a) on tumor cells. Conjugation of R₈ with antibodies was performed by maleimide-thiol chemistry using thiol groups generated by reducing the disulfide bonds of the antibody. The R₈-conjugated NuB2 was then reacted with succinimidyl meta-[¹²⁵I]iodobenzoate to prepare [¹²⁵I]SIB-NuB2(I) (0.92 R₈/NuB2) and [¹²⁵I]SIB-NuB2(III) (3.38 R₈/NuB2). Both SIB-NuB2(I) and SIB-NuB2(III) exhibited size-exclusion HPLC elution profiles and immunoreactivity to CD20-positive cells similar to those of NuB2. NuB2(I) also possessed isoelectric focusing (IEF) profile similar to NuB2. However, NuB2(III) registered a broad IEF band toward higher pI. When incubated with CD20-positive cells, [¹²⁵I]SIB-NuB2(I) and [¹²⁵I]SIB-NuB2(III) exhibited 1.4 and 4.0 times higher cell-associated radioactivity than [¹²⁵I]SIB-NuB2. After the cells were washed and reincubated in a fresh medium for 3 h, [¹²⁵I]SIB-NuB2(I) and [¹²⁵I]SIB-NuB2(III) exhibited significantly higher cell-associated radioactivity than [¹²⁵I]SIB-NuB2. In biodistribution studies in normal mice, while both [¹²⁵I]SIB-NuB2(I) and [¹²⁵I]SIB-NuB2 exhibited similar biodistribution profiles, [¹²⁵I]SIB-NuB2(III) showed faster clearance from the blood and higher hepatic radioactivity levels than [¹²⁵I]SIB-NuB2. In SCID mice bearing CD20-positive xenografts, [¹³¹I]SIB-NuB2(I) exhibited significantly higher radioactivity in xenografts than those of [¹²⁵I]SIB-NuB2 with no significant increase being observed in other tissues. The findings indicate that appropriate R₈ modification of antibodies satisfies both specific targeting ability of antibody and strong cell-association property of R₈, which was reflected in the increased radioactivity levels in tumor. These findings supported the applicability of this approach to enhance target-specific accumulation of radiolabeled antibodies.

Absorption and Excretion of Colestilan in Healthy Subjects

Colestilan, an anion-exchange resin binding both phosphate and bile-acid anions, is under development for the treatment of hyperphosphataemia and dyslipidaemia, which occur in the majority of end-stage renal disease patients. This study using 14C-colestilan was conducted to investigate the absorption and excretion of colestilan in humans. Following a 28-day run-in period with administration of colestilan 3 g three times daily, 12 subjects received a single oral dose of 14C-colestilan 100 mg (approximately 4.0 MBq) and colestilan 3 g under fasted conditions on the morning of day 1. A total of 9 g of colestilan was administered three times daily on days 1-4. Total radioactivity levels in whole blood (at 4, 8, 12 and 24 hours and then at 24-hour intervals) and in the urine and faeces (from 0 to 24 hours and then at 24-hour intervals) were monitored up to 216 hours postdose (day 10). Total radioactivity measured in all whole-blood samples was below the lower limit of quantification (0.025 microg equivalent of 14C-labelled colestilan/mL of whole blood). Total radioactivity assessed in all urine samples was also below the lower limit of quantification (0.003 microg equivalent/mL for urine), except at 0-24 hours postdose, when 0.01% of the radioactive dose was excreted by all subjects. This level was below the predetermined water soluble impurity level of 0.04%. The mean cumulative excretion of total radioactivity in the faeces was 99.66% by 216 hours postdose, excluding one subject with incomplete collection of faecal samples. These results demonstrate that colestilan is not absorbed from the gastrointestinal tract and is completely excreted in the faeces.

Pharmacokinetics (PK) of [14C]-BIBW 2992 after administration of a single dose of 15 mg [14C]-BIBW 2992 oral solution in healthy male volunteers

14607 Background: BIBW 2992 is a novel oral, potent and irreversible inhibitor of EGFR and HER2. This study investigated the PKs and the mass balance of a single oral dose of [14C]-radiolabelled BIBW 2992 in healthy male volunteers. Safety and tolerability of BIBW 2992 were also investigated. Methods: This was a Phase I, open label, single dose study with [14C]-radiolabelled BIBW 2992 performed at one study centre in 8 healthy male subjects receiving one single dose of 15 mg of BIBW 2992 containing 2.25 MBq of [14C]-radiolabelled BIBW 2992. Observation period for the collection of samples of blood, urine and feces was at least 120 hours. Plasma and urine concentrations of BIBW 2992 were analyzed using high performance liquid chromatography coupled to tandem mass spectrometry. [14C]-radioactivity levels in plasma, whole blood, urine and feces were analyzed by Liquid Scintillation Counting methods. Results: The major route of elimination of total [14C]-radioactivity was found to be via feces (85.4%) with an overall recovery of [14C]-radioactivity of 89.5%. For BIBW 2992 as well as for [14C]-plasma and whole blood radioactivity, maximum plasma concentrations were reached around 6 hours after dosing. The gMean terminal half-life (t1/2) was 33.9 hours for BIBW 2992, 118 hours for [14C]-radioactivity in plasma and 195 hours for [14C]-radioactivity in whole blood. A relatively high apparent total body clearance (CL/F) for BIBW 2992 was determined. The gMean CL/F values for the [14C]- radioactivity in plasma and whole blood were low. BIBW 2992 as well as [14C]-radioactivity in plasma and in whole blood exhibited a high apparent volume of distribution (Vz/F) during the terminal phase. BIBW 2992 was generally tolerated with only mild adverse events of short duration (all CTCAE Grade 1) reported. Conclusions: BIBW 2992 was safe and well tolerated at a single dose of 15 mg. The major route of elimination of BIBW 2992 is via feces. The data suggest the presence of one or more metabolites of BIBW 2992 in plasma and in whole blood with a longer terminal half-life than BIBW 2992. Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Expert Testimony Other Remuneration Boehringer Ingelheim Pharma GmbH & Co. KG

Experimental manipulations of afferent immune responses influence efferent immune responses to brain tumors.

Tumors grow more readily in the brain than in the periphery, in part due to immune privilege. Differences in both afferent and efferent components of the immune response contribute to this lower level of responsiveness. On the afferent side, despite the lack of lymphatic vessels in the brain, antigens from brain arrive in lymph nodes and spleen by several routes, and the route taken may influence the type of response generated. Work with viruses and soluble antigens in mice has shown that the intracerebral location and the volume of the inoculation influence the strength of the cytotoxic T cell response. We examined whether these factors influence the T cell response against experimental brain tumors in mice. Placement of tumor cells in the cerebral ventricles instead of the parenchyma generated an immune response sufficient to increase survival time. A large volume of an intraparenchymal infusion of tumor cells caused spread of cells to the ventricles, and resulted in longer survival time relative to a small volume infusion. Infusion of the same dose of radiolabeled tumor cells in either a small volume or a large volume allowed tracking of potential tumor antigens to the periphery. Both modes of infusion resulted in similar levels of radioactivity in blood, spleen and kidney. Unexpectedly, cells infused intraparenchymally in a small volume, compared to a large volume, resulted in (1) more radioactivity in cervical lymph nodes (parotid and deep cervical lymph nodes), (2) a greater number of CD11b+/Gr1+ myeloid suppressor cells in the tumors, and (3) fewer CD8+ cells within the tumor mass. Consistent with these observations, providing a stronger afferent stimulus by giving a concurrent subcutaneous injection of the same tumor cells infused into the brain increased CD8+ T cell infiltration of the tumor in the brain. These results suggest that the immune response elicited by antigens that drain predominantly to the cervical lymph nodes may be less effective than responses elicited at other lymph nodes, perhaps due to immunosuppressive cells. Directing therapies to the optimal peripheral sites may improve immune responses against brain tumors.

N-isopropyl-4-[123I]iodoamphetamine (123I-IMP) products: a difference in radiochemical purity, unmetabolized fraction, and octanol extraction fraction in arterial blood and regional brain uptake in rats

N-isopropyl-4-[123I]iodoamphetamine (123I-IMP) is a lipophilic compound utilized for cerebral blood flow (CBF) measurement with single photon emission computed tomography (SPECT). Two different 123I-IMP products (IMP(A) and IMP(B)) are commercially available. We examined the radiochemical purity, unmetabolized fraction, and octanol extraction fraction in arterial blood, and the regional brain uptake of IMP(A) and IMP(B) in a rat model. IMP(B) (96.4% +/- 0.08%, P < 0.05) showed significantly higher radiochemical purity than IMP(A) (95.5% +/- 0.20%). The mean unmetabolized fraction in arterial blood taken at 10 min after intravenous administration of IMP(B) (69.5% +/- 4.4%, P < 0.01) was significantly higher than that of IMP(A) (59.6% +/- 2.6%). The mean octanol extraction fraction of IMP(B) (75.0% +/- 1.3%, P < 0.01) was also significantly higher than that of IMP(A) (67.2% +/- 0.8%). The mean levels of radioactivity in arterial blood sampled at 10 min after injection and mean regional brain radioactivity (cerebral cortices, basal ganglia, brain stem, and cerebellum) at 10-12 min after injection were not significantly different between IMP(A) and IMP(B). The present study indicates differences in the radiochemical purity and the unmetabolized and octanol extraction fraction in arterial blood between the two commercially available 123I-IMP products. The appropriate octanol extraction fractions for IMP(A) and IMP(B) should be determined in humans and employed for quantitative CBF measurement in clinical SPECT.

Pharmacokinetics and Distribution of [35S]Methylsulfonylmethane following Oral Administration to Rats

Methylsulfonylmethane (MSM) is a sulfur-containing compound found in a wide range of human foods including fruits, vegetables, grains, and beverages. More recently, it has been marketed as a dietary supplement worldwide. The objective of this study was to evaluate the pharmacokinetic profile and distribution of radiolabeled MSM in rats. Male Sprague-Dawley rats were administered a single oral dose of [35S]MSM (500 mg/kg), and blood levels of radioactivity were determined at different time points for up to 48 h. Tissue levels of radioactivity at 48 and 120 h and urine and fecal radioactivity levels were measured at different time points for up to 120 h following [35S]MSM administration to rats. Oral [35S]MSM was rapidly and efficiently absorbed with a mean tmax of 2.1 h, Cmax of 622 microg equiv/mL, and AUC0-inf of 15124 h.microg equiv/mL. The t1/2 was 12.2 h. Soft tissue distribution of radioactivity indicated a fairly homogeneous distribution throughout the body with relatively lower concentrations in skin and bone. Approximately 85.8% of the dose was recovered in the urine after 120 h, whereas only 3% was found in the feces. No quantifiable levels of radioactivity were found in any tissues after 120 h, indicating complete elimination of [35S]MSM. The results of this study suggest that [35S]MSM is rapidly absorbed, well distributed, and completely excreted from the body.

THE EVALUATION OF RADIOLABELED ARTESUNATE ON TISSUE DISTRIBUTION IN RATS AND PROTEIN BINDING IN HUMANS

The present study reports the tissue distribution, pharmacokinetics, mass balance, and elimination of [(14)C] artesunate (AS) following single intravenous administration in rats. Protein binding was performed with rat and human plasma. Radioactivity and drug levels in blood, plasma, tissues, urine, and feces up to 192 hours were collected and measured. The mean terminal half-life of plasma (76 h) and blood (105 h) radioactivity was prolonged compared with that of unchanged AS (0.43 h) and dihydroartemisinin (0.75 h), an active metabolite of AS. Drug was widely distributed after 1 hour in select tissues. After 24 hours, the radioactivity rapidly declined in all tissues except spleen until 96 hours. Only 1% of total radioactivity was detected in brain tissue. AS revealed a higher binding capacity with human and rat plasma proteins (73-81%). The radioactivity in whole blood was higher (two to fourfold) than that in plasma throughout the period of the treatment, suggesting that AS binding to RBCs may relate to its powerful antimalarial activity.

Disposition and Biotransformation of the Acetylenic Retinoid Tazarotene in Humans

Oral tazarotene, an acetylenic retinoid, is in clinical development for the treatment of psoriasis. The disposition and biotransformation of tazarotene were investigated in six healthy male volunteers, following a single oral administration of a 6 mg (100 microCi) dose of [14C]tazarotene, in a gelatin capsule. Blood levels of radioactivity peaked 2 h postdose and then rapidly declined. Total recovery of radioactivity was 89.2+/-8.0% of the administered dose, with 26.1+/-4.2% in urine and 63.0+/-7.0% in feces, within 7 days of dosing. Only tazarotenic acid, the principle active metabolite formed via esterase hydrolysis of tazarotene, was detected in blood. One major urinary oxidative metabolite, tazarotenic acid sulfoxide, accounted for 19.2+/-3.0% of the dose. The majority of radioactivity recovered in the feces was attributed to tazarotenic acid representing 46.9+/-9.9% of the dose and only 5.82+/-3.84% of dose was excreted as unchanged tazarotene. Thus following oral administration, tazarotene was rapidly absorbed and underwent extensive hydrolysis to tazarotenic acid, the major circulating species in the blood that was then excreted unchanged in feces. A smaller fraction of tazarotenic acid was further metabolized to an inactive sulfoxide that was excreted in the urine.

Biodistribution and excretion of radioactivity after the administration of 166Ho-chitosan complex (DW-166HC) into the prostate of rat

The objective of this study was to determine the fate of the 166Ho-chitosan complex (DW-166HC) in rats by examining its absorption, distribution and excretion after administration into the prostate. About 100 microCi of DW-166HC [containing 0.1875 mg of Ho(NO3)3.5H2O and 0.25 mg of chitosan] was administered intraprostatically. The level of radioactivity in blood, urinary and faecal excretion, and radioactivity distribution were examined. To determine the effect of chitosan in DW-166HC, 166Ho nitrate alone [0.1875 mg of Ho(NO3)3.5H2O] was administered into the prostate of male rats, and radioactivity distribution was examined using whole-body autoradiography. After administration of DW-166HC into the prostate, cumulative urinary and faecal excretion over the period 0-72 h was 0.35% and 0.11%, respectively. The radioactivity at the administration site was extremely high at all time points up to 144 h (>98% of injected dose). The small amount of radioactivity which did transfer from the administration site distributed mainly to the liver, spleen, kidney cortex and bone. Compared with the DW-166HC group, the group that received 166Ho nitrate alone displayed three- to fourfold higher levels of radioactivity in the main tissues, including liver, spleen, kidney cortex and bone, at 24 h after administration (P < 0.05). The results of this study show clearly that most of the administered DW-166HC remained at the administration site. It is concluded that the chitosan complex may be used to retain 166Ho within a limited area in cancer of the prostate.

Establishing Schedules for Repeated Doses of Strontium and for Concurrent Chemotherapy in Hormone-Resistant Patients With Prostate Cancer

Strontium-89 (Sr-89) alone or with concurrent chemotherapy has a role in the treatment of patients with prostate cancer (PCP). The schedules for repeated doses of Sr-89 or for concurrent chemotherapy is undetermined. The objective of this study was to measure the effective half-life (Te) of Sr-89 using a detector available in a nuclear research facility. Blood and urine samples obtained from PCP treated with Sr-89 (Metastron, Amersham, U.K.) were measured for radioactivity with a High Pure Germanium (HPGe) detector in a gamma spectrometry system (Eurisys, France). Twenty-five urine and 22 blood samples were obtained from 8 patients during a period of 160 days after Metastron injection. Sr-89 radioactivity levels in blood and urine were quite low (<8.2 x 10(-3) microCi/mL) in all patients after 21 days, whereas Sr-85 (available in 0.5% of Metastron) urine and, to a lesser extent, blood radioactivity levels were moderately high and could be detected up to 160 days. Based on Sr-85 urine levels, the calculated Sr-89 Te ranged from 9.6 to 20.7 days. Sr-89 Te can be routinely calculated in PCP based on HPGe detection of Sr-85 radioactivity levels in urine. This measurement can establish schedules for either repeated doses of Sr-89 or concurrent chemotherapy.