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BACKGROUND: Pregnant women have increased susceptibility to Plasmodium falciparum malaria and acquire protective antibodies over successive pregnancies. Most studies that investigated malaria antibody responses in pregnant women are from high transmission areas in sub-Saharan Africa, while reports from Latin America are scarce and inconsistent. The present study sought to explore the development of antibodies against P. falciparum and Plasmodium vivax antigens in pregnant women living in a low transmission area in the Brazilian Amazon. METHODS: In a prospective cohort study, plasma samples from 408 pregnant women (of whom 111 were infected with P. falciparum, 96 had infections with P. falciparum and P. vivax, and 201 had no Plasmodium infection) were used to measure antibody levels. Levels of IgG and opsonizing antibody to pregnancy-specific variant surface antigens (VSAs) on infected erythrocytes (IEs), 10 recombinant VAR2CSA Duffy binding like (DBL domains), 10 non-pregnancy-specific P. falciparum merozoite antigens, and 10 P. vivax antigens were measured by flow cytometry, ELISA, and multiplex assays. Antibody levels and seropositivity among the groups were compared. RESULTS: Antibodies to VSAs on P. falciparum IEs were generally low but were higher in currently infected women and women with multiple P. falciparum episodes over pregnancy. Many women (21%-69%) had antibodies against each individual VAR2CSA DBL domain, and antibodies to DBLs correlated with each other (r ≥ 0.55, p < 0.0001), but not with antibody to VSA or history of infection. Infection with either malaria species was associated with higher seropositivity rate for antibodies against P. vivax proteins, adjusted odds ratios (95% CI) ranged from 5.6 (3.2, 9.7), p < 0.0001 for PVDBPII-Sal1 to 15.7 (8.3, 29.7), p < 0.0001 for PvTRAg_2. CONCLUSIONS: Pregnant Brazilian women had low levels of antibodies to pregnancy-specific VSAs that increased with exposure. They frequently recognized both VAR2CSA DBL d

Contains fulltext : 231705.pdf (Publisher’s version ) (Open Access), The extracellular matrix comprises a network of macromolecules such as collagens, proteoglycans and glycoproteins. VWA1 (von Willebrand factor A domain containing 1) encodes a component of the extracellular matrix that interacts with perlecan/collagen VI, appears to be involved in stabilizing extracellular matrix structures, and demonstrates high expression levels in tibial nerve. Vwa1-deficient mice manifest with abnormal peripheral nerve structure/function; however, VWA1 variants have not previously been associated with human disease. By interrogating the genome sequences of 74 180 individuals from the 100K Genomes Project in combination with international gene-matching efforts and targeted sequencing, we identified 17 individuals from 15 families with an autosomal-recessive, non-length dependent, hereditary motor neuropathy and rare biallelic variants in VWA1. A single disease-associated allele p.(G25Rfs*74), a 10-bp repeat expansion, was observed in 14/15 families and was homozygous in 10/15. Given an allele frequency in European populations approaching 1/1000, the seven unrelated homozygote individuals ascertained from the 100K Genomes Project represents a substantial enrichment above expected. Haplotype analysis identified a shared 220 kb region suggesting that this founder mutation arose >7000 years ago. A wide age-range of patients (6-83 years) helped delineate the clinical phenotype over time. The commonest disease presentation in the cohort was an early-onset (mean 2.0 ± 1.4 years) non-length-dependent axonal hereditary motor neuropathy, confirmed on electrophysiology, which will have to be differentiated from other predominantly or pure motor neuropathies and neuronopathies. Because of slow disease progression, ambulation was largely preserved. Neurophysiology, muscle histopathology, and muscle MRI findings typically revealed clear neurogenic changes with single isolated cases displaying additional myopathic process. We speculate that a few findings of my

The congenital myasthenic syndromes (CMS) represent aheterogeneous group of diseases with abroad spectrum of phenotypes. The common characteristic is an inherited genetic defect of the neuromuscular junction. Although in some patients the specific gene defect remains to be detected, the increasing identification of causative genes in recent years has already provided unique insights into the functionality of structural proteins at the neuromuscular junction. Neonatal and early childhood onset is observed in most CMS subtypes; however, late onset in adolescence or adulthood also occurs and establishing the diagnosis at these stages imposes particular challenges. To enable appropriate therapeutic interventions for an at least in principle treatable condition, determining the genetic cause is warranted. In this overview, the critical clinical and diagnostic features of the different CMS subtypes are presented and illustrated using typical cases. Furthermore, specific diagnostic clues are outlined. Finally, the overlap between CMS and muscular dystrophies is discussed. Illustrating characteristic patient examples, the essential clinical and additional diagnostic findings of various CMS subtypes and special diagnostic indications are presented.

Purpose We investigated whether propidium iodide (PI) enhances DNA damaging effects of ionizing and non-ionizing radiation species (X-rays, alpha-, beta-, auger electron emission and light of various wavelengths, respectively). This biophysical experimental setting allowed us, furthermore, to investigate whether Cherenkov emission can be detected by photodynamic effects and increased DNA damage. Material and methods Conformation changes of plasmid DNA were detected and quantified by gelelectrophoresis and fluorescence imaging. Hydrogen peroxide, stannous dichloride, and dimethylsulfoxide were used as chemical modulators, Tc-99m, Re-188, Ra-223, and x-ray (32 kV and 200 kV) reflected radiotoxicity and light (lambda = 254 nm, 366 nm and 530-575 nm) induced phototoxicity. Results Radiotracers and x-rays induced dose dependent DNA damage. PI did not serve as radiosensitizer in radioisotopes, while a low effect was detected in X-rays. The phototoxicity was dependent on the wavelengths of light. Light with a wavelength range of 530-575 nm in combination with PI resulted in direct DNA damage. The yield of Cherenkov emission was far below the photon emission of light irradiation and not distinguishable from general radiotoxicity. Conclusions PI binds to plasmid DNA, is not chemotoxic, and increases radiotoxicity only to minor extent. Phototoxicity and its stimulation by PI is dependent on the wavelength of the light. No kind of energy deposition was capable of inducing an Auger electron cascade. Furthermore, no increase in DNA damage induced by photodynamic effects from Cherenkov emission was detectable.

Chimeric antigen receptor (CAR) T cells have shown impressive therapeutic potential. Due to the lack of direct control mechanisms, therapy-related adverse reactions including cytokine release- and tumor lysis syndrome can even become life-threatening. In case of target antigen expression on non-malignant cells, CAR T cells can also attack healthy tissues. To overcome such side effects, we have established a modular CAR platform termed UniCAR: UniCAR T cells per se are inert as they recognize a peptide epitope (UniCAR epitope) that is not accessible on the surface of living cells. Bifunctional adapter molecules termed target modules (TM) can cross-link UniCAR T cells with target cells. In the absence of TMs, UniCAR T cells automatically turn off. Until now, all UniCAR TMs were constructed by fusion of the UniCAR epitope to an antibody domain. To open up the wide field of low-molecular weight compounds for retargeting of UniCAR T cells to tumor cells, and to follow in parallel the progress of UniCAR T cell therapy by PET imaging we challenged the idea to convert a PET tracer into a UniCAR-TM. For proof of concept, we selected the clinically used PET tracer PSMA-11, which binds to the prostate-specific membrane antigen overexpressed in prostate carcinoma. Here we show that fusion of the UniCAR epitope to PSMA-11 results in a low-molecular weight theranostic compound that can be used for both retargeting of UniCAR T cells to tumor cells, and for non-invasive PET imaging and thus represents a member of a novel class of theranostics.

Regardless of its high positron energy, 68Ga-labeled PSMA ligands have become standard of care in metabolic prostate cancer imaging. 64Cu, a radionuclide with a much longer half-life (12.7 h), is available for PSMA labeling allowing imaging much later than 68Ga. In this study, the diagnostic performance of 64Cu-labeled PSMA was compared between early and late scans. Sixteen men (median age: 70 y) with prostate cancer in different stages underwent 64Cu-PSMA-617-PET/CT 2 and 22 hours post tracer injection. Pathologic and physiologic uptakes were analyzed for both points of time. Pathologic tracer accumulations occurred in 12 patients. Five patients presented with pathologic uptake in 17 different lymph nodes, two patients showed pathologic bone uptake in nine lesions, and seven patients had pathologic PSMA uptake in eight prostatic lesions. Physiologic uptake of the renal parenchyma, urine bladder, and salivary glands decreased over time, while the physiologic uptake of liver and bowel increased. In the present study, 64Cu-PSMA-617-PET demonstrated to be feasible for imaging prostate cancer for both the primary tumor site and metastases. Later imaging showed no additional, clinically relevant benefit compared with the early scans. At least the investigated time points we chose did not vindicate the additional expenditure.

64Cu wird für die nuklearmedizinische Anwendung diskutiert. Neben der Kernumwandlung über Elektroneneinfang werden auch Positronen und Auger-Elektronen (AE) emittiert. Um deren Wirkungspotential an DNA zu untersuchen, wurden zunächst zwei Pyrenderivate mit unterschiedlicher Spacerlänge zwischen Pyreneinheit und DOTA-Chelator mit [64Cu]CuCl2 radiomarkiert. Die induzierten Strahlenschäden an pUC19 Plasmid-DNA wurden in Abhängigkeit von der Dosis untersucht.

We report the case of a 32-year-old woman with severely elevated serum creatine kinase (CK; 80,000 U/l) and progressive proximal pareses. As muscular biopsy showed inflammatory infiltrates, polymyositis was suspected and immunosuppressive treatment was initiated. However, clinical improvement could not be achieved. Gene sequencing of the DYSF-gene showed a previously unreported homozygous mutation. In summary, elevated serum CK and inflammatory infiltrates in the muscle biopsy are not specific for polymyositis, but may also occur in degenerative diseases (muscular dystrophy), such as dysferlinopathy.

Chronic inflammatory demyelinating polyneuropathy (CIDP) is a chronic progressive or relapsing autoimmune neuropathy with heterogeneous clinical presentation. Symptoms typically include symmetrical, proximal and/or distal paresis and sensory loss. Atypical CIDP variants are increasingly recognized, including subtypes with rapid onset as well as variants with pure sensory, focal or marked asymmetrical deficits. Diagnosis is established by compatible symptoms, characteristic electrophysiological features and cerebrospinal fluid analysis. In unequivocal cases, inflammatory infiltrates in sural nerve biopsy support the diagnosis. Recent studies suggest that diagnostic imaging techniques such as MRI and nerve ultrasound may become useful tools for establishing the diagnosis. First-line therapies include immunoglobulines, steroids, and plasmapheresis. Immunosuppressant agents and monoclonal antibodies are used in therapy-refractory cases or as cortison-saving agents.

Chemotherapy-induced peripheral neuropathy (CIPN) is a common and relevant side effect of antineoplastic agents such as cisplatin, paclitaxel, vincristine and bortezomib. Over the last years, significant progress has been achieved in elucidating the underlying pathomechanisms of CIPN using both in vivo and in vitro models. These studies suggest that mitochondrial toxicity, disturbed axonal transport, toxic effects on Schwann cells and activation of the immune system contribute to the pathogenesis of CIPN. This review provides an overview of the current pathogenetic concepts of CIPN. In addition, experimental approaches that aim at preventing or ameliorating neurotoxic effects of antineoplastic agents are discussed.

Ziel/Aim: Cetuximab ist ein EGFR affiner Antikörper. Bei Markierungen mit Tc-99m werden hohe Temperaturen und Reduktionsmittel benötigt, um Kopplungen an Biomolekülen zu erzielen. Mit dem Komplexbildner 2,2',2''-(1,4,7-triazonane-1,4,7-triyl)triacetic acid (NOTA) ist eine schonende Komplexierung mit Tc-99m-Tricarbonyl möglich. Mit einem doppelt funktionalisierten Alexa488-Cetuximab-NOTA-Konjugat wurden Markierungsparameter angepasst und erste Zellstudien durchgeführt. Radioaktivität und Fluoreszenz konnten mit dem gleichen Antikörper parallel in der Zelle detektiert werden. Methodik/Methods: Die Markierung des Antikörpers mit Tc-99m[(CO)3(H2O)3]+ wurde durch Anpassung der Reaktionsparameter und Reinigungsverfahren optimiert. Aussagen über Ausbeute, Qualität und Stabilität wurden mit Radio-HPLC und Radio-DC getroffen. Die Integrität des Antikörpers nach der Markierung wurde über SDS-Page untersucht. Zelluläre Studien mit EGFR-positiven A431- und EGFR-negativen MDA-Zellen beinhalteten Uptake und Bindungsstudien (Gamma-Counter) bei 4°C/37°C. Die Detektion der Fluoreszenz fand über ein invers messendes Fluoreszenzmikroskop (Zeiss) statt. Ergebnisse/Results: Eine Komplexierung Tc-99m[(CO)3(H2O)3]+ mit NOTA konnte erzielt werden. Die Markierung des modifizierten Antikörpers zeigte eine radiochemische Reinheit von > 95%. Nach 24h konnten noch 75% des markierten Antikörpers nachgewiesen werden. Zelluläre Untersuchungen ergaben max. spezifische Aufnahmen bei A431-Zellen nach 1h (48%/0,5Mio.Zellen/2MBq) mit ca. 30% Membran gebundenem Anteil, welcher nach 24h abnimmt (1,5%/0,5Mio.Zellen/2MBq). Ein fluoreszenzmikroskopischer Bindungsnachweis konnte erbracht werden. Schlussfolgerungen/Conclusions: Eine Markierung des modifizierten Cetuximabs mit Tc-99m[(CO)3(H2O)3]+ konnte entwickelt werden. Geeignete Parameter und Reinigungsverfahren wurden etablie

The no carrier added (NCA) radionuclide 197(m)Hg is accessible through proton induced nuclear reactions on gold.The decay properties of both simultaneous produced nuclea risomers 197mHg and 197Hg like convenient half life, low energy gamma radiations for imaging, Auger and conversion electrons for therapy are combined with unique chemical and physical properties of mercury and its compounds. Gold as a monoisotopic element has a natural abundance of 100% 197Au superseding expensive enrichment for the target material. Additionally, the high thermal conductivity of gold enables high beam current irradiations. For separation of target material a liquid–liquid extraction method was applied.

Progressive, atrophic, asymmetrically distributed flaccid paresis of arm and hand muscles represents a frequent symptom of neuromuscular diseases that can be attributed to injury of the arm nerves, the plexus or the cervical roots. A timely and exact diagnosis is mandatory; however, the broad spectrum of differential diagnoses often represents a diagnostic challenge. A large variety of neuromuscular disorders need to be considered, encompassing autoimmune mediated inflammatory neuropathic conditions, such as multifocal motor neuropathy, as well as chronic degenerative and nerve compression disorders. This review provides an overview of the most frequent disorders of the upper plexus and cervical roots and summarizes the characteristic clinical features as well as electrodiagnostic and laboratory test results. In addition the diagnostic value of magnetic resonance imaging and sonography is discussed.

Our objective was to develop and evaluate Ga-68- and Cu-64-labeled albumin conjugates for PET imaging of sentinel lymph nodes. Four different albumin conjugates were prepared starting from NOTA-HSA.The lymph node uptake of Ga-68- and Cu-64-labeled albumin conjugates was investigated after subcutaneous injection into the foot pad of Wistar rats. A pig model was utilized for further biological evaluation of the lymph node uptake. For all the four conjugates, radiolabeling with Ga-68 and Cu-64 resulted in >95% radiochemical yield. Denatured and mannosylated Ga-68 NOTA-HSA revealed the highest popliteal lymph node uptake in rats (2.78%±0.38% IDand 6.13%±1.13%ID 10 min and 60 min p.i., resp.).Thepopliteal lymph node reached its maximumactivity after approximately 120 min and remained constant for denatured and mannosylated Cu-64 NOTA-HSA at least up to 240 min p.i. In a pig model, 2% of the injected dose of this compound was found in the sentinel lymph node 60 min after subcutaneous injection. In conclusion, PET imaging of sentinel lymph nodes with Ga-68- and Cu-64-labeled denatured NOTA-Man-HSA could be successfully demonstrated and deserves further investigations.

Targeted regulation of protein levels is an important tool to gain insights into the role of proteins essential to cell function and development. In recent years, a method based on mutated forms of the human FKBP12 has been established and used to great effect in various cell types to explore protein function. The mutated FKBP protein, referred to as destabilization domain (DD) tag when fused with a native protein at the N- or C-terminus targets the protein for proteosomal degradation. Regulated expression is achieved via addition of a compound, Shld-1, that stabilizes the protein and prevents degradation. A limited number of studies have used this system to provide powerful insight into protein function in the human malaria parasite Plasmodium falciparum. In order to better understand the DD inducible system in P. falciparum, we studied the effect of Shld-1 on parasite growth, demonstrating that although development is not impaired, it is delayed, requiring the appropriate controls for phenotype interpretation. We explored the quantified regulation of reporter Green Fluorescent Protein (GFP) and luciferase constructs fused to three DD variants in parasite cells either via transient or stable transfection. The regulation obtained with the original FKBP derived DD domain was compared to two triple mutants DD24 and DD29, which had been described to provide better regulation for C-terminal tagging in other cell types. When cloned to the C-terminal of reporter proteins, DD24 provided the strongest regulation allowing reporter activity to be reduced to lower levels than DD and to restore the activity of stabilised proteins to higher levels than DD29. Importantly, DD24 has not previously been applied to regulate proteins in P. falciparum. The possibility of regulating an exported protein was addressed by targeting the Ring-Infected Erythrocyte Surface Antigen (RESA) at its C-terminus. The tagged protein demonstrated an important modulation of its expression.

Ziel/Aim: In einer Vielzahl menschlicher Tumore ist die Expression des Epidermal-Growth-Factor-Receptor (EGFR) nachweisbar und geht mit einer schlechteren Prognose onkologisch behandelter Patienten einher. Der EGFR wird durch seine natürlichen Liganden oder ligandenunabhängig durch Bestrahlung aktiviert. Zu den Substanzen, die den EGFR hemmen gehören monoklonale Antikörper (mAb) z.B. Cetuximab (C225). Die Markierung des C-225 mit dem Beta-Strahler Y-90 ermöglicht eine zielgerichtete Induktion von DNA-Schäden. Es sollte untersucht werden, welchen Einfluss der Zusatz von kaltem C225 auf den Uptake von Y-90-C225 in FaDu-Monolayer-Zellen hat. Methodik/Methods: FaDu-Zellen wurden in 6-Well-Mikrotiterplatten (MTP) ausgesät (1x105 und 3x105 Zellen pro well). Die Inkubation erfolgte mit Y-90-C225 über 24 h bei 37°C mit unterschiedlichen Dosen (0-6 Gy, Geant4 Monte Carlo Toolkits). Y-90-C225 wurde mit unmarkiertem C225 auf C225-Konzentrationen von 1 µg/ml und 5 µg/ml eingestellt und Proben ohne Zusatz von unmarkiertem C225 mitgeführt. Die Bestimmung des zellulär gebundenen Y-90-C225 (Uptake) erfolgte durch Messung der Radioaktivität des ungebunden Y-90-C225 sowie des gebundenen Y-90-C225 im Zelllysat an einem Gammacounter (CobraTM II). Ergebnisse/Results: Der Uptake von Y-90-C225 in FaDu-Zellen zeigte eine Abhängigkeit von der Zellzahl. Der absolute Uptake von Y-90-C225 ohne Zusatz von unmarkiertem C-225 stieg bei Dosissteigerung bzw. Konzentrationserhöhung des mAb nur marginal an, die relativen Werte (bezogen auf die eingesetzte Radioaktivität) zeigten bei 1 Gy den höchsten Uptake (1 Gy: 18,63%, 6 Gy: 3,6 %). Die Kompetition von Y-90-C225 und unmarkiertem C225 um die Bindungsstellen auf der Zellmembran konnte durch die Reduzierung des Y-90-C225-Uptakes mit zunehmendem Anteil von unmarkiertem C225 an der Gesamtantikörperkonzentration nachgewiesen werden. Schlussfolgerungen/Conclusions: Bei höheren Dosen bzw. Antikörperkonzentrationen wurde kein höherer Y-90-C225-Uptake erzie

To handle the present shortage of Mo-99 in the nuclear medicine community, we offer alternative preparations for lung scintigraphy, Ga-68-labelled DOTA microspheres (DOTA MS) and labelled aerosol, analogously to well-known Technegas. Over the last three decades, several trials have been made to label HSA microspheres with Ga-68 in an efficient manner and with stability. Biodegradable albumin microspheres (HSAM) have been widely applied for lung perfusion scintigraphy (Tc-99m HSAM) or labelled with Re-188 for internal radiation therapy. We present in vitro and in vivo stability data of Ga-68 DOTA HSAM in Wistar rats. DOTA-Bz-NCS was reacted with HSAM and labelled with Ga-68 from a Ge-68/Ga-68 generator by a simple and reproducible method in high yields. The in vitro stability of Ga-68-labelled DOTA-HSAM was checked by incubation in human plasma and by challenging with DTPA. After intravenous injection into a tail vain, the Ga-68 DOTA MS was rapidly filtered from venous blood and accumulated almost quantitatively in the lungs. Radioactivity level then remains constant for at least 30 min (PET studies with microPET P4). Ga-68-labelled particles showed high in vitro stability and almost quantitative accumulation in the lungs after intravenous application. We consider that Ga-68 DOTA HSAM is a potential radiotracer for lung and other organ perfusion studies. On the other hand, for ventilation, Ga-68 Galligas aerosol for PET imaging is simple to produce in a Technegas apparatus which can produce Ga-68 eluate concentrated in acetone/HCl. It was stable in vivo (tested over 3.5 h) and allows PET imaging in man with 5–10 MBq Ga-68 only.

Introduction: Microparticles derived from denatured human serum albumin (DOTA-derivatized human serum albumin microspheres, or DOTA-HSAM) are attractive carriers of radionuclides for both therapeutic and diagnostic purposes. In this article, we describe a labeling procedure for diagnostic (Ga-68) and therapeutic (Y-90, Lu-177) radionuclides and report on the results of stability studies of these products. Methods: DOTA-HSAM was labeled in 0.5 M ammonium acetate buffer, pH 5.0, containing 0.02 mg/ml detergent. After adding the radionuclide, the mixture was shaken for 15 min at 90°C. Labeling yields and in vitro stability were determined by thin-layer chromatography. For determination of the in vivo stability of Ga-68 and Y-90 DOTA-HSAM, the particles were injected intravenously in Wistar rats. Results: Labeling yields up to 95% in the case of Ga-68 and Lu-177 were achieved. Ga-68-labeled DOTA-HSAM showed high in vitro and in vivo stability. The amount of particle-bound radioactivity of Lu-177 DOTA-HSAM declines slowly in a linear manner to approximately 72% after 13 days. For Y-90, the labeling yield decreased with increasing radioactivity level. We presume radiolysis as the reason for these findings. Conclusion: The labeling of DOTA-HSAM with different radionuclides is easy to perform. The radiation-induced cleavage of the labeled chelator together with the rather short half-life of radioactivity fixation in vivo (3.7 days) is, in our opinion, opposed to therapeutic applications of DOTA-HSAM. On the other hand, the high stability of Ga-68 DOTA-HSAM makes them an attractive candidate for the measurement of regional perfusion by PET.

13C, 18F and 123I fatty acids (FA) are used for myocardial imaging. Recently, our group showed that 99mTc-labeled "4+1" FA are extracted into the rat and guinea pig myocardium. The present study evaluates determinants of myocardial uptake and whole body biodistribution of these FA derivatives. Studies were performed with isolated perfused hearts of Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) with a FAT/CD36 deficiency, as well as with heart type FA binding protein knockout mice (H-FABP-/-) and H-FABP+/+. Eight "4+1"-99mTc- FA were applied for 3 min followed by 1 min washout. A mathematical model was used to analyze FA dynamics and binding to proteins. Whole body distribution was studied in rats with and without Tween 80. In vitro fractionation studies with 99mTc- FA assessed red blood cell uptake as well as association with plasma lipoproteins (VLDL, LDL and HDL). Myocardial extraction was 19.0-33.0 % of the infused dose in isolated WKY and 15.2-26.4 % in SHR hearts. However, H-FABP-/- showed a marked reduction of tracer extraction (2.8 ± 0.6%ID vs. 17 ± 2%ID P < 0.001). Uptake in red blood cells (< 1.2%ID) and incorporation into lipoproteins were negligible. Incubation of 99mTc-FA with albumin reduced ventricular extraction (P < 0.001) into the range of established iodinated FA tracers. Tween 80 improved the heart-to-liver ratio in the biodistribution studies. Myocardial uptake of "4+1"-99mTc- FA derivatives is dependent on H-FABP. These substances may therefore provide a new tool to specifically assess regional myocardial changes of H-FABP.

Our group has synthesized technetium labeled fatty acids which are extracted into the myocardium and sequestered due to H-FABP binding. In this paper we further address the detailed subcellular distribution and potential myocardial metabolism of "4+1" 99mTc-fatty acids (FA). Experiments were conducted using isolated hearts of wistar rats as well as wildtype and H-FABP-/--mice. Myocardium samples underwent subcellular fractionation (subsarcolemmal and intermyofibrillar mitochondria, cytosol with microsomes as well as nuclei and crude membranes) and analysis by TLC and HPLC. The largest fraction of tissue radioactivity was associated with the cytosol (79.69 ± 8.88% of the infused dose =ID). 9.07 ± 0.95% and 3.43 ± 1.38% of the ID were associated with the subsarcolemmal and the intermyofibrillar mitochondrial fractions, respectively. In the rat heart, etomoxir, an inhibitor of the carnitin-palmitoyl transferase I, did not significantly decrease radioactivity associated with the mitochondrial fractions, whereas the myocardial extraction of 123I-IPPA (13.26% vs. 49.49% in control) and the radioactivity associated with the subsarcolemmal and the intermyofibrillar mitochondrial fractions was blunted. The percentage of the ID in the mitochondrial and crude fractions increased with the number of NH-amide groups of the FA derivative. Absence of H-FABP significantly decreased the radioactivity count in the cytosolic fraction (P < 0.001). No metabolic products of a 99mTc "4+1" FA could be detected in any isolated hearts. Myocardial 99mTc "4+1" FA extraction reflects binding to H-FABP and membrane structures (including the mitochondrial membrane). However, the compounds do not undergo mitochondrial metabolism, because they do not reach the mitochondrial matrix.

INTRODUCTION Radiolabelled particles are an attractive tool in the therapy of malignancies of the liver. Hepatic tumours receive their blood supply mainly from the arterial circulation, unlike normal liver tissue which is supplied by the portal vein. The intra-arterial application of microspheres with particle size > 10 m are trapped in the capillary bed and release a local radiotoxic dose with low levels of toxicity to the normal liver (1). The aim of this study was to determine the in vitro (incubation in human plasma and DTPA-Challenge) and in vivo characteristics of radiolabelled human serum albumin (HSA) microspheres and the valuation of suitability for radiotherapeutical treatment of liver malignancies. Covalent attachment of DOTA chelators onto the surface of the spheres opens up an easy access to Y-86-radiolabelled HSA microspheres. For in vivo experiments the labelled spheres were injected into the tail vein of Wistar rats. After 1, 12, 24 and 48 h the animals were sacrificed and the radioactivity concentration of isolated organs and tissue was determined. Furthermore, to investigate the influence of the surface structure we have used three different batches of HSA microspheres (smooth to rough; diameter 20-30 µm). RESULTS AND DISCUSSION Approximately 25 DOTA molecules per molecule HSA could be attached as estimated from elemental analysis of DOTA-HSA microspheres, independently of the surface characteristics of the spheres. They were Y-86 labelled under optimized conditions in 96 ± 1 % yield. No significant differences between smooth- and rough-surfaced spheres were found. In DTPA challenge experiments 98 % of the radioactivity was still particle-associated after 24 hours incubation at 37 °C. In vitro studies in human plasma resulted in 95 ± 4 % particle-associated radioactivity after 48 h incubation at 37 °C. In the in vivo experiments radiolabelled smooth and rough microspheres were completely trapped in the lungs, which served in our experiments as mod

Our group previously synthesized 99mTc-labeled fatty acids suitable for myocardial metabolism and flow imaging. In this set of experiments 28 new analogues were synthesized according to the “4+1” mixed ligand approach with some specific differences. Conventional “4+1” 99mTc-fatty acids are built in the sequence Tc-chelate, alkyl chain and carboxylic group. We developed compounds with a new design with the sequence carboxylic group, alkyl chain, Tc-chelate, and lipophilic tail. Therefore the 99mTc-chelate was transferred to a more central position of the compound, aiming towards an improved myocardial profile and an accelerated liver clearance. In this context several functional groups incorporated in the lipophilic tail section were tested to evaluate their influence on the compound’s character. In addition to biodistribution studies in vivo the myocardial first-pass extraction of the compounds was tested in an isolated Langendorff rat heart model. A satisfactory myocardial uptake up to 20% of the injected dose (% ID) in the perfused heart and a fast liver clearance in vivo with only 0.29% ID/g at 60 min post injection demonstrates that the induced molecular modifications affect the kinetics of 99mTc-radiolabeled fatty acid compounds favorably. From the data set rules for estimating the biodistribution of fatty acids tracers are deduced.

Ziel/Aim: Radiolabelled fatty acids displaying efficient myocardial uptake and adequate retention are attractive candidates for the clinical evaluation of regional discrepancies in the cardiac metabolism and energy turnover (1). The aim of this work is the substitution of the accelerator produced radiolabel I-123 for FA against the low price generator radionuclide Tc-99m (2). Methodik/Methods: Synthesis and physico-chemical characterization of the new Tc-99m labelled FA derivatives were followed by in vitro and in vivo experiments for their biological evaluation. Experiments on isolated perfused rat hearts provide a simple and well reproducible approach to detect promising Tc-99m compounds for biodistribution experiments. Biodistribution and metabolism studies in male Wistar rats after a single intravenous injection of various Tc-99m-labelled FA were investigated (3). Ergebnisse/Results: A very rapid disappearance of Tc-99m tracer concentration from the blood circulation was observed for all compounds. C11S FA showed the highest uptake with 2.0% ID/g tissue (alternatively 3.6% by addition of Tween 80) in the rat heart. Liver and kidneys are mainly responsible for fast removal of Tc-99m derivatives from the circulation. At 5 min after injection, approximately 50% of the injected dose is liver-associated. A substantial fraction is cleared by the kidneys and approximately 6% of the injected dose is kidney-associated at 5 min p.i. Overall, fast systemic clearance of Tc-99m species after injection of Tc-99m labelled FA in the rat is explained by the rapid hepatobiliary and renal elimination of Tc-99m species. On the other hand, the low mean accumulation of Tc-99m compounds in the thyroid gland (<0.07 %ID at 5 min; <0.07 %ID at 60 min) and the stomach (<1.7 %ID at 5 min; < 3.9% at 60 min), respectively, which express the sodium-iodide transporter is indicative of a very low abundance of pertechnetate in vivo. Schlussfolgerungen/Conclusions: Experiments on isolated perfuse

Aim: The preparation and stability of a new 188Re-S4-complex (S4 = (1-aza-18-crown-6)(O)C-C(SH)-C(SH)-C(O)NH-(CH2)3–NH-(CH2)3–NHC(O)-C(SH)-C(SH)-C(O)(1-aza-18-crown-6) was studied at therapeutic relevant radioactive concentrations. The results were compared with 188Re-MAG3 (MAG3 = mercaptoacetyltriglycine) and 188Re-DMSA preparations (DMSA = dimercaptosuccinic acid) performed with the same highly concentrated [188Re]perrhenate solution (12-15 GBq/ml). Methods: The 188Re complexes were prepared by direct reduction of perrhenate (188Re-S4-complex) as well as via the 188Re-EDTA precursor complex (188Re-MAG3, 188Re-DMSA). The preparations were stabilised with 15 mg of ascorbic acid and analysed after 1, 2, and 24 hours by TLC and HPLC. Additionally, in vitro and in vivo stability studies were performed with the purified complexes. Results: After stabilisation with 15 mg of ascorbic acid, all of the complexes were nearly stable under nitrogen for hours, and only 2-8 % of perrhenate was observed after 24 h. In contrast, only the 188Re-S4 complex was completely stable in vitro and in all investigated in vivo samples after separation of ligand excess and reducing agent by HPLC. Conclusion: The bridging amine group or free carboxylic groups of the S4-ligand framework make available reactive positions for coupling biomolecules to the chelate. Thus it appears that the new 188Re-S4 complexes offer the possibility of stable and high specific activity labelling of biomolecules for therapeutic application.

In an effort to develop 99mTc-labelled fatty acids (FAs) for myocardial metabolism and flow imaging, several 99mTc analogues according to the ‘3+1’ and the ‘4+1’-mixed ligand approach were synthesized and myocardial extraction was evaluated in non-working isolated guinea pig hearts. Biodistribution patterns in guinea pigs were determined exemplarily of one FA analogue.

Ziel/Aim: Gegenstand der Arbeiten war die In-vitro- und In-vivo-Charakterisierung von radioaktiv beladenen Humanserumalbumin-Mikrosphären (HSAM). Ziel war dabei die Bewertung der Partikel hinsichtlich ihrer Eignung zur radiotherapeutischen Behandlung von Krebserkrankungen der Leber. Methodik/Method: An vorgefertigte HSAM (3 Chargen mit unterschiedlicher Oberflächenrauigkeit; mittlerer Durchmesser 25 µm) wurden oberflächlich DOTA-Chelatoren kovalent gebunden. Anschließend erfolgte die Markierung mit Y-86, indem die DOTA-HSAM in einer Lösung des Radionuklides suspendiert wurden. Die In-Vitro-Charakterisierung der markierten Partikel erfolgte durch Inkubation in Humanplasma und Challenge mit DTPA. Die In-vivo-Stabilität der Y-86-DOTA-HSAM wurde bestimmt durch Bioverteilungsstudien in gesunden Wistarratten. Nach intravenöser Applikation der Y-86 markierten Partikel kommt es zum vollständigen Trapping der Mikrosphären in der Lunge, die in useren Untersuchungen als Zielorgan diente. Aus dem Verschwinden der Radioaktivität aus der Lunge können Rückschlüsse auf die Stabilität der Markierung und damit verbunden auf die Stabilität der HSA-Partikel gezogen werden. Ergebnisse/Results: DOTA konnte in Form eines Isothiocyanat-Derviates an Oberflächenaminogruppen der HSAM gebunden werden. Unter optimierten Bedingungen enthielten 1 mg Mikrosphären 2 x 10-7 mol DOTA. Die Markierung der Partikel mit Y-86 gelang in hohen reproduzierbaren Ausbeuten (96 ± 1 %, n = 7) nach 15minütigem Schütteln der suspendierten DOTA-HSAM in Acetatpuffer (pH = 6,5). Die markierten Mikrosphären zeigten hohe Stabilität in Humanplasma und in Anwesenheit von DTPA. Bei den Bioverteilungsstudien fanden wir große Unterschiede zwischen den einzelnen Mikrosphärenchargen mit unterschiedlicher Oberflächenrauigkeit. Partikel mit glatter Oberfläche zeigten die höchste Stabilität. Hier war das Aktivitätsniveau in der Lunge über 48 Stunden annähernd konstant. Schlussfolgerungen/Consequences: Die Kopplung von

The preparation and stability of a novel 188Re-S4-complex (S4 = bridged DMSA ligand) was studied at therapeutic relevant radioactive concentrations. The results were compared with an 188Re-MAG3 and an 188Re-DMSA preparation which were obtained from 188Re-EDTA by a combined re-oxidation/ligand exchange reaction. The highly concentrated [188Re]perrhenate solution (12-15 GBq/ml) used for preparations was received by successive elution of several 188W/188Re generators connected in series. The preparations were performed under nitrogen, stabilized by adding 15 mg of ascorbic acid and analyzed after 1, 2, and 24 hours by TLC and HPLC. Only about 8 % of perrhenate were detected after 24 h in the 188Re-S4-complex samples. The 188Re-DMSA preparations were only marginally re-oxidized during this time, whereas the stabilized 188Re-MAG3 samples remained nearly stable. Additionally, in vitro and in vivo stability studies at radioactive concentrations of 30-50 MBq/ml were performed with all the complexes. Therefore, the preparations were purified by HPLC and incubated in phosphate buffer as well as rat and human plasma. The in-vivo-stability was determined in rats. Samples of plasma, urine, liver, and intestine were taken off 1 h after application of the 188Re complex solutions. Results from these studies demonstrated that only the 188Re-S4-complex was completely stable in vitro and in all investigated in vivo samples.

Technetium-labelled fatty acids intended for myocardial metabolism imaging and the respective rhenium model complexes were synthesized according to the “4+1” mixed-ligand approach and investigated in vitro and in vivo. In “4+1” mixed-ligand Tc(III) and Re(III) complexes the metal is coordinated by both a tripodal tetradentate thiolato ligand and by a monodentate isocyano ligand bearing the biomolecule. High heart extraction in perfused heart studies (up to 26% ID) and noticeable heart uptake of the 99mTc tracer in vivo being in the order of 2% ID/g at 5 min p.i., accompanied by a good heart to blood ratio of 8, confirms that the new Tc-compounds are mimics of fatty acids.

Introduction: In an effort to develop technetium-labelled fatty acid analogues for myocardial metabolism imaging rhenium model complexes and their 99mTc analogues were synthesized according to the '4+1' mixed-ligand approach and investigated in vitro and in vivo. Materials and Methods: The rhenium model complexes were completely characterised by NMR, IR, MS, EA and the geometrical impact of the chelate unit on the integrity of the fatty acid head structure was determined by single crystal X-ray analyses. To estimate the diagnostic value of the 99mTc-labelled fatty acids the compounds were investigated using the isolated constant-flow-perfused guinea pig heart model (see presentation A. Heintz et al.), in cell-uptake experiments and in biodistribution studies using male Wistar rats (5 - 6 weeks old, body weight 151 ± 15 g). Results: The new fatty acid tracers contain the metal core in the oxidation states +3, well-wrapped in a trigonal-bipyramidal coordination moiety which is attached at the omega-position of a fatty acid chain. This structural feature is considered to be a good imitate of the well-established iodinated phenyl fatty acid. The formation of the rhenium models was accomplished by ligand exchange reactions using different pre-formed rhenium precursors. Noticable heart uptake of the 99mTc tracers being in the order of 2 % ID/g 5´p.i. and accompanied by a good heart to blood ratio of 8.6 confirms the remarkably results of the perfused heart experiments. A significant time-dependent cell uptake in HepG2 cells is shown for two representatives. Conclusion: Further species such as mice and guinea pigs will be involved to characterise in vivo patterns of those derivatives, that show high extraction rates in the isolated constant-flow-perfused guinea pig heart model. While the tracers are superior to other described Tc-fatty acid imitates with regard to good heart to blood ratios, heart to liver ratio has to be improved. For this

Aim: Determination of the biological effect of the alpha emiter 211At on cellular level as well as the assessment of dosimetric data in a tumor model in vivo. Methods: Transplantation of malignant ascitic cells in mice intraperitoneally and estimation of tumor characteristics (doubling time of the cells, mean survival of the animals following an i.p. applicatin of a defined tumor cell number). 211At labelled human serum albumine microspheres B-20 (MSP) of different activity were injected into tumor bearing mice intraperitoneally. The effectiveness of the therapy was evaluated by means of determination of the duration of cell cycle arrest as well as the microscopic analysis of the rate of abnormal mitotic cells due to radiation induced damage. Furthermore, dose dependence of survival was evaluated. Results: Three days following the intraperitoneally application of 8 x 106 tumor cells, 50 - 600 kBq 211At-MSP were applied into the abdominal cavity. Derived from the volume of ascites at this time and the administered activity, dese calculations were performed. An activity of 50 kBq caused a dose of 0.84 Gy. The increase of radiation induced effect on ascitic tumor cells was correlated with the dose. Between the duration of the cell cycle arrest and the administered activity, a dirctly proportional correlation was found. The mean survival of non treated animals was 16.9 ± 3.7 days. Teh prolongation of the survival was proportional to the activity administered. Using a dosage of 10 Gy, five animals out of 16 survived. Conclusion: Therapy of malignant ascitic cells using 211At-MSP was effective in vivo. For tumor therapy, teh alpha emitter 211At represents a highly effective alternative to usually employed beta emitters.

Aufgabe der Erfindung ist es Substanzen vorzuschlagen, die zur Diagnose und Therapie von Tumoren in der Nuklearmedizin geeignet sind und ein Verfahren zur Herstellung der Substanzen anzugeben.