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Roland Haubner

Publications and source records attributed to Roland Haubner.

At least 19 recordsLinked to original sources
PubMed2006

Noninvasive characterization of myocardial molecular interventions by integrated positron emission tomography and computed tomography.

OBJECTIVES: We sought to investigate the usefulness of integrated positron emission tomography (PET) and computed tomography (CT) for in vivo characterization of an angiogenesis-directed molecular intervention. BACKGROUND: Controversies about the effectiveness of molecular therapies for cardiovascular disease have prompted the need for more powerful noninvasive imaging techniques. METHODS: In a model of regional adenoviral transfer of the VEGF(121) gene to myocardium of healthy pigs, PET-CT using multiple molecular-directed radiotracers was employed. RESULTS: Two days after gene transfer, successful transgene expression was noninvasively confirmed by a reporter probe targeting co-expressed HSV1-sr39tk reporter gene. The CT-derived ventricular function and morphology remained unaltered (left ventricular ejection fraction 57 +/- 5% in adenovirus-injected animals vs. 53 +/- 5% in controls; p = 0.36). Increased regional perfusion was identified in areas overexpressing VEGF (myocardial blood flow during adenosine-induced vasodilation 1.47 +/- 0.49 vs. 1.14 +/- 0.27 ml/g/min in remote areas; p = 0.01), corroborating in vivo effects on microvascular tone and permeability. Finally, regional angiogenesis-associated alpha(v)beta3 integrin expression was not enhanced, suggesting little contribution to the perfusion increase. Fusion of CT morphology and tracer-derived molecular signals allowed for accurate regional localization of biologic signals. Findings were validated by control vectors, sham-operated animals, and ex vivo tissue analysis. CONCLUSIONS: Integrated PET-CT has the potential to dissect cardiovascular biologic mechanisms from gene expression to physiologic function and morphology. The VEGF overexpression in healthy myocardium increases myocardial perfusion without significant up-regulation of alpha(v)beta3 integrin adhesion molecules early after the intervention.

Animals↗
PubMed2006

Positron emission tomography using [18F]Galacto-RGD identifies the level of integrin alpha(v)beta3 expression in man.

PURPOSE: The integrin alpha(v)beta3 plays a key role in angiogenesis and tumor cell metastasis and is therefore an important target for new therapeutic and diagnostic strategies. We have developed [18F]Galacto-RGD, a highly alpha(v)beta3-selective tracer for positron emission tomography (PET). Here, we show, in man, that the intensity of [18F]Galacto-RGD uptake correlates with alpha(v)beta3 expression. EXPERIMENTAL DESIGN: Nineteen patients with solid tumors (musculoskeletal system, n = 10; melanoma, n = 4; head and neck cancer, n = 2; glioblastoma, n = 2; and breast cancer, n = 1) were examined with PET using [18F]Galacto-RGD before surgical removal of the tumor lesions. Snap-frozen specimens (n = 26) were collected from representative areas with low and intense standardized uptake values (SUV) of [18F]Galacto-RGD. Immunohistochemistry was done using the alpha(v)beta3-specific antibody LM609. Intensity of staining (graded on a four-point scale) and the microvessel density of alpha(v)beta3-positive vessels were determined and correlated with SUV and tumor/blood ratios (T/B). RESULTS: Two tumors showed no tracer uptake (mean SUV, 0.5 +/- 0.1). All other tumors showed tracer accumulation with SUVs ranging from 1.2 to 10.0 (mean, 3.8 +/- 2.3; T/B, 3.4 +/- 2.2; tumor/muscle ratio, 7.7 +/- 5.4). The correlation of SUV and T/B with the intensity of immunohistochemical staining (Spearman's r = 0.92; P < 0.0001) as well as with the microvessel density (Spearman's r = 0.84; P < 0.0001) were significant. Immunohistochemistry confirmed lack of alpha(v)beta3 expression in normal tissue (benign lymph nodes, muscle) and in the two tumors without tracer uptake. CONCLUSIONS: Molecular imaging of alpha(v)beta3 expression with [18F]Galacto-RGD in humans correlates with alpha(v)beta3 expression as determined by immunohistochemistry. PET with [18F]Galacto-RGD might therefore be used as a new marker of angiogenesis and for individualized planning of therapeutic strategies with alpha(v)beta3-targeted drugs.

Adult↗
PubMed2006

Alphavbeta3-integrin imaging: a new approach to characterise angiogenesis?

OVERVIEW: The field of angiogenesis research is one of the most rapidly growing biomedical disciplines. Great efforts are being made to develop anti-angiogenesis drugs for treatment of cancer as well as non-oncological diseases. Thus, imaging techniques allowing non-invasive monitoring of corresponding molecular processes will be of great interest. One target structure involved in the angiogenic process is the integrin alphavbeta3, which mediates the migration of activated endothelial cells during vessel formation. MATERIALS AND METHODS: A variety of radiolabelled RGD peptides have been introduced for monitoring of alphavbeta3 expression using nuclear medicine tracer techniques. OBJECTIVES: This review discusses tracer development and highlights some strategies for tracer optimisation. It summarises the preclinical and clinical data and discusses the potential of this class of tracer to characterise angiogenesis.

Animals↗
PubMed2006

[99mTc]HYNIC-RGD for imaging integrin alphavbeta3 expression.

There has been increasing interest in peptides containing the Arg-Gly-Asp (RGD) sequence for targeting of alpha(v)beta(3) integrins to image angiogenesis. [(18)F]Galacto-RGD has been successfully used for positron emission tomography applications in patients. Here we report on the preclinical characterization of a (99m)Tc-labeled derivative for single-photon emission computed tomography. c(RGDyK) was derivatized with HYNIC at the amino group of the lysine [c(RGDyK(HYNIC)) or HYNIC-RGD]. (99m)Tc labeling was performed using coligands (tricine and EDDA), as well as (99m)Tc(CO)(3)(H(2)O)(3). Radiolabeled peptides were characterized with regard to lipophilicity, protein binding and stability in buffer, serum and tissue homogenates. Integrin receptor activity was determined in internalization assays using alpha(v)beta(3)-receptor-positive M21 and alpha(v)beta(3)-receptor-negative M21L melanoma cells. Biodistribution was evaluated in normal and nude mice bearing M21, M21L and small cell lung tumors. HYNIC-RGD could be labeled at high specific activities using tricine, tricine-trisodium triphenylphosphine 3,3',3''-trisulfonate (TPPTS), tricine-nicotinic acid (NA) or EDDA as coligands. [(99m)Tc]EDDA/HYNIC-RGD, [(99m)Tc]tricine-TPPTS/HYNIC-RGD and [(99m)Tc]tricine-NA/HYNIC-RGD showed protein binding (<5%) considerably lower than [(99m)Tc](CO)(3)/HYNIC-RGD and [(99m)Tc]tricine/HYNIC-RGD. [(99m)Tc]EDDA/HYNIC-RGD revealed high in vitro stability accompanied by low lipophilicity with a log P value of -3.56, comparable to that of [(18)F]Galacto-RGD. In M21 cells for this compound, the highest level of specific and rapid cell uptake (1.25% mg protein(-1)) was determined. In vivo, rapid renal excretion, low blood retention, low liver and muscle uptakes and low intestinal excretion 4 h postinjection were observed. Tumor uptake values were 2.73% ID/g in M21 alpha(v)beta(3)-receptor-positive tumors versus 0.85% ID/g in receptor-negative tumors 1 h postinjection. Small cell lung tumors could be visualized using gamma camera imaging. [(99m)Tc]EDDA/HYNIC-RGD shows encouraging properties to target alpha(v)beta(3) receptors in vivo with high stability and favorable pharmacokinetics. Tumor uptake studies showed specific targeting of alpha(v)beta(3)-receptor-positive tumors with tumor-to-organ ratios comparable to those of [(18)F]Galacto-RGD.

Animals↗
PubMed2006

PET-based human dosimetry of 18F-galacto-RGD, a new radiotracer for imaging alpha v beta3 expression.

UNLABELLED: (18)F-Galacto-RGD is a new tracer for PET imaging of alpha v beta3, a receptor involved in a variety of pathologic processes including angiogenesis and metastasis. Our aim was to study the dosimetry of (18)F-galacto-RGD in humans. METHODS: Eighteen patients with various tumors (musculoskeletal tumors [n = 10], melanoma [n = 5], breast cancer [n = 2], or head and neck cancer [n = 1]) were examined. After injection of 133-200 MBq of (18)F-galacto-RGD, 3 consecutive emission scans from the thorax to the pelvis were acquired at 6.7 +/- 2.9, 35.6 +/- 7.6, and 70.4 +/- 12.2 min after injection. Blood samples (n = 4) for metabolite analysis were taken 10, 30, and 120 min after injection. The OLINDA 1.0 program was used to estimate the absorbed radiation dose. RESULTS: Reversed-phase high-performance liquid chromatography of serum revealed that more than 95% of tracer was intact up to 120 min after injection. (18)F-Galacto-RGD showed rapid clearance from the blood pool and primarily renal excretion. Background activity in lung and muscle tissue was low (percentage injected dose per liter at 71 min after injection, 0.56 +/- 0.15 and 0.69 +/- 0.25, respectively). The calculated effective dose was 18.7 +/- 2.4 microSv/MBq, and the highest absorbed radiation dose was in the bladder wall (0.22 +/- 0.03 mGy/MBq). CONCLUSION: (18)F-Galacto-RGD demonstrates high metabolic stability, a favorable biodistribution, and a low radiation dose. Consequently, this tracer can safely be used for noninvasive imaging of molecular processes involving the alpha v beta3 integrin and for the planning and monitoring of therapeutic approaches targeting alpha v beta3.

Adult↗
PubMed2005

Noninvasive visualization of the activated alphavbeta3 integrin in cancer patients by positron emission tomography and [18F]Galacto-RGD.

BACKGROUND: The integrin alphavbeta3 plays an important role in angiogenesis and tumor cell metastasis, and is currently being evaluated as a target for new therapeutic approaches. Several techniques are being studied to enable noninvasive determination of alphavbeta3 expression. We developed [(18)F]Galacto-RGD, a (18)F-labeled glycosylated alphavbeta3 antagonist, allowing monitoring of alphavbeta3 expression with positron emission tomography (PET). METHODS AND FINDINGS: Here we show by quantitative analysis of images resulting from a small-animal PET scanner that uptake of [(18)F]Galacto-RGD in the tumor correlates with alphavbeta3 expression subsequently determined by Western blot analyses. Moreover, using the A431 human squamous cell carcinoma model we demonstrate that this approach is sensitive enough to visualize alphavbeta3 expression resulting exclusively from the tumor vasculature. Most important, this study shows, that [(18)F]Galacto-RGD with PET enables noninvasive quantitative assessment of the alphavbeta3 expression pattern on tumor and endothelial cells in patients with malignant tumors. CONCLUSIONS: Molecular imaging with [(18)F]Galacto-RGD and PET can provide important information for planning and monitoring anti-angiogenic therapies targeting the alphavbeta3 integrins and can reveal the involvement and role of this integrin in metastatic and angiogenic processes in various diseases.

Animals↗
PubMed2005

Non-invasive imaging of cardiac transgene expression with PET: comparison of the human sodium/iodide symporter gene and HSV1-tk as the reporter gene.

PURPOSE: Genes encoding for intracellular enzymes or transmembrane proteins are suitable as reporters, but may differ in terms of applicability for cardiac imaging. The aim of this study was to compare the human sodium iodide symporter gene (hNIS) with the herpes simplex virus type 1 thymidine kinase gene (HSV1-tk) as the reporter gene in non-invasive imaging of cardiac transgene expression with positron emission tomography (PET). METHODS: Equal doses of adenoviral vectors encoding for hNIS, wild-type HSV1-tk, mutant HSV1-sr39tk or LacZ as the control gene were directly injected into the myocardium of 34 animals. Two days later, dynamic PET was performed with a clinical scanner, using reporter probes specific for the respective reporter gene. Imaging with (13)N-ammonia was also performed to identify cardiac regions of interest. RESULTS: Kinetics differed significantly: (124)I as the probe for hNIS showed rapid early uptake, remaining stable over time. Maximal myocardial concentration was 3.61+/-1.15%. The nucleoside (18)F-FHBG, as the specific probe for HSV1-sr39tk, showed increasing uptake over time, but maximal accumulation was significantly lower (1.45+/-0.54%, P=0.0009). (124)I-FIAU, as the specific probe for wild-type HSV1-tk, showed early uptake with subsequent washout. Maximal accumulation was lowest (0.63+/-0.23%, P<0.0001). Post-mortem analysis by autoradiography and gamma counting confirmed the in vivo data. CONCLUSION: Reporter genes encoding for transporter proteins such as hNIS are an attractive alternative to overexpression of intracellular enzymes for cardiac gene product imaging. hNIS yielded higher signal intensity and imaging contrast for PET than did HSV1-tk and HSV1-sr39tk. Therefore, this approach may be preferable for the future monitoring of cardiac gene- or cell-based therapy.

Animals↗
PubMed2005

Non-invasive imaging of glycoprotein VI binding to injured arterial lesions.

Glycoprotein VI (GPVI) is the major platelet collagen receptor and plays a critical role in the process of thrombosis at sites of atherosclerotic lesions. This study evaluates the feasibility of radiolabeled soluble GPVI to identify injured arterial lesions. Radiolabeling was carried out using the iodogen method and resulted in the radioiodinated GPVI in radiochemical yields between 97-100%. The biodistribution of [(125)I]GPVI was determined in normal mice and demonstrated a blood clearance halftime of approximately 5.5 hours. Vascular lesions were induced in the carotid artery in wild type and ApoE(-/-)mice. Immediately after injury radioiodinated GPVI was injected intravenously. Binding of [(123)I]GPVI to carotid lesions was assessed by szintigraphic in vivo imaging. Carotid arteries were explanted for ex vivo autoradiography and histological characterization of the lesion. In vivo and ex vivo imaging revealed substantial accumulation of radioiodinated GPVI in the injured artery wall, with a ratio of lesion to control vessel of 3:1 and 7:1, respectively. Because GPVI is the critical collagen receptor that mediates platelet adhesion to vascular lesions, soluble radiolabeled GPVI may be an agent for non-invasive imaging of thrombogenic thus, vulnerable atherosclerotic plaques.

Animals↗
PubMed2005

Myocardial kinetics of reporter probe 124I-FIAU in isolated perfused rat hearts after in vivo adenoviral transfer of herpes simplex virus type 1 thymidine kinase reporter gene.

UNLABELLED: Reporter gene imaging holds promise for noninvasive monitoring of cardiac gene therapy. We recently demonstrated that (124)I-labeled 2'-fluoro-2'-deoxy-5'-iodo-1beta-d-arabinofuranosyluracil ((124)I-FIAU) is suitable for PET of myocardial expression of herpes simplex virus type 1 thymidine kinase reporter gene (HSV1-tk). In contrast to previous studies in tumors, early specific uptake was followed by rapid washout. Myocardial kinetics of (124)I-FIAU are still poorly understood. This study aimed at a further investigation under controlled conditions using an isolated heart perfusion model. METHODS: Male Wistar rats underwent transthoracic regional injection of replication-defective adenovirus (2.5 x 10(9) plaque-forming units) containing either HSV1-tk (n = 16) or LacZ reporter gene (n = 15) into the inferior wall. Nonmanipulated rats (n = 5) served as further controls. Hearts were excised 2 d later and perfused according to the Langendorff technique with (124)I-FIAU-containing buffer (15 min, followed by 30 min of nonradioactive perfusion). Experiments were performed under baseline conditions and in the presence of thymidine (competitive substrate) or fludarabine (in vitro inhibitor of 5'-nucleotidase). Time-activity curves were acquired by external coincident detectors. The myocardial rate of (124)I-FIAU uptake (K(i)), clearance rate (K(o)), and volume of distribution (V(d) = K(i)/K(o)) were calculated. Subsequently, hearts were subjected to gamma-counting, followed by microtome slicing and autoradiography. RESULTS: The V(d) from Langendorff perfusion significantly correlated with final whole-heart tracer retention (r = 0.88, P = 0.019) and the autoradiographic area of regional myocardial activity (r = 0.89, P = 0.016). HSV1-tk hearts showed higher K(i) and V(d) of (124)I-FIAU compared with that of controls (P < 0.001) and detectable but slower washout compared with that of the LacZ group (P < 0.01). Addition of thymidine to the perfusate inhibited myocardial uptake of (124)I-FIAU by reducing V(d) and K(i) in HSV1-tk and LacZ hearts compared with the baseline. Addition of fludarabine did not result in the expected reduction of washout in HSV1-tk hearts due to inhibition of 5'-nucleotidases (which may dephosphorylate (124)I-FIAU monophosphate). It acted as an uptake inhibitor similar to thymidine, reducing V(d) in HSV1-tk hearts. CONCLUSION: Assessment of specific reporter probe kinetics after regional in vivo reporter gene transfer is feasible using the isolated perfused rat heart preparation. This model allows one to study the effects of pharmacologic interventions and may refine understanding of the reporter probe signal for in vivo imaging. Different nucleoside analogs significantly inhibit (124)I-FIAU uptake, emphasizing the importance of transporter mechanisms for reporter probe kinetics.

Adenoviridae↗
PubMed2005

Imaging of delayed-type hypersensitivity reaction by PET and 18F-galacto-RGD.

UNLABELLED: Radiolabeled cyclic peptides containing the amino acid sequence arginine-glycine-aspartate (RGD peptides) have successfully been used to image the expression of the alpha(v)beta(3) integrin in malignant tumors. However, the alpha(v)beta(3) integrin also plays an important role in angiogenesis induced by chronic inflammatory processes. Therefore, the aim of this study was to evaluate whether radiolabeled RGD peptides may also be used to assess alpha(v)beta(3) expression in inflammatory diseases. We studied a hapten-induced delayed-type hypersensitivity reaction (DTHR) as a model for inflammatory processes, since DTHRs are involved in many human autoimmune disorders. METHODS: The abdominal skin of mice was sensitized by application of 2,4,6-trinitrochlorobenzene (TNCB). One week later, a DTHR was elicited by challenging the right ear with TNCB. Application of TNCB was then repeated every 48 h to induce chronic skin inflammation. Small-animal PET and autoradiography with the alpha(v)beta(3) ligands (18)F-galacto-RGD and (125)I-gluco-RGD were performed at various times after TNCB application. The time course of tracer uptake by the treated ears was compared with histologic skin changes. RESULTS: The first challenge with TNCB caused, within 12 h, an acute inflammatory response with dense dermal infiltrates of polymorphonuclear leukocytes and lymphocytes. However, autoradiography revealed no significant increase in (125)I-gluco-RGD uptake at that time (mean uptake ratio for treated ear to untreated ear, 1.02 +/- 0.1 [SD]). Further challenges with TNCB resulted in chronic skin inflammation with markedly increased small-vessel density in the ear tissue. This was paralleled by a continuous increase in uptake of (125)I-gluco-RGD. After 13 challenges, the uptake ratio had increased to 2.30 +/- 0.27 (P < 0.005 compared with baseline). Enhanced uptake of radiolabeled RGD peptides in chronic inflammation was also demonstrated noninvasively by PET with (18)F-galacto-RGD. Pretreatment of the mice with nonradiolabeled cyclic peptide c(RGDfV) almost completely blocked uptake of (18)F-galacto-RGD by the challenged ear, thus confirming the specificity of tracer uptake. CONCLUSION: Radiolabeled RGD peptides allow a noninvasive assessment of alpha(v)beta(3) expression in inflammatory processes. PET with (18)F-galacto-RGD might become a powerful tool to distinguish between the acute and chronic phases of T cell-mediated immune responses and may represent a new biomarker for disease activity in autoimmune disorders.

Animals↗
PubMed2005

Biodistribution and pharmacokinetics of the alphavbeta3-selective tracer 18F-galacto-RGD in cancer patients.

UNLABELLED: (18)F-Galacto-RGD has been developed for PET of alpha(v)beta(3) integrin expression, a receptor involved in, for example, angiogenesis and metastasis. Our aim was to study the kinetics and biodistribution of (18)F-Galacto-RGD in cancer patients. METHODS: Nineteen patients with metastases of malignant melanoma (n = 7), sarcomas (n = 10), or osseous metastases (n = 2) were examined. After injection of 133-200 MBq (18)F-Galacto-RGD, 3 consecutive emission scans from the pelvis to the thorax or dynamic emission scans of the tumor over 60 min, followed by 1 static emission scan of the body, were acquired. Time-activity curves and standardized uptake values (SUVs) were derived by image region-of-interest analysis with image-based arterial input functions. Compartmental modeling was used to derive the distribution volume for muscle tissue and tumors. RESULTS: (18)F-Galacto-RGD showed rapid blood clearance and primarily renal excretion. SUVs in tumors ranged from 1.2 to 9.0. Tumor-to-blood and tumor-to-muscle ratios increased over time, with peak ratios of 3.1 +/- 2.0 and 7.7 +/- 4.3, respectively, at 72 min. The tumor kinetics were consistent with a 2-tissue compartment model with reversible specific binding. Distribution volume values were, on average, 4 times higher for tumor tissue (1.5 +/- 0.8) than those for muscle tissue (0.4 +/- 0.1). The data suggest that there was only minimal free and bound (specific or nonspecific) tracer in muscle tissue. CONCLUSION: (18)F-Galacto-RGD demonstrates a highly favorable biodistribution in humans with specific receptor binding. Most important, this study shows that (18)F-Galacto-RGD allows visualization of alpha(v)beta(3) expression in tumors with high contrast. Consequently, this tracer offers a new strategy for noninvasive monitoring of molecular processes and may supply helpful information for planning and controlling of therapeutic approaches targeting the alpha(v)beta(3) integrin.

Adult↗
PubMed2004

Use of the norepinephrine transporter as a reporter gene for non-invasive imaging of genetically modified cells.

BACKGROUND: The norepinephrine transporter (NET) is a high-affinity transporter for catecholamines. Its expression is almost exclusively restricted to the sympathetic nervous system. In this study we evaluated whether the NET can be used as a reporter gene for non-invasive imaging of genetically modified cells with radiolabeled probes. METHODS: Human A431, HT1080 and murine CMS-5 cells were retrovirally transduced with bovine NET cDNA. Transduced and parental cells were incubated in vitro with [(131)I]meta-iodobenzylguanidine ([(131)I]MIBG). The specificity of tracer uptake was determined by adding the NET inhibitor imipramine. Rat PC12 cells served as positive controls. Parental and A431NET cells were xenotransplanted into nude mice and tumor uptake of [(123)I]MIBG in vivo was determined after tracer administration. RESULTS: In vitro stably transduced cells showed a 66- to 120-fold higher [(131)I]MIBG uptake than parental cells. Incubation with imipramine reduced [(131)I]MIBG uptake of transduced cells to the level found in parental cells. More than 70% of the initial radioactivity was retained in all transduced cell lines after 2 h incubation with tracer-free medium. [(131)I]MIBG uptake in PC12 cells, which express the NET endogenously, was 20- to 28-fold lower than in transduced cells. In vivo, A431NET tumors demonstrated a 33-fold higher [(123)I]MIBG uptake than parental tumors. Gamma camera images 24 h after tracer injection showed no tracer uptake in parental A431 tumors, but clear images of A431NET tumors. CONCLUSIONS: Transduction of tumor cells with NET cDNA causes highly specific uptake and significant retention of catecholamine analogs in vitro and in vivo. These characteristics make the NET suitable as a reporter gene for non-invasive monitoring of gene transfer.

3-Iodobenzylguanidine↗
PubMed2004

Radiolabeled tracers for imaging of tumor angiogenesis and evaluation of anti-angiogenic therapies.

A variety of therapeutic strategies in oncology are focused on the inhibition of tumor-induced angiogenesis. Thus, there is a keen interest in methods which allow non-invasive monitoring of molecular targets involved in angiogenesis which would support information for planning and controlling corresponding therapies. Moreover, such techniques would provide an insight into the formation of new sprouting blood vessels, the involved processes and regulatory mechanisms in patients. At the moment, development of radiotracer based techniques is mainly concentrated on three different targets which include peptidic and non-peptidic alpha v beta 3-integrin binding antagonists, matrix metalloproteinase inhibitors and single chain anti-fibronectin antibody fragments. Development of radiolabeled MMP inhibitors is based on either the decapeptide Cys-Thr-Thr-His-Trp-Gly-Phe-Thr-Leu-Cys resulting from a phage display library or small molecular weight compounds. The in vitro data for these tracers are very promising. However, more detailed in vivo data are necessary to evaluate the potency of MMP-inhibitors for in-vivo imaging. The radiolabelled anti-ED-B single chain antibody fragment scFv L-19 shows selective accumulation in the tumor vasculature in a murine tumour model. In a first patient study a selective localisation of the (123)I-labeled tracer in lesions of different tumours was found. On the basis of the lead structure cyclo(-Arg-Gly-Asp-dPhe-Val) a variety of different radiolabeled RGD-peptides has been developed for the non-invasive determination of the alpha v beta 3 expression. These developments include peptides labeled with minimum structural alteration, peptide carbohydrate conjugates, peptidomimetics based on the RGD-structure as well as heterodimeric, homodimeric and homotetrameric ligand systems. Many of the tracers show high alpha v beta 3-affinity and selectivity in vitro and receptor selective tumour accumulation with high image contrast in different murine tumour models. Further studies have to demonstrate that this approach can be translated to clinical settings allowing visualisation of alpha v beta 3-positive tumours and alpha v beta 3 expression during tumour-induced angiogenesis in patients.

Angiogenesis Inhibitors↗
PubMed2004

Coexpression of herpesviral thymidine kinase reporter gene and VEGF gene for noninvasive monitoring of therapeutic gene transfer: an in vitro evaluation.

UNLABELLED: Coexpression of a reporter gene and a therapeutic gene may allow for noninvasive monitoring of cardiac gene therapy. We sought to evaluate the usefulness of an adenoviral vector expressing mutant herpesviral thymidine kinase reporter gene (HSV1-sr39tk) and vascular endothelial growth factor (VEGF) 121 in independent expression cassettes (Ad4tk). METHODS: Accumulation of 14C-2'-fluoro-5-methyl-1-beta-D-arabinofuranosyluracil (FIAU) and 9-(4-18F-fluoro-3-hydroxymethylbutyl)guanine (FHBG) as reporter probes, and secretion of VEGF into medium, were determined for Ad4tk-infected H9c2 rat cardiac cells in vitro. RESULTS: In vitro tracer uptake increased with increasing vector concentration and over time. It was comparable to cells infected with adenovirus expressing only wild-type HSV1-tk (reporter probe: 14C-FIAU) or mutant HSV1-sr39tk (reporter probe: 18F-FHBG). No significant uptake was observed in cells infected with adenovirus expressing VEGF alone. With increasing vector concentration, Ad4tk-infected cells increasingly released VEGF into medium. VEGF production correlated significantly with cellular reporter probe uptake (r = 0.93; P = 0.0003). CONCLUSION: The usefulness of a vector coexpressing HSV1-tk and VEGF for noninvasive imaging of expression of a therapeutic transgene has been demonstrated in vitro. This approach may allow for future in vivo monitoring of cardiac angiogenesis gene therapy.

Animals↗
PubMed2004

PET of cardiac transgene expression: comparison of 2 approaches based on herpesviral thymidine kinase reporter gene.

UNLABELLED: PET of reporter gene expression holds promise for noninvasive monitoring of gene therapy. Previously, 2 approaches based on the herpes simplex virus type 1 thymidine kinase gene (HSV1-tk) have been successfully applied to the heart. Wild-type HSV1-tk was imaged with (124)I-labeled 2'-fluoro-2'-deoxy-5-iodo-1-beta-D-arabinofuranosyl-5-iodouracil (FIAU), and a mutant HSV1-tk (HSV1-sr39tk) was imaged with (18)F-labeled 9-[4-fluoro-3-(hydroxymethyl)butyl]guanine (FHBG). The aim of this study was to compare these 2 combinations with regard to specificity, imaging contrast, and reporter probe kinetics using dynamic PET in small and large animals. METHODS: Similar titers of adenovirus-expressing wild-type HSV1-tk (Ad(tk)), mutant HSV1-sr39tk (Ad(sr39tk)), or control genes were directly injected into the myocardium of 24 rats and 8 pigs. Two days later, dynamic PET was performed with a clinical scanner during the 120 min after injection of (124)I-FIAU (Ad(tk) animals and controls) or (18)F-FHBG (Ad(sr39tk) animals and controls). Imaging with (13)N-ammonia was performed to identify cardiac regions of interest. RESULTS: In rats, significant cardiac (124)I-FIAU accumulation occurred in images obtained early (10-30 min) after Ad(tk) injection. Because of tracer washout, however, no difference between Ad(tk)-injected animals and controls was seen in the images obtained later. For (18)F-FHBG, specific myocardial accumulation greater than background levels was detected in Ad(sr39tk)-injected animals at early imaging and, in contrast to (124)I-FIAU accumulation, increased over time until the latest imaging (105-120 min). At maximum, cardiac (18)F-FHBG concentration showed a 4.15 +/- 1.65-fold increase compared with controls (105-120 min), and cardiac (124)I-FIAU concentration reached a maximal increase of 1.34 +/- 0.38-fold compared with controls (10-30 min, P = 0.0014). Global cardiac reporter probe kinetics in rats were confirmed by regional myocardial analysis in pig hearts. Transgene expression was specifically visualized by both approaches. The highest target-to-background ratio of (124)I-FIAU in Ad(tk)-infected pig myocardium was 1.50 +/- 0.20, versus 2.64 +/- 0.49 for (18)F-FHBG in Ad(sr39tk)-infected areas (P = 0.01). In vivo results were confirmed by ex vivo counting and autoradiography. CONCLUSION: Both reporter gene/probe combinations were feasible for noninvasive imaging of cardiac transgene expression in different species. Specific probe kinetics suggest different myocardial handling of pyrimidine (FIAU) and acycloguanosine (FHBG) derivatives. The results favor (18)F-FHBG with mutant HSV1-sr39tk because of continuous accumulation over time and higher imaging contrast.

Animals↗
PubMed2003

Noninvasive imaging of transgene expression by use of positron emission tomography in a pig model of myocardial gene transfer.

BACKGROUND: Radionuclide imaging of reporter gene expression may be useful for noninvasive monitoring of clinical cardiac gene therapy. Experience until now, however, has been limited to small animals. METHODS AND RESULTS: To evaluate feasibility in a clinically applicable setting, pigs were studied by conventional positron emission tomography (PET) 2 days after regional intramyocardial injection of control adenovirus or adenovirus carrying herpesviral thymidine kinase reporter gene (HSV1-tk). Myocardial blood flow was quantified by use of [13N]ammonia. Subsequently, kinetics of the reporter substrate [124I]-2'-fluoro-2'-deoxy-5-iodo-1-beta-d-arabino-furanosyluracil (FIAU) were assessed over a period of 2 hours. Areas infected with adenovirus expressing HSV1-tk showed significantly elevated FIAU retention during the first 30 minutes after injection. At later times, washout was observed, and retention was not different from that in areas infected with control virus or remote myocardium. Early in vivo FIAU uptake correlated with ex vivo images, autoradiography, and immunohistochemistry for reporter gene product after euthanasia. After intramyocardial injection of both adenoviruses, myocardial blood flow was mildly elevated compared with that in remote areas, consistent with histological signs of regional inflammation. CONCLUSIONS: In vivo quantification of regional myocardial transgene expression is feasible with clinical PET methodology, the radioiodinated reporter probe FIAU, and the HSV1-tk reporter gene. Radioactivity efflux after specific initial uptake was not observed previously in tumor studies, suggesting that tissue-specific differences in nucleoside metabolism influence reporter probe kinetics. By coregistering reporter gene expression with additional biological parameters such as myocardial blood flow, PET allows for noninvasive characterization of the success of cardiac gene transfer along with its functional correlates.

Animals↗
PubMedDate not supplied

[18F]Galacto-RGD: synthesis, radiolabeling, metabolic stability, and radiation dose estimates.

It has been demonstrated in various murine tumor models that radiolabeled RGD-peptides can be used for noninvasive determination of alphavbeta3 integrin expression. Introduction of sugar moieties improved the pharmacokinetic properties of these peptides and led to tracer with good tumor-to-background ratios. Here we describe the synthesis, radiolabeling, and the metabolic stability of a glycosylated RGD-peptide ([18F]Galacto-RGD) and give first radiation dose estimates for this tracer. The peptide was assembled on a solid support using Fmoc-protocols and cyclized under high dilution conditions. It was conjugated with a sugar amino acid, which can be synthesized via a four-step synthesis starting from pentaacetyl-protected galactose. For radiolabeling of the glycopeptide, 4-nitrophenyl-2-[18F]fluoropropionate was used. This prosthetic group allowed synthesis of [18F]Galacto-RGD with a maximum decay-corrected radiochemical yield of up to 85% and radiochemical purity >98%. The overall radiochemical yield was 29 +/- 5% with a total reaction time including final HPLC preparation of 200 +/- 18 min. The metabolic stability of [18F]Galacto-RGD was determined in mouse blood and liver, kidney, and tumor homogenates 2 h after tracer injection. The average fraction of intact tracer in these organs was approximately 87%, 76%, 69%, and 87%, respectively, indicating high in vivo stability of the radiolabeled glycopeptide. The expected radiation dose to humans after injection of [18F]Galacto-RGD has been estimated on the basis of dynamic PET studies with New Zealand white rabbits. According to the residence times in these animals the effective dose was calculated using the MIRDOSE 3.0 program as 2.2 x 10(-2) mGy/MBq. In conclusion, [18F]Galacto-RGD can be synthesized in high radiochemical yields and radiochemical purity. Despite the time-consuming synthesis of the prosthetic group 185 MBq of [18F]Galacto-RGD, a sufficient dose for patient studies, can be produced starting with approximately 2.2 GBq of [18F]flouride. Moreover, the fast excretion, the suitable metabolic stability and the low estimated radiation dose allow to evaluate this tracer in human studies.

Amino Sugars↗