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Fibrosis accompanies various pathologies, and there is thus an unmet medical need for non-invasive, sensitive, and quantitative methods for the assessment of fibrotic processes. Currently, needle biopsy with subsequent histological analysis is routinely used for the diagnosis along with morphological imaging techniques, such as computed tomography (CT), magnetic resonance imaging (MRI), and ultrasound (US). However, none of these imaging techniques are sufficiently sensitive and accurate to detect minor changes in fibrosis. More importantly, they do not provide information on fibrotic activity on the molecular level, which is critical for fundamental understanding of the underlying biology and disease course. Molecular imaging technology using positron emission tomography (PET) offers the possibility of imaging not only physiological real-time activity, but also high-sensitivity and accurate quantification. This diagnostic tool is well established in oncology and has exhibited exponential development during the last two decades. However, PET diagnostics has only recently been widely applied in the area of fibrosis. This review presents the progress of development of radiopharmaceuticals for non-invasive detection of fibrotic processes, including the fibrotic scar itself, the deposition of new fibrotic components (fibrogenesis), or the degradation of existing fibrosis (fibrolysis). [ABSTRACT FROM AUTHOR]

Background: Beta-cell replacement methods such as transplantation of isolated donor islets have been proposed as a curative treatment of type 1 diabetes, but widespread application is challenging due to shortages of donor tissue and the need for continuous immunosuppressive treatments. Stem-cell-derived islets have been suggested as an alternative source of beta cells, but face transplantation protocols optimization difficulties, mainly due to a lack of available methods and markers to directly monitor grafts survival, as well as their localization and function. Molecular imaging techniques and particularly positron emission tomography has been suggested as a tool for monitoring the fate of islets after clinical transplantation. The integral membrane protein DGCR2 has been demonstrated to be a potential pancreatic islet biomarker, with specific expression on insulin-positive human embryonic stem-cell-derived pancreatic progenitor cells. The candidate Affibody molecule ZDGCR2:AM106 was radiolabeled with fluorine-18 using a novel click chemistry-based approach. The resulting positron emission tomography tracer [18F]ZDGCR2:AM106 was evaluated for binding to recombinant human DGCR2 and cryosections of stem-cell-derived islets, as well as in vivo using an immune-deficient mouse model transplanted with stem-cell-derived islets. Biodistribution of the [18F]ZDGCR2:AM106 was also assessed in healthy rats and pigs. Results: [18F]ZDGCR2:AM106 was successfully synthesized with high radiochemical purity and yield via a pretargeting approach. [18F]ZDGCR2:AM106 retained binding to recombinant human DCGR2 as well as to cryosectioned stem-cell-derived islets, but in vivo binding to native pancreatic tissue in both rat and pig was low. However, in vivo uptake of [18F]ZDGCR2:AM106 in stem-cell-derived islets transplanted in the immunodeficient mice was observed, albeit only within the early imaging frames after injection of the radiotracer. Conclusion: Targeting of DGCR2 is a promising approach for in vivo detection of stem-cell-derived islets grafts by molecular imaging. The synthesis of [18F]ZDGCR2:AM106 was successfully performed via a pretargeting method to label a site-specific covalently bonded fluorine-18 to the Affibody molecule. However, the rapid washout of [18F]ZDGCR2:AM106 from the stem-cell-derived islets graft indicates that dissociation kinetics can be improved. Further studies using alternative binders of similar classes with improved binding potential are warranted. [ABSTRACT FROM AUTHOR]

Background: Platelet-derived growth factor receptor beta (PDGFRβ) is a receptor overexpressed on activated hepatic stellate cells (aHSCs). Positron emission tomography (PET) imaging of PDGFRβ could potentially allow the quantification of fibrogenesis in fibrotic livers. This study aims to evaluate a fluorine-18 radiolabeled Affibody molecule ([18F]TZ-Z09591) as a PET tracer for imaging liver fibrogenesis. Results: In vitro specificity studies demonstrated that the trans-Cyclooctenes (TCO) conjugated Z09591 Affibody molecule had a picomolar affinity for human PDGFRβ. Biodistribution performed on healthy rats showed rapid clearance of [18F]TZ-Z09591 through the kidneys and low liver background uptake. Autoradiography (ARG) studies on fibrotic livers from mice or humans correlated with histopathology results. Ex vivo biodistribution and ARG revealed that [18F]TZ-Z09591 binding in the liver was increased in fibrotic livers (p = 0.02) and corresponded to binding in fibrotic scars. Conclusions: Our study highlights [18F]TZ-Z09591 as a specific tracer for fibrogenic cells in the fibrotic liver, thus offering the potential to assess fibrogenesis clearly. [ABSTRACT FROM AUTHOR]

Intrahepatic transplantation of islets of Langerhans (ITx) is a treatment option for individuals with complicated type 1 diabetes and profoundly unstable glycemic control, but its therapeutic success is hampered by deterioration of graft function over time. To improve ITx strategies, technologies to noninvasively monitor the fate and survival of transplanted islets over time are of great potential value. We used [68Ga]Ga-NODAGA-exendin-4 (68Ga-exendin) positron emission tomography (PET)/computed tomography (CT) imaging to demonstrate the feasibility of quantifying β-cell mass in intrahepatic islet grafts in 13 individuals with type 1 diabetes, nine after ITx with functional islet grafts and four control patients not treated with ITx. β-Cell function was measured by mixed-meal tolerance test. With dynamic 68Ga-exendin PET/CT images, we determined tracer accumulation in hepatic hotspots, and intrahepatic fat was assessed using MRI and spectroscopy. Quantification of hepatic hotspots showed a significantly higher uptake of 68Ga-exendin in the ITx group compared with the control group (median 0.55 [interquartile range 0.51–0.63] vs. 0.43 [0.42–0.45]). GLP-1 receptor expression was found in transplanted islets by immunohistochemistry. Intrahepatic fat was not detected in a majority of the individuals. Our study provides the first clinical evidence that radiolabeled exendin imaging can be used to monitor viable transplanted islets after intraportal ITx. Article Highlights: This clinical study researched the potential of radiolabeled exendin to follow the fate and survival of intrahepatic islet grafts. Is it feasible to quantitatively detect intrahepatic islet transplants with [68Ga]Ga-NODAGA-exendin-4 (68Ga-exendin) positron emission tomography (PET) imaging? Our study findings indicate that the imaging technique 68Ga-exendin PET can be used to monitor viable islet mass after intrahepatic islet transplantation in humans. Alongside functional measures, 68Ga-exendin PET imaging could significantly aid in the evaluation of strategies designed to improve islet engraftment, survival, and function. [ABSTRACT FROM AUTHOR]

The progressive loss of beta-cell mass is a hallmark of diabetes and has been suggested as a complementary approach to studying the progression of diabetes in contrast to the beta-cell function. Non-invasive nuclear medicinal imaging techniques such as Positron Emission Tomography using radiation emitting tracers have thus been suggested as more viable methodologies to visualize and quantify the beta-cell mass with sufficient sensitivity. The transmembrane G protein-coupled receptor GPR44 has been identified as a biomarker for monitoring beta-cell mass. MK-7246 is a GPR44 antagonist that selectively binds to GPR44 with high affinity and good pharmacokinetic properties. Here, we present the synthesis of MK-7246, radiolabeled with the positron emitter fluorine-18 for preclinical evaluation using cell lines, mice, rats and human pancreatic cells. Here, we have described a synthesis and radiolabeling method for producing [18F]MK-7246 and its precursor compound. Preclinical assessments demonstrated the strong affinity and selectivity of [18F]MK-7246 towards GPR44. Additionally, [18F]MK-7246 exhibited excellent metabolic stability, a fast clearance profile from blood and tissues, qualifying it as a promising radioactive probe for GPR44-directed PET imaging. [ABSTRACT FROM AUTHOR]

Imaging and radiotherapy targeting the glucose-dependent insulinotropic polypeptide receptor (GIPR) could potentially benefit the management of neuroendocrine neoplasms (NENs), complementing clinically established radiopharmaceuticals. The aim of this study was to evaluate a GIPR-targeting positron emission tomography (PET) radioligand with receptor-specific binding, fast blood clearance, and low liver background uptake. The peptide DOTA-bioconjugate, C803-GIP, was developed based on the sequence of the endogenous GIP(1-30) and synthetic exendin-4 peptides with selective amino acid mutations to combine their specificity for the GIPR and in vivo stability, respectively. The 68Ga-labeled bioconjugate was evaluated in vitro in terms of binding affinity, specificity, and internalization in HEK293 cells transfected with the human GIPR, GLP1, or GCG receptors and in sections of human insulinoma and NENs. In vivo binding specificity, biodistribution, and tissue background were investigated in mice bearing huGIPR-HEK293 xenografts and in a pig. Ex vivo organ distribution, pharmacokinetics, and dosimetry were studied in normal rats. [68Ga]Ga-C803-GIP was stable and demonstrated a high affinity to the huGIPR-HEK293 cells. Binding specificity was demonstrated in vitro in frozen sections of NENs and huGIPR-HEK293 cells. No specific uptake was observed in the negative controls of huGLP1R and huGCGR cells. A novel rationally designed PET radioligand, [68Ga]Ga-C803-GIP, demonstrated promising binding characteristics and specificity towards the GIPR. [ABSTRACT FROM AUTHOR]

Purpose: In the characterization of severe lung diseases, early detection of specific inflammatory cells could help to monitor patients' response to therapy and increase chances of survival. Macrophages contribute to regulating the resolution and termination of inflammation and have increasingly been of interest for targeted therapies. [68Ga]Ga-DOTA-TATE is an established clinical radiopharmaceutical targeting somatostatin receptor subtype 2 (SSTR 2). Since activated macrophages (M1) overexpress SSTR 2, the aim of this study was to investigate the applicability of [68Ga]Ga-DOTA-TATE for positron emission tomography (PET) imaging of M1 macrophages in pulmonary inflammation. Methods: Inflammation in the pig lungs was induced by warm saline lavage followed by injurious ventilation in farm pigs (n = 7). Healthy pigs (n = 3) were used as control. A 60-min dynamic PET scan over the lungs was performed after [68Ga]Ga-DOTA-TATE injection and [18F]FDG scan was executed afterward for comparison. The uptake of both tracers was assessed as mean standardized uptake values (SUVmean) 30–60-min post-injection. The PET scans were followed by computed tomography (CT) scans, and the Hounsfield units (HU) were quantified of the coronal segments. Basal and apical segments of the lungs were harvested for histology staining. A rat lung inflammation model was also studied for tracer specificity using lipopolysaccharides (LPS) by oropharyngeal aspiration. Organ biodistribution, ex vivo autoradiography (ARG) and histology samples were conducted on LPS treated, octreotide induced blocking and control healthy rats. Results: The accumulation of [68Ga]Ga-DOTA-TATE on pig lavage model was prominent in the more severely injured dorsal segments of the lungs (SUVmean = 0.91 ± 0.56), compared with control animals (SUVmean = 0.27 ± 0.16, p < 0.05). The tracer uptake corresponded to the damaged areas assessed by CT and histology and were in line with HU quantification. The [68Ga]Ga-DOTA-TATE uptake in LPS treated rat lungs could be blocked and was significantly higher compared with control group. Conclusion: The feasibility of the noninvasive assessment of tissue macrophages using [68Ga]Ga-DOTA-TATE/PET was demonstrated in both porcine and rat lung inflammation models. [68Ga]Ga-DOTA-TATE has a great potential to be used to study the role and presence of macrophages in humans in fight against severe lung diseases. [ABSTRACT FROM AUTHOR]

Biomarkers for the measurement of islets of Langerhans could help elucidate the etiology of diabetes. Synaptic vesicle glycoprotein 2 A (SV2A) is a potential marker reported to be localized in the endocrine pancreas. [11C]UCB-J is a novel positron emission tomography (PET) radiotracer that binds to SV2A and was previously evaluated as a synaptic marker in the central nervous system. Here, we evaluated whether [11C]UCB-J could be utilized as a PET tracer for the islets of Langerhans in the pancreas by targeting SV2A. The mRNA transcription of SV2A was evaluated in human isolated islets of Langerhans and exocrine tissue. In vitro autoradiography was performed on pancreas and brain sections from rats and pigs, and consecutive sections were immunostained for insulin. Sprague–Dawley rats were examined with PET-MRI and ex vivo autoradiography at baseline and with administration of levetiracetam (LEV). Similarly, pigs were examined with dynamic PET-CT over the pancreas and brain after administration of [11C]UCB-J at baseline and after pretreatment with LEV. In vivo radioligand binding was assessed using a one-compartment tissue model. The mRNA expression of SV2A was nearly 7 times higher in endocrine tissue than in exocrine tissue (p < 0.01). In vitro autoradiography displayed focal binding of [11C]UCB-J in the pancreas of rats and pigs, but the binding pattern did not overlap with the insulin-positive areas or with ex vivo autoradiography. In rats, pancreas binding was higher than that in negative control tissues but could not be blocked by LEV. In pigs, the pancreas and brain exhibited accumulation of [11C]UCB-J above the negative control tissue spleen. While brain binding could be blocked by pretreatment with LEV, a similar effect was not observed in the pancreas. Transcription data indicate SV2A to be a valid target for imaging islets of Langerhans, but [11C]UCB-J does not appear to have sufficient sensitivity for this application. [ABSTRACT FROM AUTHOR]

Due to the wide scale of inflammatory processes in different types of disease, more sensitive and specific biomarkers are required to improve prevention and treatment. Cluster of differentiation 69 (CD69) is one of the earliest cell surface proteins expressed by activated leukocytes. Here we characterize and optimize potential new imaging probes, Affibody molecules targeting CD69 for imaging of activated immune cells. Analysis of candidates isolated in a previously performed selection from a Z variant E. coli library to the recombinant extracellular domain of human CD69, identified one cross-reactive Z variant with affinity to murine and human CD69. Affinity maturation was performed by randomization of the primary Z variant, followed by selections from the library. The resulting Z variants were evaluated for affinity towards human and murine CD69 and thermal stability. The in vivo biodistribution was assessed by SPECT/CT in rats following conjugation of the Z variants by a DOTA chelator and radiolabeling with Indium-111. A primary Z variant with a Kd of approximately 50 nM affinity to human and murine CD69 was identified. Affinity maturation generated 5 additional Z variants with improved or similar affinity. All clones exhibited suitable stability. Radiolabeling and in vivo biodistribution in rat demonstrated rapid renal clearance for all variants, while the background uptake and washout varied. The variant ZCD69:4 had the highest affinity for human and murine CD69 (34 nM) as well as the lowest in vivo background binding. In summary, we describe the discovery, optimization and evaluation of novel Affibody molecules with affinity for CD69. Affibody molecule ZCD69:4 is suitable for further development for imaging of activated immune cells. [ABSTRACT FROM AUTHOR]

Due to the wide scale of inflammatory processes in different types of disease, more sensitive and specific biomarkers are required to improve prevention and treatment. Cluster of differentiation 69 (CD69) is one of the earliest cell surface proteins expressed by activated leukocytes. Here we characterize and optimize potential new imaging probes, Affibody molecules targeting CD69 for imaging of activated immune cells. Analysis of candidates isolated in a previously performed selection from a Z variant E. coli library to the recombinant extracellular domain of human CD69, identified one cross-reactive Z variant with affinity to murine and human CD69. Affinity maturation was performed by randomization of the primary Z variant, followed by selections from the library. The resulting Z variants were evaluated for affinity towards human and murine CD69 and thermal stability. The in vivo biodistribution was assessed by SPECT/CT in rats following conjugation of the Z variants by a DOTA chelator and radiolabeling with Indium-111. A primary Z variant with a Kd of approximately 50 nM affinity to human and murine CD69 was identified. Affinity maturation generated 5 additional Z variants with improved or similar affinity. All clones exhibited suitable stability. Radiolabeling and in vivo biodistribution in rat demonstrated rapid renal clearance for all variants, while the background uptake and washout varied. The variant ZCD69:4 had the highest affinity for human and murine CD69 (34 nM) as well as the lowest in vivo background binding. In summary, we describe the discovery, optimization and evaluation of novel Affibody molecules with affinity for CD69. Affibody molecule ZCD69:4 is suitable for further development for imaging of activated immune cells. [ABSTRACT FROM AUTHOR]

Pathological fibrosis of the liver is a landmark feature in chronic liver diseases, including nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH). Diagnosis and assessment of progress or treatment efficacy today requires biopsy of the liver, which is a challenge in, e.g., longitudinal interventional studies. Molecular imaging techniques such as positron emission tomography (PET) have the potential to enable minimally invasive assessment of liver fibrosis. This review will summarize and discuss the current status of the development of innovative imaging markers for processes relevant for fibrogenesis in liver, e.g., certain immune cells, activated fibroblasts, and collagen depositions. [ABSTRACT FROM AUTHOR]

Aim: To explore the early effects of dapagliflozin on myocardial function and metabolism in patients with type 2 diabetes without heart failure. Materials and Methods: Patients with type 2 diabetes on metformin treatment were randomized to double‐blind, 6‐week placebo or dapagliflozin 10 mg daily treatment. Investigations included cardiac function and structure with myocardial resonance imaging; cardiac oxygen consumption, perfusion and efficiency with [11C]‐acetate positron emission tomography (PET); and cardiac and hepatic fatty acid uptake with [18F]‐6‐thia‐heptadecanoic acid PET, analysed by ANCOVA as least square means with 95% confidence intervals. Results: Evaluable patients (placebo: n = 24, dapagliflozin: n = 25; 53% males) had a mean age of 64.4 years, a body mass index of 30.2 kg/m2 and an HbA1c of 6.7%. Body weight and HbA1c were significantly decreased by dapagliflozin versus placebo. Dapagliflozin had no effect on myocardial efficiency, but external left ventricular (LV) work (−0.095 [−0.145, −0.043] J/g/min) and LV oxygen consumption were significantly reduced (−0.30 [−0.49, −0.12] J/g/min) by dapagliflozin, although the changes were not statistically significant versus changes in the placebo group. Change in left atrial maximal volume with dapagliflozin versus placebo was −3.19 (−6.32, −0.07) mL/m2 (p =.056). Peak global radial strain decreased with dapagliflozin versus placebo (−3.92% [−7.57%, −0.28%]; p =.035), while peak global longitudinal and circumferential strains were unchanged. Hepatic fatty acid uptake was increased by dapagliflozin versus placebo (0.024 [0.004, 0.044] μmol/g/min; p =.018), while cardiac uptake was unchanged. Conclusions: This exploratory study indicates reduced heart work but limited effects on myocardial function, efficiency and cardiac fatty acid uptake, while hepatic fatty acid uptake increased, after 6 weeks of treatment with dapagliflozin. [ABSTRACT FROM AUTHOR]

The beneficial role of glucose-dependent insulinotropic polypeptide receptor (GIPR) in weight control and maintaining glucose levels has led to the development of several multi-agonistic peptide drug candidates, targeting GIPR and glucagon like peptide 1 receptor (GLP1R) and/or the glucagon receptor (GCGR). The in vivo quantification of target occupancy by these drugs would accelerate the development of new drug candidates. The aim of this study was to evaluate a novel peptide (GIP1234), based on previously reported ligand DOTA-GIP-C803, modified with a fatty acid moiety to prolong its blood circulation. It would allow higher target tissue exposure and consequently improved peptide uptake as well as in vivo PET imaging and quantification of GIPR occupancy by novel drugs of interest. A 40 amino acid residue peptide (GIP1234) was synthesized based on DOTA-GIP-C803, in turn based on the sequences of endogenous GIP and Exendin-4 with specific amino acid modifications to obtain GIPR selectivity. A palmitoyl fatty acid chain was furthermore added at Lys14 via a glutamic acid linker to prolong its blood circulation time by the interaction with albumin. GIP1234 was conjugated with a DOTA chelator at the C-terminal cysteine residue to achieve 68Ga radiolabeling. The resulting PET probe, [68Ga]Ga-DOTA-GIP1234 was evaluated for receptor binding specificity and selectivity using HEK293 cells transfected with human GIPR, GLP1R, or GCGR. Blocking experiments with tirzepatide (2 μM) were conducted using huGIPR HEK293 cells to investigate binding specificity. Ex vivo and in vivo organ distribution of [68Ga]Ga-DOTA-GIP1234 was studied in rats and a pig in comparison to [68Ga]Ga-DOTA-C803-GIP. Binding of [68Ga]Ga-DOTA-GIP1234 to albumin was assessed in situ using polyacrylamide gel electrophoresis (PAGE). The stability was tested in formulation buffer and rat blood plasma. [68Ga]Ga-DOTA-GIP1234 was synthesized with non-decay corrected radiochemical yield of 88 ± 3.7 % and radiochemical purity of 97.8 ± 0.8 %. The molar activity for the radiotracer was 8.1 ± 1.1 MBq/nmol. [68Ga]Ga-DOTA-GIP1234 was stable and maintained affinity to huGIPR HEK293 cells (dissociation constant (K d) = 40 ± 12.5 nM). The binding of [68Ga]Ga-DOTA-GIP1234 to huGCGR and huGLP1R cells was insignificant. Pre-incubation of huGIPR HEK293 cell sections with tirzepatide resulted in the decrease of [68Ga]Ga-DOTA-GIP1234 binding by close to 90 %. [68Ga]Ga-DOTA-GIP1234 displayed slow blood clearance in pigs with SUV = 3.5 after 60 min. Blood retention of the tracer in rat was 2-fold higher than that of [68Ga]Ga-DOTA-C803-GIP. [68Ga]Ga-DOTA-GIP1234 also demonstrated strong liver uptake in both pig and rat combined with decreased renal excretion. The concentration dependent binding of [68Ga]Ga-DOTA-GIP1234 to albumin was confirmed in situ by PAGE. [68Ga]Ga-DOTA-GIP1234 demonstrated nanomolar affinity and selectivity for huGIPR in vitro. Addition of a fatty acid moiety prolonged blood circulation time and tissue exposure in both rat and pig in vivo. However, the liver uptake was also increased which may make PET imaging of abdominal tissues such as pancreas challenging. The investigation of the influence of fatty acid moiety on the biological performance of the peptide ligand paved the way for further rational design of GIPR ligand analogues with improved characteristics. [Display omitted] [ABSTRACT FROM AUTHOR]

There is an unmet medical need for non-invasive, sensitive, and quantitative methods for the assessment of fibrosis. Herein, an improved collagelin analogue labelled with gallium-68 for use with positron emission tomography (PET) is presented. A cyclic peptide, c[CPGRVNleHGLHLGDDEGPC], was synthesized by solid-phase peptide synthesis, conjugated to 2-(4,7-bis(2-(tert-butoxy)-2-oxoethyl)-1,4,7-triazonan-1-yl)acetic acid, and labelled with gallium-68. High performance liquid chromatography (HPLC) was used for the quality and stability assessment of the collagelin analogue. Non-specific organ distribution, blood clearance, and excretion rates were investigated in healthy mice and rats using ex vivo organ distribution analysis and dynamic in vivo PET/CT. Mice with carbon tetrachloride (CCl4) induced liver fibrosis were used for the investigation of specific binding via in vitro frozen section autoradiography, ex vivo organ distribution, and in vivo PET/CT. A non-decay corrected radiochemical yield (48 ± 6%) of [68Ga]Ga-NOTA-PEG2-c[CPGRVNleHGLHLGDDEGPC] ([68Ga]Ga-NO2A-[Nle13]-Col) with a radiochemical purity of 98 ± 2% was achieved without radical scavengers. The 68Ga-labelling was regioselective and stable at ambient temperature for at least 3 h. The autoradiography of the cryosections of fibrotic mouse liver tissue demonstrated a distinct heterogeneous radioactivity uptake that correlated with the fibrosis scores estimated after Sirius Red staining. The blood clearance and tissue washout from the [68Ga]Ga-NO2A-[Nle13]-Col was fast in both normal and diseased mice. Dosimetry investigation in rats indicated the possibility for 4–5 PET/CT examinations per year. Radiolytic stability of the collagelin analogue was achieved by the substitution of methionine with norleucine amino acid residue without a deterioration of its binding capability. [68Ga]Ga-NO2A-[Nle13]-Col demonstrated a safe dosimetry profile suitable for repeated scanning. [ABSTRACT FROM AUTHOR]