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EJNMMI Research Nov 2023Targeting prostate-specific membrane antigen (PSMA) has been highly successful for imaging and treatment of prostate cancer. However, heterogeneity in...
Heterogeneity of prostate-specific membrane antigen (PSMA) and PSMA-ligand uptake detection combining autoradiography and postoperative pathology in primary prostate cancer.
BACKGROUND
Targeting prostate-specific membrane antigen (PSMA) has been highly successful for imaging and treatment of prostate cancer. However, heterogeneity in immunohistochemistry indicates limitations in the effect of imaging and radionuclide therapy of multifocal disease. Tc-PSMA-I&S is a γ-emitting probe, which can be used for intraoperative lesion detection and postsurgical autoradiography (ARG). We aimed to study its intraprostatic distribution and compared it with (immuno)-histopathology.
RESULTS
Seventeen patients who underwent RGS between 11/2018 and 01/2020 with a total of 4660 grids were included in the preliminary analysis. Marked intratumor and intra-patient heterogeneity of PSMA expression was detected, and PSMA negative foci were observed in all samples (100%). Heterogeneous intra-patient PSMA-ligand uptake was observed, and no significant correlation was present between the degree of heterogeneity of PSMA expression and PSMA-ligand uptake. Higher PSMA-ligand uptake was observed in GS ≥ 8 than GS < 8 (p < 0.001). The appearance of Gleason Pattern (GP) 4 was strongly associated with higher uptake (coefficient: 0.43, p < 0.001), while GP 5 also affected the uptake (coefficient: 0.07, p < 0.001).
CONCLUSION
PSMA expression and PSMA-ligand uptake show marked heterogeneity. Prostate carcinoma with GP 4 showed significantly higher uptake compared with non-neoplastic prostate tissue. Our analyses extend the scope of applications of radiolabeled PSMA-ligands to ARG for identifying high-grade disease and using its signal as a noninvasive biomarker in prostate cancer.
PubMed: 37971546
DOI: 10.1186/s13550-023-01044-8 -
Brain Research Dec 2020The neuropathological hallmark of Parkinsońs disease, multiple system atrophy and dementia with Lewy bodies is the accumulation of α-synuclein. The development of an...
The neuropathological hallmark of Parkinsońs disease, multiple system atrophy and dementia with Lewy bodies is the accumulation of α-synuclein. The development of an imaging biomarker for α-synuclein is an unmet need. To date, no selective α-synuclein imaging agent has been identified, though initial studies suggest that the tau tracer [C]PBB3 displays some degree of binding to α-synuclein. In this study, a series of compounds derived from the PBB3 scaffold were examined using fluorescence imaging and tissue microarrays (TMAs) derived from brain samples with different proteinopathies. One compound, C05-01, was selected based on its higher fluorescence signal associated with Lewy body aggregates compared with other PBB3 analogues. In vitro binding assays using human brain homogenates and recombinant fibrils indicated that C05-01 had higher affinity for α-synuclein (K/Ki 25 nM for fibrils, Ki 3.5 nM for brain homogenates) as compared with PBB3 (K 58 nM). In autoradiography (ARG) studies using fresh frozen human tissue and TMAs, [H]C05-01 displayed specific binding in cases with α-synuclein pathology. C05-01 is the first PBB3 analogue developed as a potential compound targeting α-synuclein. Despite improved affinity for α-synuclein, C05-01 showed specific binding in AD tissue with Amyloid β and tau pathology, as well as relatively high non-specific and off-target binding. Additional efforts are needed to optimize the pharmacological and physicochemical properties of this series of compounds as ligands for α-synuclein. This study also showed that the construction of TMAs from different proteinopathies provides a tool for evaluation of fluorescent or radiolabelled compounds binding to misfolded proteins.
Topics: Alzheimer Disease; Benzothiazoles; Brain; Dementia; Humans; Lewy Bodies; Parkinson Disease; alpha-Synuclein; tau Proteins
PubMed: 32956648
DOI: 10.1016/j.brainres.2020.147131 -
Theranostics 2021Myocardial infarction (MI) evokes an organized remodeling process characterized by the activation and transdifferentiation of quiescent cardiac fibroblasts to generate...
Myocardial infarction (MI) evokes an organized remodeling process characterized by the activation and transdifferentiation of quiescent cardiac fibroblasts to generate a stable collagen rich scar. Early fibroblast activation may be amenable to targeted therapy, but is challenging to identify . We aimed to non-invasively image active fibrosis by targeting the fibroblast activation protein (FAP) expressed by activated (myo)fibroblasts, using a novel positron emission tomography (PET) radioligand [Ga]MHLL1 after acute MI. One-step chemical synthesis and manual as well as module-based radiolabeling yielded [Ga]MHLL1. Binding characteristics were evaluated in murine and human FAP-transfected cells, and stability tested in human serum. Biodistribution in healthy animals was interrogated by dynamic PET imaging, and metabolites were measured in blood and urine. The temporal pattern of FAP expression was determined by serial PET imaging at 7 d and 21 d after coronary artery ligation in mice as percent injected dose per gram (%ID/g). PET measurements were validated by autoradiography and immunostaining for FAP and inflammatory macrophages. [Ga]MHLL1 displayed specific uptake in murine and human FAP-positive cells (p = 0.0208). In healthy mice the tracer exhibited favorable imaging characteristics, with low blood pool retention and dominantly renal clearance. At 7 d after coronary artery ligation, [Ga]MHLL1 uptake was elevated in the infarct relative to the non-infarcted remote myocardium (1.3 ± 0.3 1.0 ± 0.2 %ID/g, p < 0.001) which persisted to 21 d after MI (1.3 ± 0.4 1.1 ± 0.4 %ID/g, p = 0.013). Excess unlabeled compound blocked tracer accumulation in both infarct and non-infarct remote myocardium regions (p < 0.001). Autoradiography and histology confirmed the regional uptake of [Ga]MHLL1 in the infarct and especially border zone regions, as identified by Masson trichrome collagen staining. Immunostaining further delineated persistent FAP expression at 7 d and 21 d post-MI in the border zone, consistent with tracer distribution . The simplified synthesis of [Ga]MHLL1 bears promise for non-invasive characterization of fibroblast activation protein early in remodeling after MI.
Topics: Animals; Autoradiography; Cell Line, Tumor; Endopeptidases; Fibroblasts; Fibrosis; Gallium Radioisotopes; Humans; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Molecular Imaging; Myocardial Infarction; Myocardium; Positron-Emission Tomography; Tissue Distribution; Tomography, X-Ray Computed
PubMed: 34335962
DOI: 10.7150/thno.51419 -
In Vivo (Athens, Greece) 2024Since acute myeloid leukemias still represent the most aggressive type of adult acute leukemias, the profound understanding of disease pathology is of paramount...
BACKGROUND/AIM
Since acute myeloid leukemias still represent the most aggressive type of adult acute leukemias, the profound understanding of disease pathology is of paramount importance for diagnostic and therapeutic purposes. Hence, this study aimed to explore the real-time disease fate with the establishment of an experimental myelomonoblastic leukemia (My1/De) rat model using preclinical positron emission tomography (PET) and whole-body autoradiography.
MATERIALS AND METHODS
In vitro [F]F-FDG uptake studies were performed to compare the tracer accumulation in the newly cultured My1/De tumor cell line (blasts) with that in healthy control and My1/De bone marrow suspensions. Post transplantation of My1/De cells under the left renal capsule of Long-Evans rats, primary My1/De tumorigenesis, and metastatic propagation were investigated using [F]F-FDG PET imaging, whole-body autoradiography and phosphorimage analyses. To assess the organ uptake profile of the tumor-carrying animals we accomplished ex vivo biodistribution studies.
RESULTS
The tracer accumulation in the My1/De culture cells exceeded that of both the tumorous and the healthy bone marrow suspensions (p<0.01). Based on in vivo imaging, the subrenally transplanted My1/De cells resulted in the development of leukemia in the abdominal organs, and metastasized to the mesenterial and thoracic parathymic lymph nodes (PTLNs). The lymphatic spread of metastasis was further confirmed by the significantly higher %ID/g values of the metastatic PTLNs (4.25±0.28) compared to the control (0.94±0.34). Cytochemical staining of the peripheral blood, autopsy findings, and wright-Giemsa-stained post-mortem histological sections proved the leukemic involvement of the assessed tissues/organs.
CONCLUSION
The currently established My1/De model appears to be well-suited for further leukemia-related therapeutic and diagnostic investigations.
Topics: Animals; Rats; Disease Models, Animal; Fluorodeoxyglucose F18; Autoradiography; Positron-Emission Tomography; Cell Line, Tumor; Tissue Distribution; Leukemia, Myeloid, Acute; Radiopharmaceuticals; Male; Humans
PubMed: 38688644
DOI: 10.21873/invivo.13540 -
Nuclear Medicine and Biology 2022Antibody-based constructs, engineered to enter the brain using transferrin receptor (TfR) mediated transcytosis, have been successfully used as PET radioligands for...
PURPOSE
Antibody-based constructs, engineered to enter the brain using transferrin receptor (TfR) mediated transcytosis, have been successfully used as PET radioligands for imaging of amyloid-beta (Aβ) in preclinical studies. However, these radioligands have been large and associated with long circulation times, i.e. non-optimal properties for neuroPET radioligands. The aim of this study was to investigate the in vivo brain delivery of the radiolabeled nanobody VHH-E9 that binds to glial fibrillary acidic protein (GFAP) expressed by reactive astrocytes, without and with fusion to a TfR binding moiety, as potential tools to detect neuroinflammation.
METHODS
Three protein constructs were recombinantly expressed: 1) The GFAP specific nanobody VHH-E9, 2) VHH-E9 fused to a single chain variable fragment of the TfR binding antibody 8D3 (scFv8D3) and 3) scFv8D3 alone. Brain delivery of the constructs was investigated at 2 h post injection. Binding to GFAP was studied with autoradiography while in vivo brain retention of [I]VHH-E9 and [I]VHH-E9-scFv8D3 was further investigated at 8 h, 24 h and 48 h in wild-type (WT), and at the same time points in transgenic mice (ArcSwe) that in addition to Aβ pathology also display neuroinflammation.
RESULTS
At 2 h after administration, [I]VHH-E9-scFv8D3 and [I]scFv8D3 displayed 3-fold higher brain concentrations than [I]VHH-E9. In vitro autoradiography showed distinct binding of both [I]VHH-E9-scFv8D3 and [I]VHH-E9 to regions with abundant GFAP in ArcSwe mice. However, in vivo, there was no difference in brain concentrations between WT and ArcSwe at any of the studied time points.
CONCLUSIONS
Fused to scFv8D3, VHH-E9 displayed increased brain delivery. When radiolabeled and applied on brain sections, the bispecific construct was able to discriminate between WT and ArcSwe mice, but in vivo brain uptake and retention over time did not differ between WT and ArcSwe mice.
Topics: Animals; Mice; Glial Fibrillary Acidic Protein; Alzheimer Disease; Neuroinflammatory Diseases; Positron-Emission Tomography; Brain; Amyloid beta-Peptides; Mice, Transgenic
PubMed: 35487832
DOI: 10.1016/j.nucmedbio.2022.04.002 -
Chembiochem : a European Journal of... Dec 2022Tetrazine (Tz)-trans-cyclooctene (TCO) ligation is an ultra-fast and highly selective reaction and it is particularly suited to label biomolecules under physiological...
Tetrazine (Tz)-trans-cyclooctene (TCO) ligation is an ultra-fast and highly selective reaction and it is particularly suited to label biomolecules under physiological conditions. As such, a H-Tz based synthon would have wide applications for in vitro/ex vivo assays. In this study, we developed a H-labeled Tz and characterized its potential for application to pretargeted autoradiography. Several strategies were explored to synthesize such a Tz. However, classical approaches such as reductive halogenation failed. For this reason, we designed a Tz containing an aldehyde and explored the possibility of reducing this group with NaBT . This approach was successful and resulted in [ H]-(4-(6-(pyridin-2-yl)-1,2,4,5-tetrazin-3-yl)phenyl)methan-t-ol with a radiochemical yield of 22 %, a radiochemical purity of 96 % and a molar activity of 0.437 GBq/μmol (11.8 Ci/mmol). The compound was successfully applied to pretargeted autoradiography. Thus, we report the synthesis of the first H-labeled Tz and its successful application as a labeling building block.
Topics: Cell Line, Tumor; Radiopharmaceuticals; Heterocyclic Compounds; Cyclooctanes
PubMed: 36333105
DOI: 10.1002/cbic.202200539 -
PloS One 2019In the rat, oxytocin (OT) produces dose-dependent diuretic and natriuretic responses. Post-translational enzymatic conversion of the OT biosynthetic precursor forms both...
In the rat, oxytocin (OT) produces dose-dependent diuretic and natriuretic responses. Post-translational enzymatic conversion of the OT biosynthetic precursor forms both mature and C-terminally extended peptides. The plasma concentrations of these C-terminally extended peptides (OT-G; OT-GK and OT-GKR) are elevated in newborns and pregnant rats. Intravenous injection of OT-GKR to rats inhibits diuresis, whereas injection of amidated OT stimulates diuresis. Since OT and OT-GKR show different effects on the urine flow, we investigated whether OT-GKR modulates renal action by inhibition of the arginine-vasopressin (AVP) receptor V2 (V2R), the receptor involved in renal water reabsorption. Experiments were carried out in the 8-week-old Wistar rats receiving intravenous (iv) injections of vehicle, OT, OT-GKR or OT+OT-GKR combination. OT (10 μmol/kg) increased urine outflow by 40% (P<0.01) and sodium excretion by 47% (P<0.01). Treatment with OT-GKR (10 μmol/kg) decreased diuresis by 50% (P<0.001), decreased sodium excretion by 50% (P<0.05) and lowered potassium by 42% (P<0.05). OT antagonist (OTA) reduced diuresis and natriuresis exerted by OT, whereas the anti-diuretic effect of OT-GKR was unaffected by OTA. The treatment with V2R antagonist (V2A) in the presence and absence of OT induced diuresis, sodium and potassium outflow. V2A in the presence of OT-GKR only partially increased diuresis and natriuresis. Autoradiography and molecular docking analysis showed potent binding of OT-GKR to V2R. Finally, the release of cAMP from CHO cells overexpressing V2 receptor was induced by low concentration of AVP (EC50:4.2e-011), at higher concentrations of OT (EC50:3.2e-010) and by the highest concentrations of OT-GKR (EC50:1.1e-006). OT-GKR potentiated cAMP release when combined with AVP, but blocked cAMP release when combined with OT. These results suggest that OT-GKR by competing for the OT renal receptor (OTR) and binding to V2R in the kidney, induces anti-diuretic, anti-natriuretic, and anti-kaliuretic effects.
Topics: Animals; Autoradiography; Binding, Competitive; CHO Cells; Cell Line; Cricetinae; Cricetulus; Cyclic AMP; Diuresis; Electrolytes; Humans; Kidney; Molecular Docking Simulation; Natriuresis; Oxytocin; Peptides; Rats; Rats, Wistar; Receptors, Vasopressin; Urination; Vasopressins
PubMed: 31269062
DOI: 10.1371/journal.pone.0219205 -
Journal of Nuclear Cardiology :... Dec 2021Imaging Somatostatin Subtype Receptor 2 (SST) expressing macrophages by [DOTA,Tyr]-octreotate (DOTATATE) has proven successful for plaque detection. DOTA-JR11 is a SST...
BACKGROUND
Imaging Somatostatin Subtype Receptor 2 (SST) expressing macrophages by [DOTA,Tyr]-octreotate (DOTATATE) has proven successful for plaque detection. DOTA-JR11 is a SST targeting ligand with a five times higher tumor uptake than DOTATATE, and holds promise to improve plaque imaging. The aim of this study was to evaluate the potential of DOTA-JR11 for plaque detection.
METHODS AND RESULTS
Atherosclerotic ApoE mice (n = 22) fed an atherogenic diet were imaged by SPECT/CT two hours post injection of [In]In-DOTA-JR11 (~ 200 pmol, ~ 50 MBq). In vivo plaque uptake of [In]In-DOTA-JR11 was visible in all mice, with a target-to-background-ratio (TBR) of 2.23 ± 0.35. Post-mortem scans after thymectomy and ex vivo scans of the arteries after excision of the arteries confirmed plaque uptake of the radioligand with TBRs of 2.46 ± 0.52 and 3.43 ± 1.45 respectively. Oil red O lipid-staining and ex vivo autoradiography of excised arteries showed [In]In-DOTA-JR11 uptake at plaque locations. Histological processing showed CD68 (macrophages) and SST expressing cells in plaques. SPECT/CT, in vitro autoradiography and immunohistochemistry performed on slices of a human carotid endarterectomy sample showed [In]In-DOTA-JR11 uptake at plaque locations containing CD68 and SST expressing cells.
CONCLUSIONS
The results of this study indicate DOTA-JR11 as a promising ligand for visualization of atherosclerotic plaque inflammation.
Topics: Animals; Heterocyclic Compounds, 1-Ring; Indium Radioisotopes; Inflammation; Mice; Plaque, Atherosclerotic; Receptors, Somatostatin
PubMed: 32026330
DOI: 10.1007/s12350-020-02046-y -
Pharmacological Research Nov 2021To enable non-invasive real-time quantification of vasopressin 1A (V1A) receptors in peripheral organs, we sought to develop a suitable PET probe that would allow...
OBJECTIVES
To enable non-invasive real-time quantification of vasopressin 1A (V1A) receptors in peripheral organs, we sought to develop a suitable PET probe that would allow specific and selective V1A receptor imaging in vitro and in vivo.
METHODS
We synthesized a high-affinity and -selectivity ligand, designated compound 17. The target structure was labeled with carbon-11 and tested for its utility as a V1A-targeted PET tracer by cell uptake studies, autoradiography, in vivo PET imaging and ex vivo biodistribution experiments.
RESULTS
Compound 17 (PF-184563) and the respective precursor for radiolabeling were synthesized in an overall yield of 49% (over 7 steps) and 40% (over 8 steps), respectively. An inhibitory constant of 0.9 nM towards the V1A receptors was measured, while excellent selectivity over the related V1B, V2 and OT receptor (IC >10,000 nM) were obtained. Cell uptake studies revealed considerable V1A binding, which was significantly reduced in the presence of V1A antagonists. Conversely, there was no significant blockade in the presence of V1B and V2 antagonists. In vitro autoradiography and PET imaging studies in rodents indicated specific tracer binding mainly in the liver. Further, the pancreas, spleen and the heart exhibited specific binding of [C]17 ([C]PF-184563) by ex vivo biodistribution experiments.
CONCLUSION
We have developed the first V1A-targeted PET ligand that is suitable for subtype-selective receptor imaging in peripheral organs including the liver, heart, pancreas and spleen. Our findings suggest that [C]PF-184563 can be a valuable tool to study the role of V1A receptors in liver diseases, as well as in cardiovascular pathologies.
Topics: Animals; Autoradiography; Benzodiazepines; CHO Cells; Carbon Radioisotopes; Cricetulus; Female; Ligands; Liver; Male; Mice; Myocardium; Pancreas; Positron-Emission Tomography; Radiopharmaceuticals; Rats, Wistar; Receptors, Vasopressin; Spleen; Triazoles; Rats
PubMed: 34536549
DOI: 10.1016/j.phrs.2021.105886 -
Life (Basel, Switzerland) Feb 2023Inorganic nanoparticles of boron-rich compounds represent an attractive alternative to boron-containing molecules, such as boronophenylalanine or boranes, for BNCT...
Inorganic nanoparticles of boron-rich compounds represent an attractive alternative to boron-containing molecules, such as boronophenylalanine or boranes, for BNCT applications. This work describes the synthesis and biological activity of multifunctional boron carbide nanoparticles stabilized with polyacrylic acid (PAA) and a gadolinium ()-rich solid phase. A fluorophore (DiI) was included in the PAA functionalization, allowing the confocal microscopy imaging of the nanoparticles. Analysis of the interaction and activity of these fluorescent -containing nanoparticles (FGdBNPs) with cultured cells was appraised using an innovative correlative microscopy approach combining intracellular neutron autoradiography, confocal, and SEM imaging. This new approach allows visualizing the cells, the FGdBNP, and the events deriving from the nuclear process in the same image. Quantification of by neutron autoradiography in cells treated with FGdBNPs confirmed a significant accumulation of NPs with low levels of cellular toxicity. These results suggest that these NPs might represent a valuable tool for achieving a high boron concentration in tumoral cells.
PubMed: 36836786
DOI: 10.3390/life13020429