Purpose
To evaluate the relevance of CEST frequency selectivity in simultaneous in vivo imaging of both of chondrosarcoma’s phenotypic features, that are, its high proteoglycan concentration and its ...hypoxic core.
Methods
Swarm rat chondrosarcomas were implanted subcutaneously in NMRI nude mice. When tumors were measurable (12‐16 days postoperative), mice were submitted to GAG, guanidyl, and APT CEST imaging. Proteoglycans and hypoxia were assessed in parallel by nuclear imaging exploiting 99mTc‐NTP 15‐5 and 18F‐FMISO, respectively. Data were completed by ex vivo analysis of proteoglycans (histology and biochemical assay) and hypoxia (immunofluorescence).
Results
Quantitative analysis of GAG CEST evidenced a significantly higher signal for tumor tissues than for muscles. These results were in agreement with nuclear imaging and ex vivo data. For imaging tumoral pH in vivo, the CEST ratio of APT/guanidyl was studied. This highlighted an important heterogeneity inside the tumor. The hypoxic status was confirmed by 18F‐FMISO PET imaging and ex vivo immunofluorescence.
Conclusion
CEST MRI simultaneously imaged both chondrosarcoma properties during a single experimental run and without the injection of any contrast agent. Both MR and nuclear imaging as well as ex vivo data were in agreement and showed that this chondrosarcoma animal model was rich in proteoglycans. However, even if tumors were lightly hypoxic at the stage studied, acidic areas were highlighted and mapped inside the tumor.
Abstract
With the emergence of disease modifying osteoarthritis drugs (DMOAD), imaging methods to quantitatively demonstrate their efficacy and to monitor osteoarthritis progression at the functional ...level are urgently needed. Our group showed that articular cartilage can be quantitatively assessed in nuclear medicine imaging by our radiotracer
99m
Tc-NTP 15-5 targeting cartilage proteoglycans. In this work, surgically induced DMM mice were treated with sprifermin or saline. We investigated cartilage remodelling in the mice knees by
99m
Tc-NTP 15-5 SPECT-CT imaging over 24 weeks after surgery, as wells as proteoglycan biochemical assays. OA alterations were scored by histology according to OARSI guidelines. A specific accumulation of
99m
Tc-NTP 15-5 in cartilage joints was evidenced in vivo by SPECT-CT imaging as early as 30 min post-iv injection. In DMM,
99m
Tc-NTP 15-5 accumulation in cartilage within the operated joints, relative to contralateral ones, was observed to initially increase then decrease as pathology progressed. Under sprifermin,
99m
Tc-NTP 15-5 uptake in pathological knees was significantly increased compared to controls, at 7-, 12- and 24-weeks, and consistent with proteoglycan increase measured 5 weeks post-surgery, as a sign of cartilage matrix remodelling. Our work highlights the potential of
99m
Tc-NTP 15-5 as an imaging-based companion to monitor cartilage remodelling in OA and DMOAD response.
As evidenced by the number of publications and patents published in the last years, the radiosynthesis of 6‐18Ffluoro‐3,4‐dihydroxy‐L‐phenylalanine (18FFDOPA) using the nucleophilic 18FF‐ process ...remains currently a challenge for the radiochemists scientific community even if promising methods for the radiofluorination of electron‐rich aromatic structures were recently developed from arylboronate, arylstannane or iodonium salt precursors. In such context, based on the use of an iodonium triflate salt precursor, we optimized a fast and efficient radiofluorination route fully automated and free from any base, cryptand or metal catalyst for the radiosynthesis of 18FFDOPA. Using this method, this clinically relevant radiotracer was produced in 64 min, 27–38 % RCY d.c. (n = 5), >99 % RCP, >99 % ee., and high Am 170–230 GBq/µmol. In addition, this optimization study clearly highlighted the important role of a triflate‐hydrogen carbonate counterion exchange during the radiolabeling process to achieve high fluorine‐18 incorporation yields.
18FFDOPA challenge: A new radiosynthetic route to 18FFDOPA was developed and fully automated from a key iodonium triflate salt precursor. This radiotracer was produced via a “minimalist approach” (see figure) involving nucleophilic 18FF– process, without any use of base, cryptand or metal catalyst, in 64 min, 27–38 % RCY, >99 % RCP, > 99 % ee., and high Am 170–230 GBq.µmol–1 (n = 5).
Purpose
This study aims to perform dosimetry for 99mTcNTP15‐5 radiotracer used in imaging of articular cartilage in rabbits and humans. The radiotracer (covered by a world patent WO 01/00621 A1) has ...been proposed in the previous years for the study of cartilage in osteoarthritis diseases. A sensitive imaging approach is essential to quantify osteoarthritis progression and monitor response to new therapies. 99mTcNTP15‐5 binds to cartilage proteoglycans whose decreased content is associated to a loss of biomedical function of cartilage. We have implemented the whole dosimetry study concerning this new radiotracer for rabbits and humans using the GATE Monte Carlo platform.
Materials and methods
Absorbed doses to critical organs are determined using the MIRD formalism. Biodistribution data are obtained by organ sampling, measuring the activity in organs for three rabbits sacrificed at various times postadministration, and by SPECT/CT imaging at different times after injection. Most important sources are cartilages (in knees and intervertebral discs), due to localization together with the liver and kidneys due to excretion of the agent. S‐values are calculated from rabbit’s CT scan and human CT scan using the GATE v8.0 Monte Carlo platform. Cumulated activity in humans is extrapolated from animals using the %kg‐dose/g method. Particular attention is given to dose calculation in bones, bone marrow and organs at risk.
Results
The dosimetry performed in rabbits shows highest absorbed doses for liver and kidneys with respectively 22.5 and 43.8 µGy per MBq of injected activity. In humans, we found absorbed doses for a maximum injected activity of 15 MBq/kg, that is, 1050 MBq for an adult of 70 kgs of 9.03 mGy for kidneys and 4.16 mGy for knee cartilages. Effective dose is 2.69 µSv/MBq.
Conclusions
The dosimetry profile of 99mTcNTP15‐5 in the context of preclinical trials is of major importance in order to make sure that organs at risk are not overexposed. GATE provides all the capability needed to calculate dose profiles for internal dosimetry. The extrapolation of the dose for a human model is a first step towards clinical trials.
It has been suggested that chemoresistance of chondrosarcoma (CHS), the cartilage tumor, is caused by the phenotypic microenvironmental features of the tumor tissue, mainly the chondrogenic ...extracellular matrix (ECM), and hypoxia. We developed and characterized a multicellular tumor spheroid (MCTS) of human chondrosarcoma HEMC-SS cells to gain insight into tumor cell biology and drug response. At Day 7, HEMC-SS spheroids exhibited a homogeneous distribution of proliferative Ki-67 positive cells, whereas in larger spheroids (Day 14 and Day 20), proliferation was mainly localized in the periphery. In the core of larger spheroids, apoptotic cells were evidenced by TUNEL assay, and hypoxia by pimonidazole staining. Interestingly, VEGF excretion, evidenced by ELISA on culture media, was detectable from Day 14 spheroids, and increased as the spheroids grew in size. HEMC-SS spheroids synthesized a chondrogenic extracellular matrix rich in glycosaminoglycans and type-2 collagen. Finally, we investigated the sensitivity of Day 7 and Day 14 chondrosarcoma MCTS to hypoxia-activated prodrug TH-302 and doxorubicin compared with their 2D counterparts. As expected, TH-302 exhibited higher cytotoxic activity on larger hypoxic spheroids (Day 14) than on non-hypoxic spheroids (Day 7), with multicellular resistance index (MCRI) values of 7.7 and 9.1 respectively. For doxorubicin, the larger-sized spheroids exhibited higher drug resistance (MCRI of 5.0 for Day 7 and 18.3 for Day 14 spheroids), possibly due to impeded drug penetration into the deep layer of spheroids, evidenced by its auto-fluorescence property. We have developed a model of human chondrosarcoma MCTS that combines an ECM rich in glycosaminoglycans with a high hypoxic core associated with VEGF excretion. This model could offer a more predictive in vitro chondrosarcoma system for screening drugs targeting tumor cells and their microenvironment.
A multi-proxy approach using trench sediment analysis on the southwestern coast of Spain (Los Lances Bay, Andalusia) provides a high-resolution record of the 1755 Lisbon tsunami and offers new means ...to interpret tsunami deposits. A combination of sedimentological (grain size, sorting, AMS), micromorphological (X-ray tomography and radiography, thin sections) and geochemical analyses (X-ray microfluorescence, ICP-AES) combines to provide a comprehensive reconstruction of the different phases of tsunami deposition and flow dynamics. Vertical variations in sediment texture, fabric and composition of the 1755 tsunami deposits in Los Lances are then discussed with reference to existing literature on diagnostic criteria commonly used for identifying and interpreting tsunami deposits. Finally possible leads for future investigation are proposed.
► Multi-proxy approach of tsunami deposits, applied to core samples. ► 2D and 3D X-ray imagery of tsunami deposits. ► Micromorphology of tsunami deposits. ► Anisotropy of magnetic susceptibility of tsunami deposits. ► Geochemistry (ICP-AES and X-ray microfluorescence) of tsunami deposits.
Osteoporosis is a bone pathology leading to increase fractures risk and challenging quality of life. Since current treatments could exhibit deleterious side effects, the use of food compounds derived ...from plants represents a promising innovative alternative due to their potential therapeutic and preventive activities against human diseases. In this study, we investigated the ability of the polyphenol fisetin to counter osteoporosis and analyzed the cellular and molecular mechanisms involved. In vivo, fisetin consumption significantly prevented bone loss in estrogen deficiency and inflammation mice osteoporosis models. Indeed, bone mineral density, micro-architecture parameters and bone markers were positively modulated by fisetin. Consistent with in vivo results, we showed that fisetin represses RANKL-induced osteoclast differentiation and activity as demonstrated by an inhibition of multinucleated cells formation, TRAP activity and differentiation genes expression. The signaling pathways NF-κB, p38 MAPK, JNK and the key transcription factors c-Fos and NFATc1 expressions induced by RANKL, were negatively regulated by fisetin. We further showed that fisetin inhibits the constitutive proteasomal degradation of MKP-1, the phosphatase that deactivates p38 and JNK. Consistently, using shRNA stable cell lines, we demonstrated that impairment of MKP-1 decreases fisetin potency. Taken together, these results strongly support that fisetin should be further considered as a bone protective agent.
The mechanisms linking fat intake to bone loss remain unclear. By demonstrating the expression of the free fatty acid receptor G-coupled protein receptor 40 (GPR40) in bone cells, we hypothesized ...that this receptor may play a role in mediating the effects of fatty acids on bone remodeling. Using micro-CT analysis, we showed that GPR40−/− mice exhibit osteoporotic features suggesting a positive role of GPR40 on bone density. In primary cultures of bone marrow, we showed that GW9508, a GRP40 agonist, abolished bone-resorbing cell differentiation. This alteration of the receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation occurred via the inhibition of the nuclear factor κB (NF-κB) signaling pathway as demonstrated by decrease in gene reporter activity, inhibitor of κB kinase (IKKα/β) activation, inhibitor of κB (IkBα) phosphorylation, and nuclear factor of activated T cells 1 (NFATc1) expression. The GPR40-dependent effect of GW9508 was confirmed using shRNA interference in osteoclast precursors and GPR40−/− primary cell cultures. In addition, in vivo administration of GW9508 counteracted ovariectomy-induced bone loss in wild-type but not GPR40−/− mice, enlightening the obligatory role of the GPR40 receptor. Then, in a context of growing prevalence of metabolic and age-related bone disorders, our results demonstrate for the first time in translational approaches that GPR40 is a relevant target for the design of new nutritional and therapeutic strategies to counter bone complications.
Background: Long chain fatty acids have been shown to activate the membrane-bound receptor GPR40.
Results: GPR40 agonist alters bone-resorbing cell differentiation through inhibition of the NF-κB system.
Conclusion: GPR40 exerts protective effects in vivo on bone tissue.
Significance: GPR40 is a nutritional and therapeutic target opening up new avenues for clinical investigations in terms of metabolic and age-related bone disorders.
The development of 64Cu-based immuno-PET radiotracers requires the use of copper-specific bifunctional chelators (BFCs) that contain functional groups allowing both convenient bioconjugation and ...stable copper complexes to limit in vivo bioreduction, transmetallation and/or transchelation. The excellent in vivo kinetic inertness of the pentaazamacrocyclic 64CuCu-15-5 complex prompted us to investigate its potential for the 64Cu-labelling of monoclonal antibodies (mAbs), compared with the well-known NODAGA and DOTA chelators. To this end, three NODAGA, DOTA and 15-5-derived BFCs, containing a pendant azadibenzocyclooctyne moiety, were synthesised and a robust methodology was determined to form covalent bonds between them and azide-functionalised trastuzumab, an anti-HER2 mAb, using strain-promoted azide-alkyne cycloaddition. Unlike the DOTA derivative, the NODAGA- and 15-5-mAb conjugates were radiolabelled with 64Cu, obtaining excellent radiochemical yields, under mild conditions. Although all the radioimmunoconjugates showed excellent stability in PBS or mouse serum, 64CuCu-15-5- and 64CuCu-NODAGA-trastuzumab presented higher resistance to transchelation when challenged by EDTA. Finally, the immunoreactive fraction of the radioimmunoconjugates (88–94%) was determined in HER-2 positive BT474 human breast cancer cells, confirming that the bioconjugation and radiolabelling processes implemented had no significant impact on antigen recognition.
Isocitrate dehydrogenases (IDHs) are metabolic enzymes commonly mutated in human cancers (glioma, acute myeloid leukaemia, chondrosarcoma, and intrahepatic cholangiocarcinoma). These mutated variants ...of IDH (mIDH) acquire a neomorphic activity, namely, conversion of α-ketoglutarate to the oncometabolite D-2-hydroxyglutarate involved in tumourigenesis. Thus, mIDHs have emerged as highly promising therapeutic targets, and several mIDH specific inhibitors have been developed. However, the evaluation of mIDH status, currently performed by biopsy, is essential for patient stratification and thus treatment and follow-up. We report herein the development of new radioiodinated and radiofluorinated analogues of olutasidenib (FT-2102) as tools for noninvasive single photon emission computed tomography (SPECT) or positron emission tomography (PET) imaging of mIDH1 up- and dysregulation in tumours. Nonradiolabelled derivatives 2 and 3 halogenated at position 6 of the quinolinone scaffold were synthesised and tested in vitro for their inhibitory potencies and selectivities in comparison with the lead compound FT-2102. Using a common organotin precursor, (S)-125I2 and (S)-18F3 were efficiently synthesised by radio-iododemetallation and copper-mediated radiofluorination, respectively. Both radiotracers were stable at room temperature in saline or DPBS solution and at 37 °C in mouse serum, allowing future planning of their in vitro and in vivo evaluations in glioma and chondrosarcoma models.
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