Evaluation of the Active Targeting of Melanin Granules after Intravenous Injection of Dendronized Nanoparticles.

The biodistribution of dendronized iron oxides, NPs10@D1_DOTAGA and melanin-targeting NPs10@D1_ICF_DOTAGA, was studied in vivo using magnetic resonance imaging (MRI) and planar scintigraphy through [177Lu]Lu-radiolabeling. MRI experiments showed high contrast power of both dendronized nanoparticles (DPs) and hepatobiliary and urinary excretions. Little tumor uptake could be highlighted after intravenous injection probably as a consequence of the negatively charged DOTAGA-derivatized shell, which reduces the diffusion across the cells' membrane. Planar scintigraphy images demonstrated a moderate specific tumor uptake of melanoma-targeted [177Lu]Lu-NPs10@D1_ICF_DOTAGA at 2 h post-intravenous injection (pi), and the highest tumor uptake of the control probe [177Lu]Lu-NPs10@D1_DOTAGA at 30 min pi, probably due to the enhanced permeability and retention effect. In addition, ex vivo confocal microscopy studies showed a high specific targeting of human melanoma samples impregnated with NPs10@D1_ICF_Alexa647_ DOTAGA.

Characterization of pro-invasive mechanisms and N-terminal cleavage of ANXA1 in melanoma.

Annexin A1 deregulation is often associated with cancer. Indeed we have shown that annexin A1 is overexpressed in melanoma and promotes metastases by formyl peptide receptor stimulation and MMP2 expression. Here, we demonstrated in different melanoma cell lines that annexin A1-MMP2 induction is mediated by MAPK and STAT3 pathways. To decipher endogenous annexin A1 action mode, we showed that annexin A1 is externalized in A375 cells and cleaved by a membrane-associated serine protease, allowing the release of a pro-invasive annexin A1 peptide in the extra cellular environment. Finally, a biochemical and proteomic approach allowed to enrich eight out of 12 members of the annexin family and to identify an original annexin A1 cleavage site localized between Ser(28) and Lys(29). Altogether, these data identify signaling pathways involved in annexin A1 pro-invasive role and suggest that externalized full-length annexin A1 interacts with formyl peptide receptors in a juxtacrine manner while ANXA 2-28 release allows autocrine and paracrine interaction.

Internal dosimetry through GATE simulations of preclinical radiotherapy using a melanin-targeting ligand.

The GATE Monte Carlo simulation platform based on the Geant4 toolkit is under constant improvement for dosimetric calculations. In this study, we explore its use for the dosimetry of the preclinical targeted radiotherapy of melanoma using a new specific melanin-targeting radiotracer labeled with iodine 131. Calculated absorbed fractions and S values for spheres and murine models (digital and CT-scan-based mouse phantoms) are compared between GATE and EGSnrc Monte Carlo codes considering monoenergetic electrons and the detailed energy spectrum of iodine 131. The behavior of Geant4 standard and low energy models is also tested. Following the different authors' guidelines concerning the parameterization of electron physics models, this study demonstrates an agreement of 1.2% and 1.5% with EGSnrc, respectively, for the calculation of S values for small spheres and mouse phantoms. S values calculated with GATE are then used to compute the dose distribution in organs of interest using the activity distribution in mouse phantoms. This study gives the dosimetric data required for the translation of the new treatment to the clinic.

In vivo experimental imaging of osteochondral defects and their healing using (99m)Tc-NTP 15-5 radiotracer.

PURPOSE: A rabbit model of osteochondral defects (OD) and spontaneous healing was longitudinally followed over 12 weeks, by in vivo joint scintigraphy using (99m)Tc-NTP 15-5, and histology. METHODS: We used two models, one with one OD (OD1 group) in the femoral condyle of one knee and the other with two ODs (OD2 group) in the femoral condyle of one knee, with the contralateral knees serving as the reference. A serial longitudinal imaging study was performed with the scintigraphic ratio (SR, operated knee uptake/contralateral knee uptake) determined at each time-point. RESULTS: ODs were imaged as radioactive defects. The SR was decreased with respective to controls, with values of 0.73 ± 0.08 and 0.65 ± 0.07 in the OD1 and OD2 groups, respectively, at 4 weeks after surgery. Histology of both OD groups revealed the presence of repair tissue characterized by a small amount of sulphated glycosaminoglycans and collagen. CONCLUSION: (99m)Tc-NTP 15-5 imaging provided quantitative criteria useful for in vivo evaluation of cartilage trauma and healing.

Anti-melanoma efficacy of internal radionuclide therapy in relation to melanin target distribution.

Targeted internal radionuclide therapy (TRT) could be an efficient, specific way to treat disseminated melanoma. Based on a previous pharmacomodulation study, we selected a quinoxaline-derived molecule (ICF01012) for its melanin specificity and kinetic properties suitable for TRT. Here, we determined the efficacy of [(131)I]ICF01012 radiotherapy in vitro and in vivo in relation to melanogenesis using human melanoma models. [(125)I]ICF01012 uptake was first assessed in relation to melanin content. We found that melanin distribution in different models was representative of pathology seen in human tumours: melanin content was high in the extracellular space of SKMel3 tumours, and accumulated primarily in melanophages in M4Beu tumours. Targeted [(131)I]ICF01012 radiotherapy had a strong anti-tumoural efficacy in pigmented versus unpigmented tumours, regardless of target distribution and content. This study supports the use of melanin targeting with (131)I-labelled iodoquinoxaline for effective treatment of melanoma.

N-(4-iodophenyl)-N'-(2-chloroethyl)urea as a microtubule disrupter: in vitro and in vivo profiling of antitumoral activity on CT-26 murine colon carcinoma cell line cultured and grafted to mice.

The antitumoral profile of the microtubule disrupter N-(4-iodophenyl)-N'-(2-chloroethyl)urea (ICEU) was characterised in vitro and in vivo using the CT-26 colon carcinoma cell line, on the basis of the drug uptake by the cells, the modifications of cell cycle, and beta-tubulin and lipid membrane profiles. N-(4-iodophenyl)-N'-(2-chloroethyl)urea exhibited a rapid and dose-dependent uptake by CT-26 cells suggesting its passive diffusion through the membranes. Intraperitoneally injected ICEU biodistributed into the grafted CT-26 tumour, resulting thus in a significant tumour growth inhibition (TGI). N-(4-iodophenyl)-N'-(2-chloroethyl)urea was also observed to accumulate within colon tissue. Tumour growth inhibition was associated with a slight increase in the number of G2 tetraploid tumour cells in vivo, whereas G2 blockage was more obvious in vitro. The phenotype of beta-tubulin alkylation that was clearly demonstrated in vitro was undetectable in vivo. Nuclear magnetic resonance analysis showed that cells blocked in G2 phase underwent apoptosis, as confirmed by an increase in the methylene group resonance of mobile lipids, parallel to sub-G1 accumulation of the cells. In vivo, a decrease of the signals of both the phospholipid precursors and the products of membrane degradation occurred concomitantly with TGI. This multi-analysis established, at least partly, the ICEU activity profile, in vitro and in vivo, providing additional data in favour of ICEU as a tubulin-interacting drug accumulating within the intestinal tract. This may provide a starting point for researches for future efficacious tubulin-interacting drugs for the treatment of colorectal cancers.

Isotopic effect study of propofol deuteration on the metabolism, activity, and toxicity of the anesthetic.

The use of isotopic substitution to delay the oxidative metabolism of the anesthetic propofol 1 was studied. The aromatic hydrogens of propofol 1 were replaced by deuterium to produce the mono- and trideuterated derivatives 4 and 5. In vitro metabolic studies on human hepatic microsomes showed no isotopic effect in the para hydroxylation of propofol, and 1, 4, and 5 display similar hypnotic activity and toxicity in mice.

Scintigraphic in vivo assessment of the development of pulmonary intravascular macrophages in liver disease: experimental study in rats with biliary cirrhosis.

STUDY OBJECTIVES: In regard to nuclear medicine literature reporting lung uptake of colloidal radiopharmaceuticals in patients with liver diseases, it has been hypothesized that liver abnormalities could trigger induction of pulmonary intravascular macrophages (PIMs) in humans normally lacking them. Recently, experimental induction of PIMs in rats in which they are not normally prevalent has been demonstrated to be at the origin of pulmonary hemodynamic alterations with an increased susceptibility to ARDS. If such induction may occur in humans, the risk of pulmonary hemodynamic alterations has to be considered and detected. This study demonstrates in a rodent model of biliary cirrhosis that scintigraphy of phagocytic function as commonly used for liver exploration is a suitable strategy for staging PIM development. DESIGN: Sixty rats were randomized as follows: bile duct section (n = 40), sham operation (n = 10), and no operation (n = 10). The rats were submitted to scintigraphy of phagocytic function every 5 days over 35 days for the assessment of radiocolloid uptake within lung and liver. At day 35, radioactivity of blood was counted and immunohistochemistry was performed on lung specimens. RESULTS: As disease progressed, radiopharmaceutical uptake decreased within the liver, while increasing considerably in the lung. At day 35, lung uptake averaged about 66% as compared to 3% before surgery. Lung histologic findings revealed numerous intravascular mononuclear cells closely related to the monocyte-macrophage lineage. CONCLUSION: Scintigraphy of phagocytic function commonly used for liver scanning could be a suitable strategy for the diagnosis of the induction of PIMs under pathologic situations.

T2 relaxation time as a marker of brain myelination: experimental MR study in two neonatal animal models.

The progress of myelination in the brain was evaluated by visualization of grey/white matter differentiation on magnetic resonance (MR) images and quantitative analysis of MR data. In vivo quantitative MR imaging was used to monitor the T2 transverse relaxation time changes associated with cerebral development and myelination. The progress of myelination was evaluated using two neonatal animal models, the monkey and the dog, known to mature at very different rates. Three beagles were studied from birth to 4 months of age and nine baboons from 1 to 30 months of age. The T2 values in the frontal, parietal and occipital white matter were calculated and the changes in these values with age were followed. Brain maturation in both species was found to correspond to decreasing T2 values in both grey and white matter. This decrease was observed both in the dog brain and, despite slower maturation, in the baboon brain, and appeared to fit with the myelination process in these models. Exploiting the physicochemical parameters of water in tissues via T2 determination is a convenient and reliable strategy for the documentation of brain development in both experimental approaches and clinical situations.

Potential of T2 relaxation time measurements for early detection of radiation injury to the brain: experimental study in pigs.

PURPOSE: To investigate the MR T2 relaxation time and histologic changes after a single-fraction 25-Gy dose of radiation to the brain of pigs. METHODS: The right hemisphere of 10 Meishan pigs was irradiated with a single dose of 25 GY at the 90% isodose, using a 12-MeV electron beam. T2 relaxation time was measured within three regions of interest in the brain: those that had received 90%, 70%, and 40% of the total dose, respectively. T2 kinetics over time was compared with histologic studies. RESULTS: Brain T2 values were noted to increase within the irradiated areas. T2 kinetics were analyzed in three phases: an immediate transient phase and two long-lasting phases. These two long-lasting phases were correlated with the detection of ventricular compression and necrosis, respectively. The T2 increase within the 90% region of interest was 19%, 22%, and 26% for phases I, II, and III, respectively. T2 measurements within other regions of interest were not significant. CONCLUSION: Although our results suggest a dose threshold for T2 variations, brain T2 values increased after irradiation at a level at which disease could not be seen on conventional MR images. This illustrates the value of using conventional MR imaging in a quantitative manner to assess molecular tissue abnormalities at earlier stages of developing diseases.

Macrophage targeting with technetium-99m labelled J001 acylated poly-galactoside for scintigraphy of inflammation: optimization and assessment of imaging specificity in experimental arthritis.

J001, an acylated poly-(1,3)-galactoside purified from the membrane of Klebsiella pneumoniae, associates selectively with macrophages via the binding to CD11b and CD14 molecules. Inflammatory foci known to recruit macrophages could thus be imaged with technetium-99m labelled J001. This study aims to define the optimal scintigraphic protocol for 99mTc-J001 imaging and to evaluate the specificity of J001 scans. A dose range study was conducted in rabbits with immunological arthritis using six different specific activities ranging from 370 to 11840 MBq·mg-1 while the intravenously injected activity was constant (37 MBq) Radiochemical purity for each preparation was documented together with the in vivo stability of the 99mTc-J001 complex using exclusion-diffusion radioHPLC of serum collected 1 h after radiopharmaceutical administration. Scintigraphic images were recorded at 2, 3 and 4h and analysed using indexes calculated from regions of interest. Specificity of the macrophage imaging was assessed by comparison with scans obtained after administration of 99mTcO4(- )or 99mTc-albumin nanocolloids. A protocol of plasma transfusion was also used to inject 99mTc-J001 after complete removal of radioactive colloids likely to be generated during the labelling. For the higher specific activities (5920 and 11840 MBq.mg-1), radiochemical purity degradation and in vitro 99mTc transchelation were noted. To prevent transchelation and 99mTc bond hydrolysis likely to impair imaging specificity, 1480 MBq.mg-1 corresponding to 25microg injected J001 was found to be the optimal usable specific activity. Results obtained with the various tracers support the hypothesis that macrophage targeting is the main factor involved in the J001 imaging of arthritis.