[18F]-Radiolabelled Nanoplatforms: A Critical Review of Their Intrinsic Characteristics, Radiolabelling Methods, and Purification Techniques.

A wide range of nano-objects is found in many applications of our everyday life. Recognition of their peculiar properties and ease of functionalization has prompted their engineering into multifunctional platforms that are supposed to afford efficient tools for the development of biomedical applications. However, bridging the gap between bench to bedside cannot be expected without a good knowledge of their behaviour in vivo, which can be obtained through non-invasive imaging techniques, such as positron emission tomography (PET). Their radiolabelling with [18F]-fluorine, a technique already well established and widely used routinely for PET imaging, with [18F]-FDG for example, and in preclinical investigation using [18F]-radiolabelled biological macromolecules, has, therefore, been developed. In this context, this review highlights the various nano-objects studied so far, the reasons behind their radiolabelling, and main in vitro and/or in vivo results obtained thereof. Then, the methods developed to introduce the radioelement are presented. Detailed indications on the chemical steps involved are provided, and the stability of the radiolabelling is discussed. Emphasis is then made on the techniques used to purify and analyse the radiolabelled nano-objects, a point that is rarely discussed despite its technical relevance and importance for accurate imaging. The pros and cons of the different methods developed are finally discussed from which future work can develop.

Preclinical study of 212Pb alpha-radioimmunotherapy targeting CD20 in non-Hodgkin lymphoma.

BACKGROUND: Despite therapeutic advances, Non-Hodgkin lymphoma (NHL) relapses can occur. The development of radioimmunotherapy (RIT) with α-emitters is an attractive alternative. In this study, we investigated the potential of α-RIT in conjunction with 212Pb-rituximab for the treatment of NHL. METHODS: EL4-hCD20-Luc cells (mouse lymphoma cell line) were used for in vitro and in vivo studies. Biodistribution and efficacy studies were performed on C57BL/6 mice injected intravenously with 25 × 103 cells. RESULTS: 212Pb-rituximab (0.925-7.4 kBq/mL) inhibit proliferation of EL4-hCD20-Luc cells in vitro. Biodistribution of 203/212Pb-rituximab in mice showed a significant tumour uptake and suggested that the liver, spleen, and kidneys were the organs at risk. For efficacy studies, mice were treated at either 11 days (early stage) or 20-30 days after injection of tumour cells (late stage). Treatment with 277.5 kBq 212Pb-rituximab significantly prolonged survival. Even at an advanced tumour stage, significant tumour regression occurred, with an increase in the median survival time to 28 days, compared with 9 days in the controls. CONCLUSIONS: These results show the efficacy of 212Pb-rituximab in a murine syngeneic lymphoma model, in terms of significant tumour regression and increased survival, thereby highlighting the potency of α-RIT for the treatment of NHL.

212Pb α-Radioimmunotherapy Targeting CD38 in Multiple Myeloma: A Preclinical Study.

Multiple myeloma (MM) is a plasma cell cancer and represents the second most frequent hematologic malignancy. Despite new treatments and protocols, including high-dose chemotherapy associated with autologous stem cell transplantation, the prognosis of MM patients is still poor. α-radioimmunotherapy (α-RIT) represents an attractive treatment strategy because of the high-linear-energy transfer and short pathlength of α-radiation in tissues, resulting in high tumor cell killing and low toxicity to surrounding tissues. In this study, we investigated the potential of α-RIT with 212Pb-daratumumab (anti-hCD38), in both in vitro and in vivo models, as well as an antimouse CD38 antibody using in vivo models. Methods: Inhibition of cell proliferation after incubation of the RPMI8226 cell line with an increasing activity (0.185-3.7 kBq/mL) of 212Pb-isotypic control or 212Pb-daratumumab was evaluated. Biodistribution was performed in vivo by SPECT/CT imaging and after death. Dose-range-finding and acute toxicity studies were conducted. Because daratumumab does not bind the murine CD38, biodistribution and dose-range finding were also determined using an antimurine CD38 antibody. To evaluate the in vivo efficacy of 212Pb-daratumumab, mice were engrafted subcutaneously with 5 × 106 RPMI8226 cells. Mice were treated 13 d after engraftment with an intravenous injection of 212Pb-daratumumab or control solution. Therapeutic efficacy was monitored by tumor volume measurements and overall survival. Results: Significant inhibition of proliferation of the human myeloma RPMI8226 cell line was observed after 3 d of incubation with 212Pb-daratumumab, compared with 212Pb-isotypic control or cold antibodies. Biodistribution studies showed a specific tumoral accumulation of daratumumab. No toxicity was observed with 212Pb-daratumumab up to 370 kBq because of lack of cross-reactivity. Nevertheless, acute toxicity experiments with 212Pb-anti-mCD38 established a toxic activity of 277.5 kBq. To remain within realistically safe treatment activities for efficacy studies, mice were treated with 185 kBq or 277.5 kBq of 212Pb-daratumumab. Marked tumor growth inhibition compared with controls was observed, with a median survival of 55 d for 277.5 kBq of 212Pb-daratumumab instead of 11 d for phosphate-buffered saline. Conclusion: These results showed 212Pb-daratumumab to have efficacy in xenografted mice, with significant tumor regression and increased survival. This study highlights the potency of α-RIT in MM treatment.

Radiolabelled polymeric IgA: from biodistribution to a new molecular imaging tool in colorectal cancer lung metastases.

By radiolabelling monomeric (m) and polymeric (p) IgA with technetium 99m (99mTc), this study assessed IgA biodistribution and tumour-targeting potency. IgA directed against carcinoembryonic antigen (CEA), a colorectal cancer marker, was selected to involve IgA mucosal tropism. Ig was radiolabelled with 99mTc-tricarbonyl after derivatisation by 2-iminothiolane. 99mTc-IgA was evaluated by in vitro analysis. The biodistributions of radiolabelled anti-CEA mIgA, pIgA and IgG were compared in normal mice. Anti-CEA pIgA tumour uptake was studied in mice bearing the WiDr caecal orthotopic graft. IgA radiolabelling was obtained with a high yield, was stable in PBS and murine plasma, and did not alter IgA binding functionality (Kd ≈ 25 nM). Biodistribution studies in normal mice confirmed that radiolabelled pIgA - and to a lesser extent, mIgA - showed strong and fast mucosal tropism and a shorter serum half-life than IgG. In caecal tumour model mice, evaluation of the anti-CEA-pIgA biodistribution showed a high uptake in lung metastases, confirmed by histological analysis. However, no radioactivity uptake increase in the tumoural caecum was discerned from normal intestinal tissue, probably due to high IgA caecal natural tropism. In microSPECT/CT imaging, 99mTc-IgA confirmed its diagnostic potency of tumour in mucosal tissue, even if detection threshold by in vivo imaging was higher than post mortem studies. Contribution of the FcαRI receptor, studied with transgenic mouse model (Tsg SCID-CD89), did not appear to be determinant in 99mTc-IgA uptake. Pre-clinical experiments highlighted significant differences between 99mTc-IgA and 99mTc-IgG biodistributions. Furthermore, tumoural model studies suggested potential targeting potency of pIgA in mucosal tissues.

Anti-CD20 Immunoglobulin G Radiolabeling with a 99mTc-Tricarbonyl Core: In Vitro and In Vivo Evaluations.

In recent years, the diagnostic and therapeutic uses of radioisotopes have shown significant progress. Immunoglobulin (Ig) appears to be a promising tracer, particularly due to its ability to target selected antigens. The main objective of this study is to optimize and assess an Ig radiolabeling method with Technetium 99m (99mTc), an attractive radioelement used widely for diagnostic imaging. Monoclonal anti-CD20 IgG was retained to study in vitro and in vivo radiolabeling impact. After IgG derivatization with 2-iminothiolane, IgG-SH was radiolabeled by an indirect method, using a 99mTc-tricarbonyl core. Radiolabeling stability was evaluated over 24h by thin-layer chromatography. IgG integrity was checked by sodium dodecyl sulfate-polyacrylamide gel electrophoresis coupled with Western blot and autoradiography. The radiolabeled Ig's immunoaffinity was assessed in vitro by a radioimmunoassay method and binding experiments with cells (EL4-hCD20 and EL4-WT). Biodistribution studies were performed in normal BALB/c mice. Tumor uptake was assessed in mice bearing EL4-hCD20 and EL4-WT subcutaneous xenografts. With optimized method, high radiolabeling yields were obtained (95.9 ± 3.5%). 99mTc-IgG-SH was stable in phosphate-buffered saline (4°C and 25°C) and in serum (37°C), even if important sensitivity to transchelation was observed. IgG was not degraded by derivatization and radiolabeling, as shown by Western blot and autoradiography results. 99mTc-anti-CD20 IgG-SH immunoaffinity was estimated with Kd = 35 nM by both methods. In vivo biodistribution studies for 48h showed significant accumulation of radioactivity in plasma, liver, spleen, lungs and kidneys. Planar scintigraphy of mice bearing tumors showed a significant uptake of 99mTc-anti-CD20 IgG-SH in CD20+ tumor versus CD20- tumor. Radiolabeling of derivatized IgG with 99mTc-tricarbonyl was effective, stable and required few antibody amounts. This attractive radiolabeling method is "antibody safe" and preserves Ig affinity for antigen, as shown by both in vitro and in vivo experiments. This method could easily be used with noncommercial IgG or other antibody isotypes.

Adverse reactions to radiopharmaceuticals in France: analysis of the national pharmacovigilance database.

BACKGROUND: Radiopharmaceuticals are regarded as safe by the nuclear medicine community, but up to now, no survey has been conducted with from the perspective of pharmacovigilance. OBJECTIVE: To describe the adverse reactions to radiopharmaceuticals (ARRPs) reported to the French Pharmacovigilance Database (FPVD). METHODS: We selected and described all reports encompassing at least one radiopharmaceutical in the FPVD. The annual incidence of reported ARRPs used in diagnosis was also estimated. RESULTS: From 1989 to 2013, 304 reports of ARRPs were identified (43.0% serious, 12 deaths) in 54.6% women and 45.4% men; the median age was 58 years. Five therapeutic radiopharmaceuticals ((131)I-sodium iodide, (131)I-lipiodol, (89)Sr-chloride, (153)Sm-lexidronam, and (90)Y-ibritumomab-tiuxetan) were involved in 48 reports (97 adverse reactions: 86.6% serious, 9 deaths). Pulmonary disorders represented 44.3% of ARRPs used for therapy, mainly related to (131)I-lipiodol. There were 34 diagnostic radiopharmaceuticals involved in 256 reports (451 adverse reactions: 38.1% serious, 3 deaths); 8 diagnostic products ((99m)Tc-oxidronate, (18)F-fluorodeoxyglucose, (99m)Tc-tin pyrophosphate, (99m)Tc-tetrofosmin, (99m)Tc-dimercaptosuccinic acid, (201)Tl-chloride, (99m)Tc-sestamibi, and (111)In-pentetate) accounted for two-thirds of ARRPs. The most frequent adverse reactions were skin (34.4%), general (18.2%), nervous (9.0%), and gastrointestinal disorders (7.0%). There were 25 cases of altered images and 10 medication errors. The annual incidence of reported adverse reactions ranged from 1.2 × 10(-5) to 3.4 × 10(-5) diagnostic administrations. CONCLUSIONS: Reported ARRPs occurred rarely and were more serious in the therapeutic than in the diagnostic field. The notification of ARRPs was able to provide new guidance for safe use, as was the case for (131)I-lipiodol. Therefore, it is important to report ARRPs to a pharmacovigilance system.

Effect of levodopa on pain threshold in Parkinson's disease: a clinical and positron emission tomography study.

Patients suffering from Parkinson's disease (PD) frequently experienced painful sensations that could be in part due to central modification of nociception. We compared pain threshold before and after administration of levodopa in PD patients and in controls, and investigated cerebral activity with positron emission tomography (PET) during experimental nociceptive stimulation. Pain threshold was determined using thermal stimulation during two randomized conditions: off and on. We performed H(2) (15)O PET analysis of regional cerebral blood flow on subjects while they received alternate randomized noxious and innocuous stimuli during off and on conditions. In off condition, pain threshold in nine PD patients was significantly lower than in nine controls. Administration of levodopa significantly raised pain threshold in PD patients but not in controls. During off condition, there was a significant increase in pain-induced activation in right insula and prefrontal and left anterior cingulate cortices in PD compared to control group. Levodopa significantly reduced pain-induced activation in these areas in PD. This study shows that pain threshold is lower in PD patients but returns to normal ranges after levodopa administration. Moreover, PD patients have higher pain-induced activation in nociceptive pathways, which can be reduced by levodopa.

Comparison of pharmacological activities of Neuropeptide FF1 and Neuropeptide FF2 receptor agonists.

The pharmacological effects of Neuropeptide FF (NPFF) analogs exhibiting different selectivities towards Neuropeptide FF1 (NPFF1) and Neuropeptide FF2 (NPFF2) receptors were investigated after supraspinal administration in mice. Injected into the third ventricle, VPNLPQRF-NH2, which is selective for Neuropeptide FF1 receptor induced a hypothermia while EFWSLAAPQRF-NH2 and SPAFLFQPQRF-NH2 which are selective for Neuropeptide FF2 receptor, did not. Furthermore, EFWSLAAPQRF-NH2 significantly increased the body temperature when compared to saline treated mice, indicating that Neuropeptide FF1 receptor could be responsible for hypothermia while Neuropeptide FF2 mediated an hyperthermic effect. After administration into the lateral ventricle, 1DMe ([D.Tyr1,(N.Me)Phe3]NPFF), a weakly selective Neuropeptide FF2 receptor agonist, exerted a clear anti-opioid effect in the tail flick test. The selective Neuropeptide FF1 receptor agonist VPNLPQRF-NH2 did not induce significant anti-opioid actions but rather increased, dose-dependently, morphine analgesia while EFWSLAAPQRF-NH2, the highest selective Neuropeptide FF2 receptor analog, induced the same pharmacological effect as VPNLPQRF-NH2 at comparable doses. These features indicate that the pro- and anti-opioid actions are not strictly related to the selectivity towards Neuropeptide FF2 or Neuropeptide FF1 receptor. Our data demonstrate that Neuropeptide FF1 and Neuropeptide FF2 receptors differently modulate nervous system functions.

Pharmacological characterization of human NPFF(1) and NPFF(2) receptors expressed in CHO cells by using NPY Y(1) receptor antagonists.

Neuropeptide FF (NPFF) belongs to an opioid-modulatory system including two precursors (pro-NPFF(A) and pro-NPFF(B)) and two G-protein coupled receptors (NPFF(1) and NPFF(2)). The pharmacological and functional profiles of human NPFF(1) and NPFF(2) receptors expressed in Chinese hamster ovary (CHO) cells were compared by determining the affinity of several peptides derived from both NPFF precursors and by measuring their abilities to inhibit forskolin-induced cAMP accumulation. Each NPFF receptor recognizes peptides from both precursors with nanomolar affinities, however, with a slight preference of pro-NPFF(A) peptides for NPFF(2) receptors and of pro-NPFF(B) peptides for NPFF(1) receptors. BIBP3226 ((R)-N(2)-(diphenylacetyl)-N-[(4-hydroxyphenyl)-methyl]-argininamide) and BIBO3304 ((R)-N(2)-(diphenylacetyl)-N-[4-(aminocarbonylaminomethyl)-benzyl]-argininamide trifluoroacetate), two selective neuropeptide Y (NPY) Y(1) receptor antagonists, display relative high affinities for NPFF receptors and exhibit antagonist properties towards hNPFF(1) receptors. The structural determinants responsible for binding of these molecules to NPFF receptors were investigated and led to the synthesis of hNPFF(1) receptor antagonists with affinities from 40 to 80 nM. Our results demonstrate differences in pharmacological characteristics between NPFF(1) and NPFF(2) receptors and the feasibility of subtype-selective antagonists.

Dissociation of pharmacological pro- and anti-opioid effects by neuropeptide FF analogs.

Neuropeptide FF (NPFF) and its analog 1DMe ([D-Tyr(1),(NMe)Phe(3)]NPFF) have been shown to reverse or potentiate morphine analgesia in rat depending on the supraspinal or spinal site of injection. The properties, in the mouse tail-flick test, of 1DMe and its related compound Nic-1DMe (Nicotinoyl-Pro-1DMe) were investigated after their local (i.c.v. and i.t.) and systemic administration. Whereas Nic-1DMe and 1DMe exhibit the same affinity and selectivity towards NPFF(1) and NPFF(2) receptors, Nic-1DMe, in contrast to 1DMe, is unable to inhibit morphine-induced analgesia after i.c.v. and i.p. administration. Conversely, after i.t. and i.p. administration, both neuropeptide FF analogs could potentiate morphine analgesia. Differences in disposition parameters between 1DMe and Nic-1DMe are evidenced, suggesting that the two neuropeptide FF analogs could stimulate differentially supraspinal neuropeptide FF receptors. The predominant activation of spinal neuronal pathways by Nic-1DMe could explain the selective pro-opioid action of this compound after i.t., i.c.v. and i.p. administration.