[en] The bismuth-213 is an alpha- and beta-emitting radioelement of very short physical half-life (45 min) obtained by means of a (²²⁵Ac-²¹³Bi) generator. Given its radiotoxicity, this element presents an interest in radioimmunotherapy (RIT). At present, the DTPA derivatives alone are used in radiolabelling of antibodies for RIT. This study presents the complexing properties of other chelates potentially usable to this goal. Four original chelating agents were synthesized in order to choose the families giving the best results in complexing the ²¹³Bi: the tri-ethylene-tetra-amino-hexa-carboxylic acid (HETA), the 1, 4, 7, 10-tetra-aza-cyclo-dodecane-1, 4, 7, 10-tetraacetic- 1- Gly-L-p-nitro Phe-amide (DOTA-pept), the 1, 4, 8, 11-tetrakis [(S)-2 hydroxy-propyl]-1, 4, 8, 11-tetra-aza-cyclo-tetra-decane (THEOH), and the ethylenediamine di-acetate di-acetamide-bis-thiophenol (EDTA-TH). Given the physical characteristics of ²¹³Bi and the goal of our research, the studies of complexation were conducted by C.C.M. on silica, in highly diluted solution, with fixed chelates-²¹³Bi incubation time (15 min), and variable temperature and concentrations. Analysis of the results have been done by means of a phosphor-imager by measuring the number of pixels associated to every chromatographic spot. The obtained results show that the poly-aza poly-carboxylic derivatives and poly-aza di-thio dicarboxylic derivatives are the only able ones to complex rapidly and quantitatively the ²¹³Bi

No abstract available

[en] The efficiency of the radioimmunotherapy (RIT) using beta emitters has been clinically proved in treatments of refractory forms of lymphoma. The alpha-emitting radioelements of short half-life are also good potential candidates for RIT, applicable to tumor targets accessible rapidly to the molecules of the radio-immuno-conjugates of size compatible with the short range of alpha particles (50 to 80 μm). The goal of this study is to demonstrate the feasibility of such an approach on a model of myeloma multiply targeted by specific antibodies (B-B4) coupled to bismuth-213 with a chelating agent (benzyl-DTPA). The efficiency of the alpha RIT was evaluated in vitro by means of different techniques analyzing the cellular mortality (the method of limited dilution), the effects on DNA (the testing of micro-nuclei), the analysis of radio-induced apoptosis (the test with acridine orange) and finally the study of non-specific irradiation on population of cells of hematopoietic system un-recognized by the B-B4 benzyl-DTPA) immuno-conjugate. The first results have shown besides the technical feasibility of the project a strong dose dependent cellular mortality with a survival falling rapidly from 28% to around 1 o/oo for a single doubling of the dose from 14.8 kBq / 10⁵ cells (0.4 μCi) to 29.6 kBq/10⁵ cells (0.8 μCi). The cellular mortality was total at 300 kBq/10⁵ cells (8 μCi). The cells in an apoptosis state were evidenced at rates up to 40% for a dose of 7.4 kBq/10⁵ cells (0.2 μ Ci). New experiments will permit confirming these first results and determining the irradiation range having in view a utilization in protocols of purging of the myeloma cells on pockets obtained after plasmaphereses

[en] This article describes the principles of the positron emission tomography said phenotypic. Vectors considered for this kind of approach are peptides or hormones. Different radioisotopes are tackled for diagnosis (¹¹¹In, ⁶⁸Ga) or therapy(¹⁷⁷Lu, ⁹⁰Y). The comparisons are made with PET-F.D.G. (N.C.)

[en] The principle of radioimmunotherapy is to vectorize radioisotopes to the vicinity of tumor cells where radiation can destroy these targets. The most currently established protocols use beta-emitters. Recently, new RIT approaches using alpha radiation have been developed. These types of radiation, unlike beta emissions, have a high LET, i.e. the energy deposit related to the emission of an alpha particle is high and is performed over a short pathway of several tens of lm. Thus, this type of emission allows very localized irradiation to be performed, while preserving surrounding tissues, and cellular toxicity is achieved with only a few disintegrations at the cell surface. Unlike beta-emitters, alpha-emitters, because of their short range of action, seem more suitable for the treatment of diffuse cancers or micro metastasis subsequent to ablation of a primary tumour. However, the few radionuclides required for cell destruction in this case require very specific targeting of tumor cells. To date, alpha RIT is still at a pre-clinical stage of development: the radiolabeling methods need to be optimized to ensure in vivo stability of the radio-pharmaceuticals. The modalities of administration of radiolabeled antibodies in animal models require also to be improved for delivering higher doses to tumor targets. A comprehensive analysis of the specific events occurring at cell or tissue level in response to alpha irradiation would be of great interest in order to define the best therapeutic association, chemotherapy or the possibilities of initiating immune response to tumors in patients treated by alpha-RIT. (authors)

[en] Full text of publication follows. We have developed the use of Sac7d archaeal polypeptide and its homologues as a non-antibody scaffold from which artificial affinity proteins (Affitins) can be derived with a number of favorable properties. Affitins show affinity (sub-nanomolar) and specificity that compare well with those of antibodies [Ref.1]. They are thermally (up to 90 C. degrees) and chemically stable (pH 0-12+, denaturants), well expressed in E. coli (up to 200 mg/L), lack disulfide bridge and have a size compatible with chemical synthesis (7 kDa). We have demonstrated their use as reagents for intra-cellular inhibition [Ref.1], affinity purification [Ref.2], immuno-localization [Ref.3], protein chip array [Ref.4] and biosensors [Ref.5]. We have also shown that Affitins are plastic enough to tolerate several mutagenesis schemes while their fold and their favorable properties are conserved [Ref.6]. Compared to Affitins, monoclonal antibodies are 20 times larger, less stable and more complex molecules. Furthermore, the remarkable stability properties of Affitins make them suited for demanding labeling protocols that are usually used for peptides. All together, these results show that Affitins should be well suited for biomedical applications where fine tuning of the affinity reagent properties is needed. References: [Ref.1] Mouratou, B. et al., (2007), Proc Natl Acad Sci U S A 104, 17983-17988; [Ref.2] Krehenbrink, M. et al. (2008), J Mol Biol 383, 1058-1068; [Ref.3] Buddelmeijer, N. et al. (2009), J Bacteriol 191, 161-168; [Ref.4] Cinier, M. et al. (2009), Bioconjug. Chem. 20, 2270-2277; [Ref.5] Miranda, F. F. et al. (2011), Biosens. Bioelectron. 26, 4184-4190; [Ref.6] Behar G.et al. (2013), Protein Eng Des Sel. 26(4):267-75. (authors)

[en] The full text of the publication follows. Purpose: The purpose of this study was to analyze the radiosensitivity of human lymphocytes. T cells, whether activated or not, seems to be a good model to study the radiosensitivity of normal tissues. To clarify the mechanism of action of these radiations, we study genes modulation that provides a response to alpha particles. Materials and methods: Lymphocytes from healthy donors were isolated and half were subjected to polyclonal activation monitored by expression of CD25. Both the non-activated T lymphocytes (NATL) and activated lymphocytes (ATL) were incubated with BSA-CHX-DTPA coupled to ²¹³Bi (185 kBq/ml) or with BSA-SAB coupled to ²¹¹At (74 kBq/ml). Tests were performed on the cells of four different donors to assess mortality, redox activity, cell cycle distribution and DNA double strands breaks. Gene expression was investigated using the DNA micro-arrays. Results: Forty-eight hours after irradiation with ²¹³Bi (185 kBq/ml), mortality was 29.9 ±2.2% for NATL and 12.4 ± 7.6% for ATL. With ²¹¹At (74 kBq/ml), mortality was 44.3 ± 8.8% for NATL and 16.5 ± 8.2% for ATL. For the two alpha radionuclides irradiation, the cell cycle distribution of NATL remained quiescent and alpha irradiation did not trigger their activation; for ATL, irradiation gets blocked in G2/M. An increase of 15% in the number of DNA double strands was observed in two populations of lymphocytes at 42 hours when T cells were irradiated with ²¹³Bi, but the increase is less important when T cells were irradiated with ²¹¹At. Six genes were up regulated at least 1,5-fold: CDKN1A, DRAM, FAS, MDM2, PLK2, TNFRSF10B. Conclusion: In conclusion, we identified molecular and cellular mechanisms that cells trigger after alpha irradiation. Preliminary data indicate the involvement of pathways of apoptosis, autophagy and cell cycle. (authors)

[en] ARRONAX, acronym for ''Accelerator for Research in Radiochemistry and Oncology at Nantes Atlantique'', is a high energy and high intensity cyclotron. It will turn into operation in September 2009 in Nantes (France). It is mainly devoted to the production of radionuclides for medicine. A priority list based on the capability of the machine as well as on the need expressed by the European medical community through a questionnaire has been set. It contains isotopes for imaging (⁸²Sr/⁸²Rb and ₆₈Ge/⁶⁸Ga generators and ⁶⁴Cu, ⁴⁴Sc, ⁵⁵Co) and for therapeutic use (⁶⁷Cu, ⁴⁷Sc and ²¹¹At). In this list, a special attention has been paid to dosimetry, both by giving lower scores of interest to radionuclides with undesirable emissions, such as high energy gammas, and by promoting β+/β- couples of radionuclides (⁶⁴Cu/⁶⁷Cu and ⁴⁴Sc et ⁴⁷Sc). Indeed, pre-therapeutic dosimetry is essential in the development of new radiolabeled therapeutics and PET imaging should prove more precise and accurate in calculating radiation doses received by critical organs and tumors. Arronax and its partners will also devote a large effort to develop targeted alpha-radionuclide therapy. In particular, a radioimmunotherapy project using ²¹¹At coupled to a specific monoclonal antibody will start this autumn to treat prostate cancer patients with high risk of relapse. ARRONAX is an important part of the project since it will have to produce astatine in large amounts on a regular basis. (author)

[en] The full text of the publication follows. Radioimmunotherapy (RIT) is an approach aiming at targeting the radioelements to tumours, usually through the use of antibodies specific for tumour antigens. The radiations emitted by the radioelements then induce direct killing of the targeted cells as well as indirect killing through bystander effect. Interestingly, it has been shown that ionizing radiations, in some settings of external radiotherapy, can foster an immune response directed against tumour cells. Our research team is dedicated to the development of alpha RIT, i.e RIT using alpha particle emitters, we therefore decided to study the effects of such particles on tumour cells in regards to their immunogenicity. First, we studied the effects of bismuth 213, an alpha emitter, on cellular death and autophagy in six different tumour cell lines. Then, we measured the expression of 'danger' signals and MHC molecules at the cell surface to determine whether irradiation with ²¹³Bi could cause the tumour cells to be recognized by the immune system. Finally a co-culture of dendritic cells with irradiated tumour cells was performed to test whether it would induce dendritic cells to mature. No apoptosis was detected within 48 hours after irradiation in any cell line, however half of them exhibited signs of autophagy. No increase in membrane expression of 'danger' signals was observed after treatment with ²¹³Bi, but we showed an increase in expression of MHC class I and II for some cell lines. Moreover, the co-culture experiment indicated that the immunogenicity of a human adenocarcinoma cell line (LS 174T) was enhanced in vitro after irradiation with alpha rays. These preliminary data suggest that alpha particles could be of interest in raising an immune response associated to RIT. (authors)

No abstract available