PRISCILA DE QUEIROZ SOUZA PASSOS
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Artigo IPEN-doc 28654 In vitro and in vivo response of PSMA-617 radiolabeled with CA and NCA lutetium-1772022 - BOAS, CRISTIAN A.W.V.; SILVA, JEFFERSON de J.; DIAS, LUIS A.P.; FREIRE, MARIA R.B.; BALIEIRO, LUIZA M.; SANTOS, CAROLINA S.F. dos; VIVALDINI, BIANCA F.; BENEDETTO, RAQUEL; VIEIRA, DANIEL P.; PASSOS, PRISCILA de Q.S.; MARUMO, MARIA H.; TEIXEIRA, LUIS F.S.; ARAUJO, ELAINE B. deThe PSMA-targeted radionuclide therapy has been explored since 2015 with radioisotope lutetium-177, whose β− emission range is adequate for micrometastases treatment. This radioisotope is obtained by two different production routes that directly affect the specific activity of lutetium-177 (non-carrier added and carrier added) and, consequently, the specific activity of radiopharmaceuticals, like 177Lu-PSMA-617. The influence of the specific activity of lutetium-177 on the properties of the radiopharmaceutical PSMA-617 was evaluated through pre-clinical studies. The in vitro study pointed to a lower constant of dissociation with non-carrier added lutetium-177 due to the difference in the specific activity. However, competition and internalization assays resulted in similar results for both lutetium-177. Based on these pre-clinical experiments, the total in vitro tumor cell binding and tumor uptake in vivo were similar, with no influence of the specific activity of the 177Lu-PSMA-617. Regardless the specific activity did not directly affect tumor uptake, the tumor/non-target organs ratios were higher for the radiopharmaceutical labeled with carrier added lutetium-177, which had the lowest specific activity.Resumo IPEN-doc 27446 In vitro response of 177Lu-PSMA-617 with two different specific activities2020 - VILLAS BOAS, CRISTIAN A.W.; MENGATTI, JAIR; PASSOS, PRISCILA; VIEIRA, DANIEL; ARAUJO, ELAINE B. deIntroduction: PSMA-617 radiolabeled with lutetium-177 has shown good results in compassionate studies around the world. Being a receptor-specific radiopharmaceutical, the specific activity (SA) of the preparation may represent an important factor for therapeutic efficacy. Lutetium-177 can be produced by two different routes: with ytterbium-176 (Non-carrier-added or NCA) and with lutetium-176 (Carrier-added or CA). The SA (MBq/ug) of the labeled PSMA varies accordingly to each lutetium. For NCA lutetium, the radiolabeling procedure is based on the SA of 74 MBq/ug. When the radiolabeling is performed with CA lutetium, SA is determined by the molar ratio of 2.1:1 (PSMA moles/lutetium moles declared in the certificate), resulting in lower SA than NCA. This work evaluated the influence of specific activity of 177Lu-PSMA-617 on in vitro specific binding assays (saturation, competition and internalization). Materials and Methods: Radiolabeling of PSMA-617 (ABX, Germany) with lutetium-177 was performed in heating block at 90°C for 30 minutes with sodium ascorbate (0.5 M pH 4.7) as buffer. For NCA lutetium (JSC, Russia) the radiopharmaceutical specific activity was 74 MBq/ug. For CA lutetium (IDB, Netherlands), the specific activity was 41 MBq/ug. The radiochemical purity was analyzed with HPLC. For all experiments, 6-well plates were used for adherence cells with 200,000 LNCaP per well. Molar concentration of saturation curves experiments were 0.01; 0.05; 0.6; 1.5; 3.0 and 3.5 for CA lutetium and 0.1; 0.6; 1.5; 2.0; 2.5 and 3.0 for NCA lutetium. After 1 hour of incubation at 8 ºC, supernatant was removed, then washed with PBS (phosphate buffer saline) and finally cells were burst with NaOH 1 M, and activity was measured in gama counter; the experiments were performed in octuplicate. Competition experiments were performed adding in all wells 5 nM of radiolabeled PSMA-617 and in the competition well (non-specific binding) were added an excess of 15 times (76 ug) of non radiolabeled PSMA-617. After 1 hour of incubation at 8 ºC, supernatant was removed, then washed with PBS and finally cells were burst with NaOH 1 M, and activity was measured in gama counter, these experiments were performed in triplicate. The specific binding was obtained by the difference between total binding and non-specific binding. Internalization experiments were performed at Kd of NCA and CA lutetium. After 1 hour of incubation at 37 ºC, supernatant was removed, washed with PBS, then washed again with 0.05 M glycine solution pH 2.8 and finally cells were burst with NaOH 1 M. Activity was measured in gama counter, these experiment were performed in sextuplicate. Results and discussion: The radiochemical purity were 98% and 99% for labeling with NCA and CA lutetium, respectively. Saturation curve assay with NCA lutetium shown a Kd of 0.7 ± 0.15 nM and a Bmax of 857 ± 55.79 pMol/ng, and with CA lutetium resulted in a Kd of 1.71 ± 0.45 nM and a Bmax of 1156 ± 113.8 pMol/ng. The variation between both Kd curves were statistically different (P value = 0.0058). Competition assay demonstrated an effective blocking for both types of lutetium, for NCA unpaired T test shown a P value of 0.0011. For CA lutetium, the unpaired test disclosed a P value of 0.0258. The comparison between both results revealed a P value of 0.01 at the specific binding. Internalization assay shown for both types of lutetium similar results, 27.1 ± 2.45% and 30.6 ± 4.97%, for CA and NCA lutetium, respectively, and was not statistically significant (P value = 0.17). Conclusions: These experiments demonstrated that the type of lutetium (CA or NCA) directly affects in vitro binding of 177Lu-PSMA-617 to receptors expressed in LNCaP cells. It was statistically demonstrated that the higher specific activity of 177Lu-PSMA-617, more radiolabeled peptide can bind to cells at saturation and competition assays.Artigo IPEN-doc 26273 Synthesis of paramagnetic iron oxide nanoparticles for application in in vitro three-dimensional biological models through electron beam irradiation and microwave reduction of iron ions2019 - PASSOS, PRISCILA de Q.S.; CORAZZA, FULVIO G.; LIMA, MAYELLE M.P.; TOMINAGA, FLAVIO K.; SAKATA, SOLANGE K.; GONÇALVES, KARINA O.; COURROL, LILIA C.; VIEIRA, DANIEL P.Three-dimensional (3D) cell culture is increasingly being used in assays to assess the safety and efficacy of new drug candidates. Tumor cell spheroids can mimic with high precision the biological complexity of cellular interactions with their tumor microenvironment. Currently, several techniques can be used to construct 3D spheroids. Among them, magnetic levitation is one of the most used in biomedical research. This technique consists in the magnetization of cells through the adsorption of magnetic nanoparticles of iron oxide (Fe3O4) that are produced by the reaction of Fe2+ and Fe3+ ions in alkaline medium. In this work, nanoparticles of paramagnetic iron oxide (PIONS) were synthesized by coprecipitation through electron beam irradiation at 15 and 30 kGy doses. After functionalization with polar amino acids, nanoparticle suspensions were characterized by physical-chemical assays that showed the successful attachment of the carboxylate groups to the iron, explaining the ability of the particles to adsorb the membranes. Cytotoxicity assay showed that the nanoparticles synthesized by microwave (MW) and electron beam had no toxicity. Others biological assays have also shown efficient adsorption of the particles by human prostate tumor cells, allowing the in vitro application of a biomimetic 3D biological model with potential utilization regarding the development and evaluation of antitumor drugs and radiopharmaceuticals for the treatment of prostate cancer.