Optimizing conditions for radiolabelling DTPA-bombesin analogues with In-111 at high specific activity

2010-01-01T00:00:00Z

Title: Optimizing conditions for radiolabelling DTPA-bombesin analogues with In-111 at high specific activity Authors: Pujatti, Priscilla Brunelli; Araújo, Elaine Bortoleti de; Mengattim, Jair Abstract: In designing radiometal-based peptides, an important factor to consider is the specific activity (SA) of the molecule. Low SA can compromise the uptake of the tracer in the tissue of interest in vivo and lead to physiological responses due to the presence of the cold peptide in the organism. However, very high SA can cause radiolysis of the compound, resulting in undesirable impurities. In this study, parameters influencing the kinetics of labelling of a DTPA-bombesin analogue (BETG5) with 111In were investigated and conditions were optimized to obtain the highest achievable SA. The effects of peptide mass, 111In activity, temperature and time of reaction in radiolabelling yield and peptide integrity were systematically categorized applying ITLC-SG and HPLC as chromatography techniques. Kinetics of 111In-labelling was optimal at pH 4.5, room temperature and 15 min of incubation. Higher radiochemical purities were obtained when 10 μg of BETG5 reacted with 37–555 MBq of radiometal (maximum SA 111 GBq/ μmol). 111In-BETG5 integrity analysis suggested that oxidation does not depend on radiolabelling conditions and can be avoided by the addition of methionine in radiolabelling medium. These optimized conditions will be applied to produce high specific activity DTPA-bombesin analogues for preclinical studies in healthy and tumour mice.

Cellular and biochemical responses induced by Biotherapics prepared from intact influenza A (H3N2) and inactivated influenza A (H3N2) virus at 12x and 30x in the MDCK cells.

2011-01-01T00:00:00Z

Title: Cellular and biochemical responses induced by Biotherapics prepared from intact influenza A (H3N2) and inactivated influenza A (H3N2) virus at 12x and 30x in the MDCK cells. Authors: Siqueira, Camila Monteiro; Mendonça, Rafaela Amaral Furtado de; Veiga, Venicio Feo da; Brunow, Mariah Celestino Marcondes; Zancan, Patrícia; Couceiro, José Nelson; Holandino, Carla Abstract: Biotherapics are homeopathic remedies prepared from organic products that are chemically undefined and can be used for treatment of diseases like influenza. There are several classes of biotherapics and, among these, there are some called "living biotherapics" or "Roberto Costa’s Biotherapics". This study aimed to compare the cellular and biochemical effects of biotherapics prepared from intact influenza virus diluted in water and the one obtained from the same viral sample inactivated by ethanol 70% (v / v), both in the potencies of 12x and 30x. Transmission electron microscopy (TEM) analyses were performed on both preparations to assess the integrity of viral particles, which showed that ethanol 70% (v/v) induced a complete denaturation of viral particles. In contrast, the integrity of virus particles was preserved when water was used as the biotherapic solvent. Cellular and biochemical alterations induced by the preparations on MDCK cells were analyzed and compared with those induced by respective controls (water 30x-treated and untreated cells). Cellular viability analyzed by MTT method showed statistically significant differences (p <0.05) in MDCK cells treated with intact biotherapic for 5 (3 stimuli) and 30 (18 stimuli) days in comparison with untreated control. TEM analysis did not show significant cellular changes when the different experimental groups were compared. The enzymatic activity of phosphofructokinase 1 (PFK), an important enzyme in the glycolytic pathway, presented a statistically significant increase (p <0.05) after 30 days of treatment when compared to control groups. The results obtained suggest that inactivation of viral sample with ethanol 70% induces lysis and disruption of viral particles. In addition, preliminary results indicated that treatment with intact biotherapic seems to induce higher variations on MDCK cells responses when compared to inactivated biotherapic-treated cells. Further analyses are ongoing, including scanning electron microscopy and quantification of the number of mitosis, in order to elucidate the mechanisms involved with biochemical and cellular responses induced by theses biotherapics. Description: 2 p.

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Optimizing conditions for radiolabelling DTPA-bombesin analogues with In-111 at high specific activity

Cellular and biochemical responses induced by Biotherapics prepared from intact influenza A (H3N2) and inactivated influenza A (H3N2) virus at 12x and 30x in the MDCK cells.