RENATA DAMIANI
23 resultados
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Resumo IPEN-doc 24397 Carbohydrate composition and site-occupancy determination in pituitary and recombinant preparations of human thyrotropin2017 - BARTOLINI, PAOLO; RIBELA, MARIA T.C.P.; DAMIANI, RENATA; SILVA, FELIPE D.; LIMA, ELIANA R.; OLIVEIRA, JOAO E.; PERONI, CIBELE N.; TORJESEN, PETER A.; SOARES, CARLOS R.Human thyrotropin (hTSH) is a glycoprotein with three potential glycosylation sites: two in the -subunit and one in the -subunit. Carbohydrate site-occupancy is frequently neglected in glycoprotein characterization, even if related to folding, trafficking, initiation of inflammation, host defence and congenital disorders of glycosylation (CDG). For the first-time N-glycoprofiling analysis was applied to site-occupancy determination of two native pituitary hTSH, in comparison with three CHO-derived preparations of hTSH, a widely used biopharmaceutical. A single methodology provided: (i) average N-glycan mass; (ii) mass fraction of each monosaccharide and of sulfate; (iii) percent carbohydrate. The results indicate that occupancy (65–87%) and carbohydrate mass (12–19%) can be 34–57% higher in recombinant hormones. The average glycan mass is 24% lower in pituitary hTSH and contains ∼3-fold fewer moles of galactose (P < 0.005) and sialic acid (P < 0.01). The number of moles of fucose per mole of hTSH was found 2.5-fold higher in the pituitary preparations. One of these native preparations, presenting the smallest glycan mass, lowest occupancy, GalNAc, sulfate, Gal and sialic acid contents, also presented the lowest in vivo bioactivity and circulatory half-life. This methodology, extremely important for comparing a recombinant biopharmaceutical to its native equivalent, can be applied to any physiologically or clinical relevant glycoprotein.Artigo IPEN-doc 22995 N-Glycoprofiling analysis for carbohydrate composition and site-occupancy determination in a poly-glycosylated protein: human thyrotropin of different origins2017 - RIBELA, MARIA T.C.P.; DAMIANI, RENATA; SILVA, FELIPE D.; LIMA, ELIANA R.; OLIVEIRA, JOAO E.; PERONI, CIBELE N.; TORJESEN, PETER A.; SOARES, CARLOS R.; BARTOLINI, PAOLOHuman thyrotropin (hTSH) is a glycoprotein with three potential glycosylation sites: two in the -subunit and one in the -subunit. These sites are not always occupied and occupancy is frequently neglected in glycoprotein characterization, even though it is related to folding, trafficking, initiation of inflammation and host defense, as well as congenital disorders of glycosylation (CDG). For the first time N-glycoprofiling analysis was applied to the site-occupancy determination of two native pituitary hTSH, in comparison with three recombinant preparations of hTSH, a widely used biopharmaceutical. A single methodology provided the: (i) average N-glycan mass; (ii) mass fraction of each monosaccharide and of sulfate; and (iii) percent carbohydrate. The results indicate that the occupancy (65%–87%) and carbohydrate mass (12%–19%) can be up to 34%–57% higher in recombinant hormones. The average glycan mass is 24% lower in pituitary hTSH and contains ~3-fold fewer moles of galactose (p < 0.005) and sialic acid (p < 0.01). One of the two native preparations, which had the smallest glycan mass together with the lowest occupancy and GalNAc, sulfate, Gal and sialic acid contents, also presented the lowest in vivo bioactivity and circulatory half-life. The methodology described, comparing a recombinant biopharmaceutical to its native equivalent, can be applied to any physiologically or clinical relevant glycoprotein.Resumo IPEN-doc 20256 Effects of butyrate and manganese on productivity, sialylation, N-glycosylation site occupancy and biological properties of CHO-derived thyrotropin2014 - DAMIANI, RENATA; OLIVEIRA, JOAO E.; ALMEIDA, BEATRIZ E.; SANT'ANA, PATRICIA M.; DALMORA, SERGIO L.; BARTOLINI, PAOLO; RIBELA, MARIA T.C.P.Resumo IPEN-doc 16441 Glycosylation and pharmacokinetics of human thyrotropin2011 - DAMIANI, R.; OLIVEIRA, J.E.; ALMEIDA, B.E.; VORAUER-UHL, K.; BARTOLINI, P.; RIBELA, M.T.C.P.Resumo IPEN-doc 18702 Sodium butyrate greatly enhances human thyrotropin (hTSH) synthesis by serum-free CHO cell culture with alterations of the carbohydrate moiety2012 - DAMIANI, R.; OLIVEIRA, J.E.; ALMEIDA, B.E.; BARTOLINI, P.; RIBELA, M.T.C.P.Resumo IPEN-doc 18703 Quantitative analysis of different preparations of human thyrotropin (hTSH): a comparison between RP-HPLC and the in vivo bioassay based on thyroxine stimulaton mice2012 - ALMEIDA, B.E.; DAMIANI, R.; OLIVEIRA, J.E.; RIBELA, M.T.C.P.; BARTOLINI, P.Resumo IPEN-doc 12453 Influence of reduced Co2 environment on the secretion yield, potency and glycan structures of recombinant thyrotropin (hTSH) from CHO cells2007 - RIBELA, MARIA T.C.P.; OLIVEIRA, JOAO E.; DAMIANI, RENATA; CARVALHO, CRISTIANE M.; LHOTA, GABRIELE; VORAUER-UHL, KAROLA; BARTOLINI, PAOLOA consistent increase of — 60%, in the secretion yield of CHO-derived hTSH was observed by changing cell 'culture CO2 conditions from 5% CO2 to air enviroment (0.03 % CO2). The quality of the product obtained under both conditions was analysed for what concerns N-glycan structures, charge isomers and biological activity in comparison with a well known commercial preparation (Thyrogen). The N-glycans identified in the three preparations were of the complex type, presenting di-, tri- and tetraantennary structures, with variable level of sialylation. Considering the latter characteristic, which is directly related to in vivo bioactivity, the three preparations have practically an identical percentage (86-88%) of sialylated structures, with some difference in percentage of di- and tii- sialylated glycan. Monosialylated glycans (N2G2S1, N2G1S1 and N2G2S1F) represent the three most abundant structures with 68-69% of all identified forms in the three preparations. The main difference was found in terms of antennarity with 8-10% more N2 structures for hTSH-IPEN produced in the absence of CO2 (-0O2) and 7-9 % more N3 structures for hTSH-IPEN (+CO2) and Thyrogen. Also for what concerns the total percentage of neutral glycans (12- 14 %), the three preparations are practically identical. No remarkable difference in charge isomers was also observed between the three preparations, the isoelectric focusing (IEF) profiles showing six well visible bands in the 5.39 - 7.35 pI range, the three major bands focusing at pI 5.85, 6.20 and 6.63. A considerably different distribution, with more acidic forms was observed, though, for two native pituitary preparations of hTSH. When analyzed via a simple and precise single-dose bioassay, a slightly significant difference (p<0.02) in activity was found between the two IPEN preparations. hTSH-IPEN (+ CO2) potency was non significantly different from that of Thyrogen, both being 1.6-1.8-fold more potent than the native pituitary reference preparation. We can conclude that, at least in the case of CHO-derived hTSH, different production processes may not greatly affect its glycan structures, charge isomer distribution or biological activity.Resumo IPEN-doc 12452 A practical RP-HPLC method for recombinant human thyrotropin (hTSH) laboratory scale purification2007 - DAMIANI, RENATA; OLIVEIRA, JOAO E.; CARVALHO, CRISTIANE M.; PERONI, CIBELE N.; BARTOLINI, PAOLO; RIBELA, MARIA T.C.P.Resumo IPEN-doc 12451 Physico-chemical characterization of alpha and beta subunit of recombinant human glycoprotein hormones: hTSH and hLH2007 - CARVALHO, CRISTIANE M.; OLIVEIRA, JOAO E.; DAMIANI, RENATA; ALMEIDA, BEATRIZ E.; CECCHI, CLAUDIA R.; BARTOLINI, PAOLO; RIBELA, MARIA T.C.P.Resumo IPEN-doc 11009 High-level secretion of growth hormone by retrovirally transduced primary human keratinocytes: prospects for an animal model of cutaneous gene therapy2005 - PERONI, C.N.; CECCHI, C.R.; DAMIANI, R.; SOARES, C.R.J.; RIBELA, M.T.C.P.; ARKATEN, R.R.; BARTOLINI, P.
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