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Študija prisilne razgradnje terapevtskega monoklonskega protitelesa
ID Vovk, Eva (Author), ID Jamnik, Polona (Mentor) More about this mentor... This link opens in a new window, ID Igor Kristian, Štern (Comentor)

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Abstract
Med razvojem terapevtskih rekombinantnih monoklonskih protiteles je prisilna razgradnja eno od ključnih orodij za pridobivanje informacij o molekuli, analitski metodi, razvoju procesa in proizvodnje. Zato je ne le priporočljivo, temveč zaželeno, da razvoj analitske metode vključuje prisilno razgrajene vzorce, saj to pomembno vpliva na razumevanje o zmožnostih in omejitvah uporabljenih metod. Namen magistrskega dela je bil izpostaviti biološko učinkovino stresnim pogojem, da bi ugotovili, ali in pri kateri izmed izbranih analitskih metod zaznamo spremembe pri določeni (tarčni) biološki učinkovini in v kolikšni meri. Izbrano biološko učinkovino smo izpostavili različnim dejavnikom: povišani temperaturi in različnim vrednostim pH, oksidantu ter reducentu z različnimi koncentracijami, različni jakosti svetlobe, enkratnemu in večkratnemu zamrzovanju in odmrzovanju, deglikozilaciji z encimom v različnih množinah, povišani množini glukoze ter kovinskima ionoma. Z različnimi kromatografskimi metodami (afinitetna kromatografija, kromatografija z obrnjeno fazo, gelska izključitvena kromatografija, kapilarna gelska elektroforeza pod nereducirajočimi in reducirajočimi pogoji) smo analizirali vzorce in ovrednotili povzročene spremembe. Pokazali smo, da so vse metode, razen afinitetne kromatografije, zmožne zaznavanja sprememb za izbrani protein. Hkrati pa smo ugotovili, da je v študiji vključen protein obstojen tudi pri ekstremnih laboratorijskih pogojih: stabilen je v širokem temperaturnem in pH-območju, pri izpostavitvi umetni svetlobi in pri večkratnem zamrzovanju in odmrzovanju.

Language:Slovenian
Keywords:biotehnologija, prisilna razgradnja, analitske metode, monoklonsko protitelo, farmacija, biofarmacevtiki
Work type:Master's thesis/paper
Typology:2.09 - Master's Thesis
Organization:BF - Biotechnical Faculty
Publisher:[E. Vovk]
Year:2019
PID:20.500.12556/RUL-110983 This link opens in a new window
UDC:602:543.54:616-097.3(043.2)
COBISS.SI-ID:9360249 This link opens in a new window
Publication date in RUL:21.09.2019
Views:1350
Downloads:321
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Secondary language

Language:English
Title:Forced degradation study of therapeutic monoclonal antibody
Abstract:
During the development of a therapeutic recombinant monoclonal antibody, the forced degradation is one of the key tools for obtaining information about the molecule, the analytical method and about the process development and production. Hence, it is recommended that the method development includes foced degradated samples for a better understanding of the method capability and limitations of the method. The aim of the master's thesis was to expose the drug substance to stress conditions in order to determine whether and which of the chosen analytical methods are stability indicating. The chosen drug substance was exposed to various stress conditions: higher temperature, different pH values, addition of an oxidant, addition of a reducing agent, exposure to light, freezing/thawing, (partial) enzymatic deglycosylation, addition of glucose and metal ions. Samples were analyzed with different chromatographic methods (affinity liquid chromatography, reversed-phase chromatography, size exclusion chromatography, capillary gel electrophoresis under non-reducing conditions and under reducing conditions) and the induced differences were evaluated. It has been shown that all the methods except the affinity chromatography are capable to detect changes in the drug substance. Furthermore, the study showed that the studied protein is quite stable under various extreme laboratory conditions: broad temperature and pH ranges, exposure to artificial light, multiple freeze/thaw cycles.

Keywords:biotehnology, forced degradation, analytical methods, monoclonal antibody, pharmacy, biopharmaceuticals

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