Fermentation of CHO cells for biomass component quantification

Knez, Špela (author), Narat, Mojca (mentor), Borth, Nicole (comentor)

Chinese Hamster Ovary (CHO) cells are the main production host for recombinant therapeutic proteins. They have desirable process performance characteristics such as high expression rates, rapid growth and required post- translational modification. To further exploit its potential, genome scale metabolic model was made for CHO-K1 cell line. Since model is lacking correct biomass values of CHO-K1, which could improve model predictions, focus of this study was to determine biomass and biomass components during a batch process in exponential phase for producing and non producing cell line, grown with and without 8 mM Glutamine. For that, bioprocesses of each condition were performed and sampled 3 times per day to obtain data for growth characteristics, extracellular metabolites and to determine biomass composition. Biomass equation and biomas formula were calculated for each day and condition. Data show cell dry mass varies between strains, yet differences are not significant. Biomass compositional analysis revealed only small differences in amino acid composition between different CHO cell lines, but the composition changes over the time course. As the macromolecular composition changes over the batch, so must the energy requirements to synthesize these macromolecules. While results presented in this thesis may show a need to determine only proteins content for each cell line to improve cell specific predictions, further studies as energy maintenance and uptake/secretion rates are still needed to improve model predictions.

• Language: English
• Keywords: Biotechnology, bioprocessing, CHO, modeling, biomass
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: BF - Biotechnical Faculty
• Publisher: [Š. Knez]
• Year: 2019
• PID: 20.500.12556/RUL-109719
• UDC: 602.42:577.152.3(043.2)
• COBISS.SI-ID: 9371769
• Publication date in RUL: 07.09.2019

Secondary language

• Language: Slovenian
• Title: Kvantifikacija komponent biomase v bioprocesu s celicami CHO

Abstract

Celice ovarijev kitajskega hrčka (CHO) so trenutno glavni gostitelj za produkcijo rekombinantnih proteinov saj imajo dobre procesne karakteristike, tj. visoka stopnja produkcije rekombinantnih proteinov, hitra rast in zaželene post translacijske modifikacije. Metabolni model genoma CHO-K1 celične linije je omogočil nadaljnjo izkoriščanje potenciala linije. Kljub obetajočim napovedim pa model ne vsebuje točnih rezultatov za biomaso celic, kar bi še izboljšalo napovedno vrednot modela. Namen te raziskave je bil preučiti spremembo biomase v bioprocesu z ali brez dodatka 8 mM Glutamina pri dveh celičnih linijah, matični CHO-K1 in rekombinantni DXB11. Tekom procesa so bili vzeti vzorci z namenom določiti celično rast, zunaj celične metabolite in sestavo biomase celic: DNA, RNA, proteine, lipide in ogljikove hidrate. Iz pridobljenih podatkov smo izračunali formulo biomase za vsak dan tekom eksponentne faze bioprocesa. Podatki kažejo, da je biomasa celic med linijami različna ter da se spreminja tekom procesa. Analiza sestave biomase je pokazala, da so med linijami manjše razlike v vsebnosti amino kislin ter, da se vsebnost spreminja tekom procesa. Energetska potreba se tako spreminja tekom procesa skupaj s sestavo makromolekul. Za izboljšanje napovedi so tako potrebne še nadaljnje analize energetske potrebe in študij hitrosti vnosa in iznosa metabolitov.

• Keywords: biotehnologija, bioprocesništvo, CHO, modeliranje, biomasa