TY - JOUR

T1 - Kinetics of NK-92 growth and functionality in pseudo-static cultures

AU - García Aponte, Oscar Fabian

AU - Kozma, Bence

AU - Egger, Dominik

AU - Kasper, Cornelia

AU - Herwig, Christoph

N1 - Funding Information: This work was performed within the framework of the Competence Center CHASE GmbH, funded by the Austrian Research Promotion Agency (grant number 868615 ). The COMET Centre CHASE is funded within the framework of COMET – Competence Centers for Excellent Technologies by BMVIT , BMDW , the Federal Provinces of Upper Austria and Vienna.

PY - 2023/7

Y1 - 2023/7

N2 - Understanding the growth and functionality variation of lymphocytes during expansion is fundamental for process control and endpoint determination. However, monitoring CQAs along lymphocyte expansion requires offline cell characterization, hindering timely identification of process deviations and transitions. We developed a culturing protocol that allowed to investigate day-to-day variation of NK-92 cells’ cytotoxicity against K-562 cells. Replacing 20% of the culture with fresh medium every day while disrupting the formed lymphocyte aggregates, morphological and functionality changes of the NK-92 cells were induced. Our experiments revealed that the adaptation to the pseudo-static protocol divided the culture to two distinctive phases with exponential growth in each. In between, there was an intermittent drop in viability. Significant functionality fluctuations were observed, despite the regular stimulation strategy. Scale-independent descriptors, such as growth rate, were calculated for each phase and the whole culture and correlated with cell morphology parameters and functionality. A novel correlation was found between, the cell size-derived, biomass volume and cytotoxicity that was stronger during the first phase, but it was apparent throughout the entire culture. This was possible to capture, only because of the day-to-day monitoring of cytotoxicity. By utilizing the pseudo-static culturing protocol and the methodology, variables, such as biomass volume, were discovered and by this, we are getting closer to find performance indicators suitable to monitor CQAs in a straightforward, non-invasive manner.

AB - Understanding the growth and functionality variation of lymphocytes during expansion is fundamental for process control and endpoint determination. However, monitoring CQAs along lymphocyte expansion requires offline cell characterization, hindering timely identification of process deviations and transitions. We developed a culturing protocol that allowed to investigate day-to-day variation of NK-92 cells’ cytotoxicity against K-562 cells. Replacing 20% of the culture with fresh medium every day while disrupting the formed lymphocyte aggregates, morphological and functionality changes of the NK-92 cells were induced. Our experiments revealed that the adaptation to the pseudo-static protocol divided the culture to two distinctive phases with exponential growth in each. In between, there was an intermittent drop in viability. Significant functionality fluctuations were observed, despite the regular stimulation strategy. Scale-independent descriptors, such as growth rate, were calculated for each phase and the whole culture and correlated with cell morphology parameters and functionality. A novel correlation was found between, the cell size-derived, biomass volume and cytotoxicity that was stronger during the first phase, but it was apparent throughout the entire culture. This was possible to capture, only because of the day-to-day monitoring of cytotoxicity. By utilizing the pseudo-static culturing protocol and the methodology, variables, such as biomass volume, were discovered and by this, we are getting closer to find performance indicators suitable to monitor CQAs in a straightforward, non-invasive manner.

KW - Adoptive Cell Therapy (ACT)

KW - ATMPs (advanced therapy medicinal products)

KW - Cytotoxicity

KW - Kinetics

KW - Natural Killer Cells

KW - NK-92

UR - http://www.scopus.com/inward/record.url?scp=85152737845&partnerID=8YFLogxK

U2 - 10.1016/j.bej.2023.108929

DO - 10.1016/j.bej.2023.108929

M3 - Article

AN - SCOPUS:85152737845

VL - 196

JO - Biochemical engineering journal

JF - Biochemical engineering journal

SN - 1369-703X

M1 - 108929

ER -