Multivariate data analysis of capacitance frequency scanning for online monitoring of viable cell concentrations in small-scale bioreactors.

Viable cell concentration (VCC) is one of the most important process attributes during mammalian cell cultivations. Current state-of-the-art measurements of VCC comprise offline methods which do not allow for continuous process data. According to the FDA's process analytical technology initiative, process monitoring and control should be applied to gain process understanding and to ensure high product quality. In this work, the use of an inline capacitance probe to monitor online VCCs of a mammalian CHO cell culture process in small-scale bioreactors (250 mL) was investigated. Capacitance sensors using single frequency are increasingly common for biomass monitoring. However, the single-frequency signal corresponds to the cell polarization that represents the viable cell volume. Therefore single-frequency measurements are dependent on cell diameter changes. Measuring the capacitance across various frequencies (frequency scanning) can provide information about the VCC and cope with changing cell diameter. Applying multivariate data analysis on the frequency scanning data successfully enabled direct online monitoring of VCCs in this study. The multivariate model was trained with data from 5 standard cultivations. The model provided a prediction of VCCs with relative errors from 5.5 to 11%, which is a good agreement with the acceptance criterion based on the offline reference method accuracy (approximately 10% relative error) and strongly improved compared with single-frequency results (16 to 23% relative error). Furthermore, robustness trials were conducted to demonstrate the model's predictive ability under challenging conditions. The process deviations in regard to dilution steps and feed variations were detected immediately in the online prediction of the VCC with relative errors between 6.7 and 13.2%. Thus in summary, the presented method on capacitance frequency scanning demonstrates its suitability for process monitoring and control that can save batches, time, and cost. Graphical abstract.

Magnetic Nanoparticle-Based Hyperthermia Mediates Drug Delivery and Impairs the Tumorigenic Capacity of Quiescent Colorectal Cancer Stem Cells.

Cancer stem cells (CSCs) are the tumor cell subpopulation responsible for resistance to chemotherapy, tumor recurrence, and metastasis. An efficient therapy must act on low proliferating quiescent-CSCs (q-CSCs). We here investigate the effect of magnetic hyperthermia (MHT) in combination with local chemotherapy as a dual therapy to inhibit patient-derived colorectal qCR-CSCs. We apply iron oxide nanocubes as MHT heat mediators, coated with a thermoresponsive polymer (TR-Cubes) and loaded with DOXO (TR-DOXO) as a chemotherapeutic agent. The thermoresponsive polymer releases DOXO only at a temperature above 44 °C. In colony-forming assays, the cells exposed to TR-Cubes with MHT reveal that qCR-CSCs struggle to survive the heat damage and, with a due delay, restart the division of dormant cells. The eradication of qCR-CSCs with a complete stop of the colony formation was achieved only with TR-DOXO when exposed to MHT. The in vivo tumor formation study confirms the combined effects of MHT with heat-mediated drug release: only the group of animals that received the CR-CSCs pretreated, in vitro, with TR-DOXO and MHT lacked the formation of tumor even after several months. For DOXO-resistant CR-CSCs cells, the same results were shown, in vitro, when choosing the drug oxaliplatin rather than DOXO and applying MHT. These findings emphasize the potential of our nanoplatforms as an effective patient-personalized cancer treatment against qCR-CSCs.