Culture of cancer spheroids and evaluation of anti-cancer drugs in 3D-printed miniaturized continuous stirred tank reactors (mCSTRs)

Abstract

Cancer continues to be a leading cause of mortality in modern societies; therefore, improved and more reliable in vitro cancer models are needed to expedite fundamental research and anti-cancer drug development. Here, we describe the use of a miniaturized continuous stirred tank reactor (mCSTR) to first fabricate and mature cancer spheroids (i.e, derived from MCF7 cells, DU145 cells, and a mix of MCF7 cells and fibroblasts), and then to conduct anti-cancer drug assays under continuous perfusion. This 3 mL mCSTR features an off-center agitation system that enables homogeneous chaotic laminar mixing at low speeds to support cell aggregation. We incubated cell suspensions for 3 days in ultra-low-adherence (ULA) plates to allow formation of discoid cell aggregates (~600 µm in diameter). These cell aggregates were then transferred into mCSTRs and continuously fed with culture medium. We characterized the spheroid morphology and the expression of relevant tumor biomarkers at different maturation times for up to 4 weeks. The spheroids progressively increased in size during the first 5 to 6 days of culture to reach a steady diameter between 600 and 800 µm. In proof-of-principle experiments, we demonstrated the use of this mCSTR in anti-cancer drug testing. Three drugs commonly used in breast cancer treatment (doxorubicin, docetaxel, and paclitaxel) were probed at different concentrations in MCF7 derived spheroids. In these experiments, we evaluated cell viability, glucose consumption, spheroid morphology, lactate dehydrogenase activity, and the expression of genes associated with drug resistance (ABCB1 and ABCC1) and anti-apoptosis (Bcl2). We envision the use of this agitated system as a tumor-on-a-chip platform to expedite efficacy and safety testing of novel anti-cancer drugs and possibly in personalized medicine applications.

Novel quinazoline-1,2,3-triazole hybrids with anticancer and MET kinase targeting properties

“…Additionally, we observed that the 3D spheroid culture of AsPC-1 cells was more resistant to tested compounds compared to the same cells when grown in 2D cultures as IC 50 values obtained for both compounds as well as cabozantinib (positive control) were elevated in comparison to monolayer cultures. This is consistent with the results observed in other studies 35 , 37 and could be explained by features such as a higher number of intercellular interactions and physical limitations, restricting the drug permeability in 3D compared to 2D cultures 38 , 39 .…”

Is Spheroid a Relevant Model to Address Fibrogenesis in Keloid Research?

“…Cancer spheroids usually overgrow over time as a consequence of the pathological cell phenotype [ 37 , 38 , 39 ]. In our study, we did not observe such overgrowth, but we measured a decrease in the spheroid size during the culture period.…”

Continuous inline monitoring of glucose in an organ-on-chip using FreeStyle™ libre glucometers

“…Organ-on-chips (Lim et al, 2021;Liu et al, 2021;Subia et al, 2021), or microphysiological systems (MPS), are micro/milli-fluidic devices used to culture microtissues and are proving valuable as platforms that emulate relevant features of tissues and organs to facilitate biomedical research (Liu et al, 2022;Ngo et al, 2023; OPEN ACCESS EDITED BY Yuxiang Liu, Worcester Polytechnic Institute, United States Zhou et al, 2023). Organ-on-chips can be used to conduct fundamental studies (Zhang et al, 2017;Sabaté del Río et al, 2023;Wang et al, 2023), to model diseases (Weltin et al, 2014;Azizgolshani et al, 2021;Fuchs et al, 2021;Gallegos-Martínez et al, 2022), to assess the effects of physical or chemical stimuli on tissues (Dornhof et al, 2022), or to screen novel pharmaceutical compounds (Criscione et al, 2023;Sabaté del Río et al, 2023).…”