Abstract
PurposeMultiple myeloma (MM) remains an incurable hematologic malignancy which ultimately develops drug resistance and evades treatment. Despite substantial therapeutic advances over the past years, the clinical failure rate of preclinically promising anti-MM drugs remains substantial. More realistic in vitro models are thus required to better predict clinical efficacy of a preclinically active compound.MethodsHere, we report on the establishment of a conical agarose 3D co-culture platform for the preclinical propagation of primary MM cells ex vivo. Cell growth was compared to yet established 2D and liquid overlay systems. MM cell lines (MMCL: RPMI-8226, U266, OPM-2) and primary patient specimens were tested. Drug sensitivity was examined by exploring the cytotoxic effect of bortezomib and the deubiquitinase inhibitor auranofin under various conditions.ResultsIn contrast to 2D and liquid overlay, cell proliferation in the 3D array followed a sigmoidal curve characterized by an initial growth delay but more durable proliferation of MMCL over 12 days of culture. Primary MM specimens did not expand in ex vivo monoculture, but required co-culture support by a human stromal cell line (HS-5, MSP-1). HS-5 induced a > fivefold increase in cluster volume and maintained long-term viability of primary MM cells for up to 21 days. Bortezomib and auranofin induced less cytotoxicity under 3D vs. 2D condition and in co- vs. monoculture, respectively.ConclusionsThis study introduces a novel model that is capable of long-term propagation and drug testing of primary MM specimens ex vivo overcoming some of the pitfalls of currently available in vitro models.
Highlights
Multiple myeloma (MM) is a plasma cell disorder with an expected number of 34,920 new cases in the United States in 2021 (American Cancer Society 2021)
MM cells are cultured in conical microwells formed of a non-adherent agarose matrix with bone marrow (BM)-derived stromal cells plated on the base of each plate
To compare and optimize ex vivo propagation of MM cells, RPMI-8226 cells were cultured in three different in vitro platforms. These platforms included a conventional 2D monolayer model, a liquid overlay approach, as well as the culture of cells in a novel 3D microwell device that allows for self-assembly of cell clusters (Figs. 1A–D, 2A). Aggregate formation in the latter model was supported by a centrifugation step which forced seeding of MM cell lines (MMCL)
Summary
Multiple myeloma (MM) is a plasma cell disorder with an expected number of 34,920 new cases in the United States in 2021 (American Cancer Society 2021). Overall response rates to induction therapy have substantially improved over the past decades and are nowadays consistently > 90%, with a pooled 5 year overall survival (OS) of 54% across all age and risk groups (American Cancer Society 2019). Despite such substantial progress, MM remains an incurable disease that develops resistance and evades treatment. MM cells are cultured in conical microwells formed of a non-adherent agarose matrix with bone marrow (BM)-derived stromal cells plated on the base of each plate This distant co-culture model allows for the diffusion of soluble cytokines between BM stromal and MM cells. In addition to reducing cost and time for more realistic preclinical drug screening, the here established 3D model holds the potential to perform drug testing in primary patient samples and to predict individual treatment responses ex vivo in real-time
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