2D and 3D thermally bioprinted human MCF-7 breast cancer cells: A promising model for drug discovery.

Abstract

2605 Background: Breast cancer (BC) is the second leading cause of cancer death following lung cancer. Bioprinting, the use of computer aided process to print biological living and non-living material to create patterns in 2D or 3D structures, is a novel technique that has been proposed to be used to develop tissue engineered solutions for a wide array of clinical applications, e.g., skin grafting. We investigate here if bioprinted breast cancer cells show some of the hallmarks of cancer tissues, and thus may represent good in vitro models for drug discovery. Methods: For this study, MCF-7 BC cells were cultured, stained, counted and turned into a bioink solution by suspending in phosphate buffered saline solution. The cells were bioprinted over a 96-well plate and pre-incubated for 18 hours in DMEM and RPMI media with 10% Fetal Bovine Serum and Charcoal Stripped Serum, respectively. After 18 hours of incubation the media was supplemented with Tamoxifen at 5µM, 10µM, 50µM, 90µM and 110µM concentrations. Cytotoxicity was measured 24 hours post-treatment using a differential nuclear staining assay and an INCell 2000 bioimager system. Results: Bioprinted cells exposed to high concentrations of Tamoxifen (90 µM and 110µM) exhibited a viability of 8.2% and 10.8%, respectively. Whereas viability of manually seeded cells at those concentrations was 0.11% and 0.05%. Viability of negative and positive controls was at 7.6% and 97.0% for the bioprinted samples and for the normally seeded cells was 4.9% and 98.8% respectively. Conclusions: In our study, we have established a novel 2D/3D breast tumor model applying bioprinting technology for drug discovery. The higher cell viability of MCF-7’s at high concentrations of Tamoxifen could be attributed to the hormesis effect and activation of chaperone proteins, e.g., HSP70 and HSP90, possibly caused by bioprinting. We hypothesize that bioprinted MCF-7 cells also show increased levels of chaperone proteins, which may in a way mimic their in vivo behavior. In this novel in vitro 2D/3D model, the bioprinted cells show a more biological relevant behavior than normally cultured cells. Insights into the cell behavior after bioprinting may elucidate how to build improved in vitro models for BC research.

Thermal Bioprinting Causes Ample Alterations of Expression of LUCAT1, IL6, CCL26, and NRN1L Genes and Massive Phosphorylation of Critical Oncogenic Drug Resistance Pathways in Breast Cancer Cells

“…On the other hand, downregulated genes in BP cells with ≥10 protein interactions are: TP53, FOS, JUN, EGR1, HIST2H2AC, and FOSB (see Table 2). TP53, aka tumor suppressor gene, is further repressed in these cells; this explains the added resolute characteristics hence death evasion (Campbell et al, 2019). A Complete list of gene analysis is available in Supplemental Information II.…”

“…Bioprinting may be causing thermal and mechanical stress to the cells resulting in a high number of stressed and weak cells that when subjected to the Annexin A5-FITC assay, became apoptotic, instead of fully convalescing. This is important when considering drug tests, BP cells shall be allowed recovery time with at least one media change prior to testing any drugs (Campbell et al, 2019).…”

3D bioprinting for reconstituting the cancer microenvironment

“… Cancer model type Cell types used Bioink or substrate used Bioprinting modalities used Ref. Glioblastoma-on-a-chip Glioma cell line U118 and endothelial cells Collagen or dECM hydrogel EBB 28 Glioma stem cell (shell); glioma cell line (core) Alginate Coaxial EBB 29 Glioma stem cells Gelatin, alginate, and fibrinogen EBB 30 Hepatoma HepG2 and glioma cell U251 Alginate DBB (Inkjet) 31 iPSC-derived human neural progenitor cells and U118 human glioma cells Scaffold-free 3D culture EBB 32 Human glioma stem cell line, U118 Sodium alginate and gelatin EBB 33 Glioblastoma-associated macrophages (GAMs) and glioblastoma cells GelMA EBB 34 Human primary umbilical cord-derived mesenchymal stromal cells (UC-MSC, referred to as MSCs), HUVEC, and human bone marrow-derived epithelial-neuroblastoma immortalized cells (SH-SY5Y) Agarose and type-I collagen DBB 100 Breast tumor model Immortalized non-tumorigenic human breast epithelial cell lines MCF-12A and MCF10A Rat tail collagen I EBB 36 MCF-7 BC cells PBS solution DBB 101 Immortalized non-tumorigenic human breast epithelial cell line, MCF-12A, and the breast carcinoma cell lines MCF-7 and MDA-MB-468 Rat tail collagen EBB 37 MCF-7 cell Gelatin-PEG DBB 38 BT474 breast cancer cells, human perinatal foreskin fibroblasts (BJ), and human adult dermal fibroblasts (HDF) Poly(ethylene glycol) diacrylate (PEGDA) LBB (optical project...…”

Cancer Cell Direct Bioprinting: A Focused Review

“…Van Pel et al combined the extrusion printing with spheroids from glioblastoma cell lines, to model the glioma invasion process that occurs in vivo [ 104 ]. On the other hand, inkjet printers can also bioprint cell suspensions to develop a breast cancer model for drug discovery and testing [ 105 ].…”