Cancer Model

Biomimicking of natural extracellular matrix (ECM) by electrospun scaffold has seen extensive research and application to use it in tissue engineering or regenerative grafts. Electrospun scaffold may also have the potential to investigate in vitro cancer cell behavior and to modify its behavior to mimic in vivo expression. This will provide an in vitro model for deeper understanding of cancer progression and drug screening for cancer treatments.

Preliminary studies using cancer cell lines have shown favorable adhesion and proliferation characteristics on electrospun scaffold. Szot et al (2011) tested the response of prostate cancer cell line (PC-3), murine renal cancer cell line (RENCA) and human breast cancer cell line (MDA-MD-231) on electrospun polycaprolactone/collagen fibers mat and compared with cultures on bacteria cellulose nanofiber mat (< 100 nm fiber diameter). Cell cultured on bacteria cellulose nanofiber mat showed poor viability and proliferation compared to electrospun fibers. Electrospun fibers with the smallest diameter of 424 nm exhibits the greatest viability and proliferation across the cancer cell lines compared to all other electrospun larger fiber diameters (1 µm, 1.6 µm and 2.2 µm).

Surface topographical effect of electrospun scaffold has been shown to influence the behavior of cancer cells. The effect of anti-cancer drugs seem to be reduced when cancer cells are cultured on electrospun scaffold compared to culture plates. Using gastric cancer cell line (MKN28), Kim et al (2009) showed lower inhibition of the cancer cells cultured on electrospun poly(3-hydroxybutyrate-co-3-hydroxyvalerate)/collagen nanofibers when anti-cancer drugs, namely, 5-FU, oxaliplatin and cisplatin are added compared to 2D cell culture plate cultures at 24 hours. However, paclitaxel and irinotecan are able to exhibit comparable cell inhibition to 2D culture. A study using breast cancer cells showed that influence of fiber alignment on the epithelial - mesenchymal transition (EMT). Saha et al (2012) tested the behavior of nonaggressive luminal cancer cells (MCF-7), aggressive basal cells (MDA-MD-231) and mammary tumor cell (H605) on electrospun aligned and randomly oriented polycaprolactone fibers with diameter of 1.8 µm and 2.0 µm respectively. Aggressive MDA-MD-231 cancer cells were found to align and elongated along the fiber axis but nonaggressive MCF-7 showed random orientation on aligned fibers. Mouse mammary tumor cells (H605) cells cultured on aligned fibers showed significant upregulation in the gene expression of Cytokeratin (Ck14), smooth muscle actin, TGFβ, Snail, fibroblast specific protein and Smad3 while less expression was observed in cells cultured on randomly oriented fibers. TGFβ and Snail are well known EMT inducers while Ck14 are known to activate aggressive basal-like breast cancer cells [Saha et al 2012]. Further, Nelson et al (2014) found that on aligned PCL nanofibers (diameter of 795 nm), MDA-MB-231 cells migrated significantly further in the presence of CXCL12 gradient compared to MCF-7 and MCF-10A (normal mammary epithelial) cells. Migration of all cell types on aligned nanofibers is greater than randomly aligned nanofibers. In the progression of breast cancer tumor, the extracellular matrix (ECM) transform from random fiber organization to highly aligned ECM which encourages migration and invasion. Aligned electrospun nanofibers scaffold may be used to mimic such fiber organization to replicate in vivo behavior such as rapid cell migration.

Breast cancer stem cells which are self-renewal and chemo-resistance are known to stay in a dormant state for many years as single cells and are resistant to therapies targeting proliferating cells. These cells have been described as having low rates of cell division, exhibit resistance to primary chemotherapy and radiation and expressing CD44+ and CD24-. Isolated MDA-MB-231 stem cells did not show significant difference in growth when cultured on fibrous scaffold and TCP. Comparison of a heterogeneous population of breast cancer cells on electrospun polycaprolactone fibers and tissue culture polystyrene (TCP) showed that aggressive breast cancer cells (MDA-MB-231) adopt a dormant phenotype with reduced growth on fibrous scaffold but the cells cultured on TCP maintains a high growth [Guiro et al 2015]. This gives electrospun scaffold the potential for enrichment of cancer stem cells for in vitro models.

In another study with breast cancer cells, Rabionet et al (2017) showed that isolated MDA-MB-231 triple negative cell line responded differently for electrospun PCL fibers with different diameters and tissue culture plate (TCP). Cell proliferation was fastest on TCP, followed by electrospun scaffold with fiber diameter of 700 nm and slowest on electrospun scaffold with 300 nm diameter fibers. However, the cell morphology was different between the two fibrous scaffolds. With 300 nm diameters fibers, the cells exhibited similar nucleus and cytoplasm architecture to 2D cultured cells. With 700 nm diameter fibers, high number of MDA-MD-231 cells showed lengthened morphology. Cells were also observed to have migrated into the depth of the scaffold made of 700 nm diameter fibers. Larger pore size of scaffold made from 700 nm diameters fiber may favor cell penetration while the smaller pore size from 300 nm diameters fiber may restrict cell movement into its depth. Therefore, the scaffold made from 700 nm fibers may provide a more 3D environment. Further studies on this scaffold showed significant aldehyde dehydrogenase-positive population increase compared to standard 2D culture. The cells on this scaffold also showed improved mammospheres forming ability, particularly after 6 days of culture which are evidence of stem cell characteristics.

Influence of material chemistry and fiber modulus on cancer cell behavior has been investigated using core-shell nanofibers. Rao et al (2013) carried out the study using primary tumor cells obtained from a patient with glioblastoma multiforme (GBM), the most common and aggressive brain tumor. To examine the effect of modulus on the cell behavior, core-shell fibers with a common shell material, polycaprolactone (PCL), and varying core materials were electrospun to form aligned fibers. It was found that the cancer cell migration was the fastest for fibers of intermediate modulus (~11 µm/h for ~ 8 MPa PCL nanofibers) and slower for fibers with higher (30 MPa) and lower (2 MPa) modulus. Material chemistry effect was examined by having a common PCL core and shell material of hyaluronic acid, collagen and matrigel. Their study showed that hyaluronic acid reduces cell attachment and migration while there was no change difference in cell migration between collagen, matrigel and PCL [Rao et al 2013].

Currently, there are very few, if any, inexpensive 3D-tumor model that replicates in vivo tumorigenesis. In the development of in vitro cancer tumors, both material and surface topography of the scaffold was found to influence the formation of tumoroids. Girard et al (2013) selected a mixture of PLGA and PLA-PEG block-copolymer for electrospinning to form a nanofibrous scaffold. PLGA and PLA were included for their better mechanical properties while PEG was thought to influence electrostatic binding properties of cells and encourage cell-cell interactions for the formation of tumoroids. Tests on electrospun PLGA only scaffolds do not form tumoroids which supports the importance of PLA-PEG block copolymer presence in the fiber. Topographical influence was supported by tumoroid formation on the electrospun nanofibers scaffold but not on film made from the same material. Having a coating of chitosan to impart a positive charge on the scaffold also resulted in the absence of tumoroids formation. Mimicking of in vivo tumorigenesis by LLC1 lung cancer cells cultured on the electrospun scaffold was verified by the induction of epithelial mesenchymal transition (EMT) which occurs during cancer metastasis. At day 3, the cells undergo EMT when cultured on PLGA/PLA-PEG nanofiber scaffold but not on PLGA scaffold or as a monolayer.

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▼ Credit and Acknowledgement