A silk fibroin based hepatocarcinoma model and the assessment of the drug response in hyaluronan-binding protein 1 overexpressed HepG2 cells

Microenvironment around tumor cells plays an important role in its malignancy or invasiveness. Hyaluronan (HA), a major component of extracellular matrix is found to be elevated in most of cancerous niche/microenvironment and performs regulatory role in the progression of tumors and metastasis. Overexpression of the hyaladherin, hyaluronan-binding protein 1 (HABP1) in the hepatocarcinoma cells (HepG2) termed as HepR21 leads to enhanced cell proliferation with increased HA 'pool' associated with HA 'cables' indicating elevated tumorous potential under 2D culture conditions. For in vitro experimentation, scaffold based three dimensional niche modeling may have greater acceptance than conventional 2D culture condition. Thus, we have examined the influence of intrinsic properties of non-mulberry tropical tasar silk fibroin on the HepR21 cells in order to develop a 3D hepatocarcinoma construction to act as model. The scaffold of tasar silk fibroin of Antheraea mylitta when efficiently loaded with transformed hepatocarcinoma cells, HepR21; exhibits enhanced adhesiveness, viability, metabolic activity, proliferation and enlarged cellular morphology in 3D compared to its parent cell line HepG2, supporting the earlier observation made in 2D system. In addition, formation of multicellular aggregates, the indicator of tumor progression is also revealed in silk based 3D culture conditions. Further, the use of 4-MU (a hyaluronan synthase inhibitor) on HepR21 cells reduces the HA level and downregulates the expression of growth promoting factors like pAKT and PKC; while upregulating the expression of the tumor suppressor p53. Thus, 4-MU efficiently reduces the tumor potency associated with increased HA pool as well as HA cables and the effect of 4-MU doubling up as an anticancer agent in 2D and 3D are also comparable. The in vitro 3D multicellular model demonstrates the insight of hepatocarcinoma progression and offers the predictability of cellular response to transfection efficacy, drug treatment and therapeutic intervention.