Liver cancer is the fifth most abundant solid cancer and in cancer-related death, which accounts for the third number worldwide. Hepatoma cell culture is a crucial system for in vitro research studies in biomedical and pharmaceutical science. However, conventional two-dimensional (2D) cell cultures could not represent the complex three-dimensional (3D) natural microenvironment of liver cancer. Especially, essential biological phenomena like adhesion, migration, gene expression, and proliferation can be more clearly reproduced by 3D culture. Currently, there is a lack of an adequate 3D culture model for studying tumor microenvironment. Therefore, engineering a suitable 3D matrix system for the Hepatoma microenvironment is a major challenge to be addressed. This will bridge between in vitro culture and in vivo tumors with the concept of applying 3 Rs Replacement, Reduction and Refinement to use animals for experiments more ethically. In the current study, a 3D scaffold system will be designed to produce hepatoma tumoroids. It is expected the postulated 3D scaffold will recapitulate the hepatoma microenvironment of hepatoma and hence it will represent the chemoresistance phenomenon to an anticancer drug that of like in vivo tumor. The possible factor involved and molecular mechanism at the genetic level will be evaluated by molecular level using PCR analysis. And the level of important genes involved in chemoresistance like interleukins (ILs), tumor necrosis factors (TNF), nuclear factor kappa-B (NF-kB), matrix metalloproteinases (MMPS), and notch signaling pathway genes will be evaluated. Level of apoptosis related genes e.g., BCL-2, BCL-xl and Bax, etc. will be also observed at the molecular level. The final strategy will be to be accomplished by inhibiting or knocking out one or more of the signaling molecule to overcome the chemoresistance face to already established drugs like 5- Fluorouracil, doxorubicin and new chemotherapeutic agents like curcumin and resveratrol.