Table 1.

Summary of 3D cancer cell culture techniques and animal models.

Technology	Technique	Description	Advantages	Disadvantages
Spheroid models	Multicellular tumor spheroids	Aggregation and compaction of suspended cancer cell lines26,27	Standardized cells; ideal for high-throughput screening (HTS); cell–cell interactions easily incorporated; and partial differentiation 24	Immortalized cell lines and culture adapted
	Tumorsphere (tumor organoids)	Clonal proliferation of cells suspended in stem-cell media 24	Enriched for cancer stem cells	Clonal cell population; only cancer stem cells
	Tumor-derived spheroids	Partial dissociation and reorganization of tumor tissue	Recreates tumor properties/ microarchitecture	Not standardized cell lines and exclusively tumor cells
	Organotypic spheroids	Mechanically diced and rounding of tumor tissue	Preserves tumor heterogeneity and microarchitecture	Not standardized cell lines
Scaffolds	Hydrogel-based scaffold	Cross-linked hydrophilic polymer network 28	Control over ECM proteins and growth factors and cell encapsulation	Poor mechanical properties
	Porous scaffolds	Various polymeric pore and fiber-forming techniques29–31	Diverse material selection and engineered microstructures	Inefficient cell seeding and variable mechanical properties
	Decellularized scaffolds	Decellularized ECM from tumor tissues 32	Mimics natural tissue properties and biocompatible	Inefficient cell seeding; immunogenic response; and technical preparation
Explant model	Tissue slice	Sectioning of surgically extracted tumor tissue	Preserves tumor heterogeneity and tissue architecture	Low throughput and challenging to maintain long term
In vivo tumor models	Cell line–derived xenograft (CDX)	Transplantation of cultured cancer cells into immunocompromised mice20,33,34	Easily established; synchronous growth; and low cost	Low genetic heterogeneity
	Patient-derived xenograft (PDX)	Surgically derived tumor transplantation of samples into immunocompromised mice 34	Retains human TME interactions at low passage numbers and serial transplantation avoids in vitro selection conditions	Human stroma loss in higher passages; high cost; time intensive; and engraftment variability20,34
	Environmentally induced model (EIM)	Induction of carcinogenesis via exposure to environmental stimuli	Relevant for tumorigenesis; captures genetic; and phenotypic heterogeneity	Difficult to determine tumor burden and long latency.20,35
	Genetically engineered mouse model (GEMM)	Induces cancer by cloning oncogenes or knocking out tumor suppressors in immunocompetent mice 34	Native TME and intact immune system	Variable gene expression and potential for random integration 34
	Humanized mouse (HM)	Engrafting human biological systems into immunocompromised mice	Incorporates aspects of the human immune system	Potential for graft rejection 33
	Other mammalian models (companion animals)	Naturally occurring tumors in animals that are genetically closer to humans than mice 36	Increased relevance compared to mouse models and more representative pharmacodynamics	Higher operational costs; longer lifespans; and specialized expertise
	Non-mammalian models	Tumor grafting on chorioallantoic membranes or zebrafish37,38	Low-cost alternatives to mammalian models and fewer ethical concerns	Labor intensive and limited to specific facets of cancer progression

ECM: extracellular matrix; TME: tumor microenvironment.