MCB: Animal models in cancer Flashcards
Types of models (general)
- Subcellular
-Cellular
-Multicellular
-In vivo
Aspects cancer cell cultures
- Very useful model systems to study aspects of cancer biology:
-Examine effects of proliferation, cell cycle phase, survival, etc.
-Examine effects of cellular environment
-Examine mechanisms of action, identifying molecular networks - Useful models for anti-cancer therapy:
-Examine interactions between drugs and radiation and establish the mechanisms underlying such interactions
-Provide insights into effects of sequence, time and dose on effects
of single agent and combined modality treatments
-How they metabolize drugs - Amenable for experimental manipulation and analysis:
-Overexpression: transfection or transduction
-Knock-out or knock-down: RNAi or CRISPR/Cas9
-Omics analysis: RNAseq, ATACseq, ChIPseq, …
Advantages of cancer cell cultures
- Uniform, well defined cell populations
- Scalable and relatively cheap
- Controlled physio-chemical environment (pH, temp, CO2 level)
- Amenable for experimental manipulation
Disadvantages cancer cell cultures
- Must be maintained in sterile aseptic conditions
- Does not recapitulate complex tumor environment
- Rapid growth rate can lead to genomic alterations
- Tumor cell lines differ from tumor cells in vivo:
adapted to survive and grow in culture (plastic)
altered proliferation, clonal growth
altered gene expression, enzyme activity
altered shape, motility, metabolism, differentiation
altered response to external signals
Culture systems better for tumor in vivo
- Primary cell explants
- Three dimensional cultures (organoids)
- Perfused cultures
- Physiological growth surfaces
- Co-cultures containing multiple cell types
- ex vivo tissue and organ cultures that self organize (CRISPR-Cas9 approach)
None of these fully model tumors in vivo - they are all still
models, with inherent limitations
Human tumor xenograft mouse model
- Only the malignant cells are human
- Tumor cells have adapted for rapid growth in mice
- Murine tumor environment: stroma, vascular bed
- Pharmacokinetics, biodistribution & clearance are mouse
- Activation and metabolism of drug may reflect metabolism by
mouse cells
Two types of immunocompromised mice used for xenografts
- Nudes - reduced T-cells/thymus due to mutation in FOXN1 gene
- SCIDs - mutation in PRKDC gene, which plays a role in repairing
double-stranded DNA breaks. This has implications for B and Tcell receptor development, which is dependent upon such double-stranded breaks repair in order to rearrange V(D)J segments.
Patient-derived xenografts (PDXs)
+Gives genetic diversity and heterogeneity within tumors
-Requires surgical implantation
-Genetic and phenotypic drift with passage
Different types of GEMMs (4)
- Constitutive KO or overexpression: affects entire organism
- Conditional KO or overexpression: promoter/cell type-dependent
- Based on Cre/loxP recombination system
- Mainly tumor initiation
- Inducible KD or overpression: tetracyclin-dependent
- Cre-ERT2 Cre lines: tamoxifen-inducible
- Mainly tumor maintenance
Random / targeted gene technology
Random gene editing technology
* Pronucleus injection
* Viral transduction
Targeted gene editing technology
* Homologous recombination (HR) in mESCs
* Recombinase-mediated cassette exchange (RMCE) in mESCs
* CRISPR/Cas9
Setup of research plan
1) To: Identify novel candidate oncogenic drivers (tumor suppressors / oncogenes) for subtypes of cancer that are difficult to treat
2) Functionally evaluate = Manipulate expression of candidate oncogenic driver genes + analyze the effects on initation/progression of cancer (in vivo and in vitro)
3) Define mechanisms of action + can we perturb their oncogenic function?
4) Develop novel therapeutic anti-cancer strategies + evaluate them using existing cancer models (in vitro and in vivo)
5) Translate results to the clinic
For every step, need reliable models that mimc certain aspects of the disease = VERY important to pick correct model
