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Cancer Biology > Cancer Mouse Models > Flashcards

Cancer Mouse Models Flashcards

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AP® Biology flashcards Biology 101 flashcards
1
Q

What does transgenic mean?

A

Being an organism whose genome have been altered by the transfer of a gene or genes from another species or breed

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2
Q

What is the process of random integration?

A

Collect one cell embryos -> inject transgene into male pronucleus -> implant injected embryos into a pseudopregnant female -> live birth, test for transgene

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3
Q

What are the advantages of random integration

A

Easy
Origin of DNA not crucial
Can test promoter or gene function in vivo

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4
Q

What are the disadvantages of random integration?

A

Non-homologous recombination - can go anywhere in the DNA
Multiple integration - can end up with a random number of copies
>1 independent transgenic line required

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5
Q

Issue with non-homologous recombination

A

Since it can be implanted anywhere, if it goes into a sight which is not transcriptionally active then you will never see it
If it goes into a gene involve involved in embryo development then you wont see it since the mouse wont be born
Could interfere with housekeeping genes
Could go into a gene which interferes with its phenotype and has nothing to do with the transgene

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6
Q

What is the process of gene targeting?

A

Introduce targeting vector by electroporation into ES cells -> Rare cell carrying targeted gene -> Positive-negative selection -> Pure population of targeted ES cells -> injected into blastocytes -> implanted into surrogate mouse -> give birth to chimeric mice -> mating between chimeric mouse and normal mouse -> give birth to some gene targeted mice and some normal mice

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7
Q

What are the 3 types of conditional gene targeting?

A

Excision
Inversion
Translocation

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8
Q

What is excision conditional gene targeting

A

cis placement of loxP sites in same directional orientation

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9
Q

What is inversion conditional gene targeting?

A

cis placement of loxP sites in opposite directional orientation

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10
Q

What is translocation conditional gene targeting

A

trans placement of loxP sites

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11
Q

How does excision conditional gene targeting work?

A

when the loxP sites are in the same orientation, in ES cells or in vivo, express Cre recombinase which causes the two sites to recombine and take out the piece of DNA in one of the loxP sites

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12
Q

How does inversion conditional gene targeting work?

A

When the loxP sites are in the right position, instead of removing the DNA, the Cre recombinase will flip it

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13
Q

Why is conditional gene targeting good?

A

Since you can do it to a specific organ or the whole body

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14
Q

What are the 4 types of genome editing?

A

Meganucleases
ZFNs
Talens
CRISPR-Cas9

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15
Q

What is the process of CRISPR-Cas9 KO?

A

Genome DNA recognised through gDNA is cleaved by Cas9 -> DNA double-strand cleavage -> Repaired through non-homologous end joining - base deletion or insertion may occur -> mouse fertilised egg

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16
Q

What is the process of CRISPR-Cas9 KI?

A

Genome DNA recognised through gDNA is cleaved by Cas9 -> DNA double-strand cleavage -> Repaired through homologous recombination - mutation on DNA is introduced -> mouse fertilised egg

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17
Q

What are genetically engineered mouse models?

A

Genetic alterations which match human tumours

Can be conditional, i.e induced

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18
Q

How do you get lung specific Kras tumours via LoxP-mediated gene activation and inactivation?

A

Put a loxP stop signal in front of the mutant gene and insert this into the genome - due to the stop signal, it will not be expressed
Give adenoviral Cre as a nasal spray - expresses Cre recombinase - when it goes to the lung, the Cre is expressed and deletes the stop signal
Express the mutated gene and develop lung tumour

19
Q

What is the advantage of loxP mediated gene activation and inactivation?

A

You can put the same mutations in humans in mice and at the specific location

20
Q

What is the disadvantage of loxP mediated gene activation and inactivation?

A

Still a mouse and not a human

21
Q

How can you change the lung specific Kras tumours to ovarian cancer?

A

Instead of using nasal spray, inject it intraperitoneally (into the abdomen) -> adenoviral Cre is taken up by the mesenchymal blood supply of the abdomen -> passes into the circulation -> model ovarian cancer

22
Q

What is a syngeneic mouse?

A

mouse tumour in a mouse

23
Q

How do you make a syngeneic mouse?

A

Take mouse tumours (which are grown as immortalised cell lines in tissue culture) -> can inject the tumour different ways, e.g. subcutaneously (under the skin), into the brain or in the blood

24
Q

What is the disadvantage of syngeneic mice?

A

You can only inject cells from the same genetic background since it expresses an immune system and therefore would reject the tumour
Have to consider the differences between a mouse and human immune system

25
Q

What is the advantage of syngeneic mice?

A

It has an immune system and this is important for immunotherapies and that it shows the effect on the immune system

26
Q

What are mouse xenografts?

A

Human tumour in a mouse

27
Q

Why are the human tumours able to go into xenograft mice?

A

Since they are immunocompromised

28
Q

Name immunocompromised mice

A

Nude mice
Non-obese diabetic mice
SCID mice

29
Q

Why are tumours in xenografts typically grown just under the skin

A

There is no immune system to stop it
Easy to measure tumour growth - require advanced technology to measure tumour growth in tumours not under the skin or you have to kill lots of mice to measure them
Cheap

30
Q

Compare xenografts and genetically engineered mouse models

A 
Xenograft:
- tumour is in the wrong environment 
- lack of immune system 
GEMM:
- mirror in vivo mutations 
- high penetrance and multiplicity 
- immune system intact
31
Q

What are patient-derived xenografts good for?

A

If a patients tumour has relapsed, you can do genomics and see what mutations have arisen since the primary tumour
Can also add it into different mice and test different therapies to see which will be the most effective

32
Q

What are the issues with patient-derived xenografts?

A

Both methods are time consuming so the patient could be death
Very expensive
Need a mouse hospital next to a human hospital
You are still using immunocompromised mice

33
Q

How can you humanise patient derived xenografts in mice?

A

Use Nog or NSG mice

Add back various parts of the immune system into the mice until they develop an almost human immune system

34
Q

What are the issues with humanised patient derived xenografts?

A

Labour intensive
Expensive
Time consuming
Need lots of mice

35
Q

What are the 3 phases of drug metabolism?

A

Phase 1 - modification
Phase 2 - conjugation
Phase 3 - drug efflux

36
Q

What is the aim of drug metabolism?

A

To make the drug more soluble

37
Q

What are the functions of P450s?

A

Steroid biosynthesis
Bile acid biosynthesis
Drug/ xenobiotic metabolism
Fatty acid metabolism

38
Q

How are P450s activated?

A

By being fed electrons which P450 OxidoReductase catalyses from NADPH

39
Q

What is the issue when comparing rat/ mouse drug metabolism systems to humans?

A

Rats/ mice have a lot more P450 enzymes

40
Q

How do you humanise mouse drug metabolism systems?

A

Remove CYP3A genes and replace them with human ones

41
Q

What is a good drug model for humans?

A

Comparing the pharmakinetics of normal mice with humanised mice so they can you see how they differ

42
Q

How can you fully humanise a mouse’s liver?

A

Genetically create a mouse so that if you remove the dietary component the mouse’s liver starts to digest itself and then you add in primary hepatocytes and regrow the liver with human hepatocytes

43
Q

What is the issue with fully humanising a mouse’s liver?

A

It is not good for the mouse
Need primary human hepatocytes, which will need to be from one human and therefore will only have that humans metabolic profile
Every time you want a new mouse you have to repeat the process

Cancer Biology (13 decks)

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