Lecture 15

Question 1

what are the 2 important strategies for disease treatment?

Answer 1

1) prevent the disease occurring

2) reverse the disease process

Question 2

what is needed for the drug discovery process?

Answer 2

• lots of stages (very expensive if it fails at one of the later stages)
• Need to know the underlying biological processes

Question 3

what are the 4 differences between using mice instead of humans for testing drugs?

Answer 3

1) vary in development - gastrulation and organogenesis
3) Physiological differences - heart size etc
3) Physical differences
4) Genetic differences - mutations in the BLM gene is fatal in mice but gives disease in humans

Question 4

what is the issue in using human cell lines?

Answer 4

adult cells don’t grow very well outside of tissue

Question 5

what are the 4 major advantages of human pluripotent stem cells over conventional cell lines?

Answer 5

normal, primary cell line
High levels of telomerase - will grow forever
Differentiate to any cellular fate
maintain normal genotype if kept in correct conditions

Question 6

what is penetrance?

Answer 6

the proportion of individuals with a specific genotype who express a particular phenotype

Question 7

what are the 6 criteria that is needed to understand feasibility of modelling the disease in vitro?

Answer 7

1) inheritence - monogenic is easier than polygenic
2) penetrance -high penetrance is easier than low
3) age of onset - developmental is easier than late onset
4) mouse model
5) phenotype - cellular phenotype is easier than pattern formation
6) tissue - easy differentiation is better than very specialised

Question 8

IPS cells are pluripotent but unlike ES cells they carry the genotype of the parent cells what is an advantage of this?

Answer 8

• this means you have captured a particular genotype potentially forever as they can grow indefinitely
• could potentially make IPS cells from different genotypes (including people with diseases)

Question 9

what are the 3 strategies for generating disease models?

Answer 9

1) derive ES cells from embryos and genetically manipulate them using gene editing to produce the disease carrying PSCs
2) derive ES cells from diseased embryos
3) use somatic cells and transform them into iPSCs and edit genes into disease carrying PSCs

Question 10

what are 3 ways we can measure phenotypes?

Answer 10

• HTS (high throughput screening) - assay is read well by well basis by some sort of signal eg. Fluorescence - screen the wells - need to know what target is likely to be affected - very fast
• HCS (high content screening) - microscopes get images used to extract phenotypic information - difficult assay (hard to set up)
• Physiological assays - used to identify physiological features of the cells - low throughput

Question 11

once you have some understanding of potential targets for disease what are 2 ways to screen small molecule modulators?

Answer 11

1) conventional drug discovery pipelines

2) using PSCs

Question 12

what is a ortholog?

Answer 12

descended from the same ancestral sequence separated by a speciation event

Question 13

how can patient derived cells be useful in cancer studies and genetic diseases?

Answer 13

cancer = use tumour derived lines 
genetic = use immortalised blood or cells

Question 14

what are wild type IPS cells?

Answer 14

unaffected cells which are used as a control to compare diseased iPS cells too which produce affected cells

Question 15

when creating disease models are are the 3 stages and the 3 outcomes?

Answer 15

stage 1 = acquisition of genotypes - both diseased iPS and wildtype iPS
stage 2 = differentiate into cell type of interest - 2 plates of affected cells from diseased iPS and one plate of unaffected from wildtype
stage 3 = phenotypic readout - 1 initially diseased with no affected phenotype after treatment - 1 diseased phenotype and 1 with no affected phenotype from control wildtype

Question 16

what are 3 ways to identify affected/ successfully treated cells out of high density wells?

Answer 16

1) antibody staining
2) reporters
3) cell counts

Question 17

when the cells are placed in the wells of a molecules cell library what is a method used to try and correct the affected ones?

Answer 17

small molecules such as siRNA

Question 18

how can you test drugs for embryo toxicology?

Answer 18

testing drugs on cells mimicking embryogenesis to identify when vulnerable periods are and there susceptibility to disease -by measuring the production of cells that represent these different stages and the screen for any deviation from ‘normal’