Block E Lecture 4: Stem Cells in Drug Development.

Question 1

What 3 germ layers do embryonic stem cells differentiate into?

Answer 1

Endoderm
Ectoderm
Mesoderm

(Slide 3)

Question 2

What can pluripotent stem cells theorectically give rise to and how do they do this?

Answer 2

Every cell type in the animal body - done via fate determination and differentiation

(Slide 4)

Question 3

What 2 phases of the drug development pipeline can stem cells and transgenic mice be used?

Answer 3

The target discovery and pre-clinical stages

(Slide 5)

Question 4

What are 3 reasons pharmaceutal companies want validity in drug discovery?

Answer 4

Drug development is expensive

High failure rate is costly

Valid screening assays and pre-clinical models ensure drug candidates are tuned to humans and their diseases, and lack side-effects

(Slide 6)

Question 5

How are stem cells used to create valid in vitro screening processes?

Answer 5

1. Somatic cells are collected from the patient
2. The cells are programmed into induced pluripotent stem cells (IPSCs) which are capabale of differentiating into any cell type
3. These IPSCs are then differentiated into desired cell types or tissues, and can be used to create 3D cultures (organoid systems) which mimic complex tiussue architecture or or 2D cultures (target cell types).
4. These can be used to test drug effects in focused screening or to analyze large-scale high-content libraries for potential therapeutics.

(Slide 7)

Question 6

What do IPS cells in the screening process provide and what is the issue with them?

Answer 6

They provide better validity, but there are ethical concerns

(Slide 9)

Question 7

What 4 things do OSKM (yamanaka factors) do to induce pluripotency in differentiated somatic cells?

Answer 7

Upregulate embryonic stem cells genes

Upregulate cell proliferation

Loosen t he chromatin structure

Downregulate differentiation genes

(Slide 11)

Question 8

What model are embryonic stem cells the basis of?

Answer 8

Transgenic mouse models of disease

(Slide 18)

Question 9

What is one way that knockout mice can be created?

Answer 9

By modifying the target gene in embryonic stem cells using a process called homologous recombination, and then selection markers are used to identify the cells in which the targeting construct has integrated into the genome.

These correctly modified embryonic stem cells, are then injected into an early-stage embryo known as a blastocyte and then implanted into the uterus of a host mother.

After birth, the mice are then cross-bred to create offspring which have the genetic modification in all cells

(Slides 19, 20 and 21 )

Question 10

What are 2 reasons why mice are used as models of disease?

Answer 10

They are often much more resistant to disease than humans

Mouse models have been created which more closely mimic human disease - known as humanised mice

(Slide 22)

Question 11

Why did scientists create the GM-CSF knockout mice and what did this result in?

Answer 11

Mice use immune cells and cytokines (such as GM-SCF) to clear infection, so scientists created a GM-SCF knockout (GM-CSF KO) mice to develop a new animal model of choronic pulmonary M. abcessus infection

GM-CSF KO mice showed a week or so of acute infection which could be cleared by antibiotics
(Slide 25)