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Drug Development > In Vivo, In Vitro And Ex Vivo Studies In Preclinical Drug Development > Flashcards

In Vivo, In Vitro And Ex Vivo Studies In Preclinical Drug Development Flashcards

1
Q

In silico methods define:

A

Experimentation performed by computers
Used alongside in vitro data to create a model and test it

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2
Q

In silico methods are used in?

A

Discovery of novel molecules with target affinity
Clarification of ADME and toxicity properties
Physiochemical characterisation

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3
Q

Examples of in silico methods

A

Machine learning
Data mining
Virtual ligand screening and affinity profiling
Pharacophores
Homolgy modelling

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4
Q

In vivo models define:

A

Experiments performed in or on whole living organisms (humans, animals, plants)
DOESN’T PROVIDE ADME results

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5
Q

In vivo model examples

A

Disease models
Behavioural studies
Safety (toxicology)
Efficiency

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6
Q

In vivo models chosen by

A

Physiological and biochemical similarities to humans
Mechanisms of drug ADME
Ethics, susceptibility to disease
Ease of handling, availability
Housing requirements, cost

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7
Q

Immunocompetence define

A

Body’s ability to produce a normal immune response after exposure to antigens

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8
Q

BALB/c

A

Albino, lab - bred strain of the house mouse

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9
Q

C57BL/6

A

Black, inbred strain, less docile than the BALB/c

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10
Q

Immunodeficiency

A

Immunocompromised, immunosuppressed
Opposite of immunocompetence: compromised immune system

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11
Q

Nude mice characteristics

A

Lack body hair
Deteriorated or no thymus → low T cells
Used for xenografts

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12
Q

NSG (NOD-scid gamma null) mice characteristic

A

Lack natural killer, T and B cells; detective cytokine signalling =) Severely immunodeficient
Used for engraftment of primary human cells

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13
Q

Humanised mice define’.

A

Immunodeficient mice transplanted with human cells/tissues
(Get IV injection)

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14
Q

Humanised mice get IV injection of:

A

Peripheral blood mononuclear cells (PBMCs) → memory T cells, but no B and myeloid cells
CD34+ human hematopoietic stem cels → enhances human Cell reconstitution

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15
Q

Humanised mice are used for?

A

Cancer, infections, autoimmune disease studies

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16
Q

Immunodeficient nude mice used for?

A

Xenograft

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17
Q

Immunodeficient NSG mice used for?

A

Engraftment of primary human cells

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18
Q

Heterotopic transplantation site

A

In different areas eg under the skin (subcutaneous)

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19
Q

Orthotopic transplantation site

A

In organ type in which tumour originated

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20
Q

Metastasis transplantation site

A

Via tail vein into circulation

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21
Q

Immunocomptence mice examples:

A

BALB/c
C57BL/6

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22
Q

Immunodeficient mice example:

A

Nude, NSG

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23
Q

Transplantation type examples

A

Heterotopic
Orthotopic
Metastasis

24
Q

Xenograft transplanted in?

A

Immunodeficient mice

25
Q

Xenograft animal model used for?

A

Cancer studies

26
Q

What type of xenografts can be done?

A

Patient derived xenografts → cells/tissues from a patients tumours
Human tumour cell lines

27
Q

Types of animal models

A

Xenograft
Genetically engineered mouse model (GEMM)
Syngeneic / allograft
In vitro methods
Ex vivo methods

28
Q

Examples of GEMM and explain:

A

Transgenic: foreign gene integrated into animal’s genome
Knockout / knock-in: targeted genes made non-functional or altered

29
Q

Syngeneic/allograft animal models transplanted into?

A

Immunocompetent mice

30
Q

Syngeneic/allograft animal model explain/ used for?

A

Tumour cells/tissues derived from same genetic background as a given mouse strain
Syngeneic mice retain intact immune system → used for immunotherapy studies

31
Q

In vitro methods tests on?

A

Isolated tissues, organs or cells performed outside of a living organism
Eg cells studied in culture media

32
Q

Ex vivo methods are?

A

Experimentation in or on tissues from an organism in an external environment with minimal alteration of natural conditions

33
Q

Ex vivo methods used in pharmacology for?

A

Testing compounds on tissues using eg. Organ baths
After treatment to investigate issues not identified in vivo

34
Q

Cell proliferation/viability assay measures:

A

Measures metabolic activity=> cell viability, proliferation and cytotoxity after exposure to stimulus or toxic agents
Changes in coloured reagent quantified by measuring absorbance (spectrophotometer)

35
Q

Cell proliferation/viability assay used in?

A

Drug sensitivity
Cytotoxicity
Response to growth factors
Cell activation

36
Q

Cell proliferation/viability assay results used to calculate.

A

IC50- half maximal inhibitory conc.
EC50- half maximal effective conc.

37
Q

Flow cytometry used to?

A

Measure physical and chemical characteristics of cells or particles

38
Q

Flow cytometry used in? Examples

A

Basic research, clinical practice/trials:

Cell counting, characteristics and function
Biomarker detection
Protein engineering detection
Diagnosis of health disorders e.g. Blood cancer

39
Q

Flow cytometry procedure:

A

Cells/particles in fluid injected into Flow cytometer analyser
One Cell at a time through laser beam → Light scatter characteristic to cells and their components
Cells are labelled with fluorescent markers → light absorbed and emitted in different wavelengths

40
Q

Fluorescent markers are?

A

Fluorophores attached to antibodies that recognise targets on or in cells

41
Q

Western blot defects?

A

Specific proteins in tissue homogenates or Cell extracts by separating a specific protein from a complex

42
Q

How is a Western blot carried out?

A

Separation by size-gel electrophoresis
Electrophoretic transfer or protein onto a membrane
Marking of target protein with a primary and secondary Ab for visualisation

43
Q

Enzyme-linked immunosorbent assay, ELISA detects?

A

Ligands (proteins) in liquid samples using Ab against proteins of interest

44
Q

ELISA procedure:

A

Wells are pre-coated with capture Ab and sample added
Sample antigens attach to a surface
Matching Ab linked to enzymes bind antigens
Enzyme’s substrate is added
Enzyme binding to substrate produces detectable signal (colour change) =) spectrophotometry

45
Q

Microscopic study of disease tissue known as

A

Histology
Used of ex vivo

46
Q

Examples of staining to show general tissue structure

A

Haematoxylin and eosin

47
Q

Haematoxylin stains

A

Cell nuclei blue

48
Q

eosin stains

A

Cytoplasm pink

49
Q

Immunohistochemistry HIC identifies?

A

Proteins in tissue section using antibody-antigen binding in biological tissues
(Visualises antibody-antigen interaction)

50
Q

How does IHC Visualises antibody-antigen interaction?

A

Chromogenic and immunofluorescence

51
Q

Chromogenic IHC

A

Ab conjugated to enzyme → colour producing reaction

52
Q

immunofluorescence IHC

A

Ab tagged to fluorophore

53
Q

IHC used in?

A
  • Cancer diagnosis, basic research
54
Q

Immunocytochemistry ICC visualises?

A

Localisation of specific proteins/antigens in cells using Ab binding and fluorescence microscopy

55
Q

ICC VS IHC

A

ICC uses samples of intact cells, whereas IHC uses sections of biological tissue

Drug Development (11 decks)

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