Lecture 4 - Molecular Targeting and Leukaemia

Question 1

What is the initiator mutation?

Answer 1

First mutation acquired by the cell

Question 2

At what stage do progressive or driver mutations occur and what do these cause?

Answer 2

After initiator mutations
Cause metastasises

Question 3

Is it easier to target overexpression or loss of function?

Answer 3

Overexpression

Question 4

How do you identify leukaemia?

Answer 4

Take blood sample, if milky then leukaemia

Question 5

What is leukaemia?

Answer 5

Unregulated proliferation of a clone of immature white blood cells, resulting in crowding normal cells out of the bone marrow
- loss of function of normal cells

Question 6

How is leukaemia classified?

Answer 6

Acute or chronic
Myeloid or Lymphoid

Question 7

Mutations in which cell type are the start of leukemia?

Answer 7

Haemopoietic stem cells (progenitor cells)

Question 8

What are the 3 clinical stages of leukaemia?

Answer 8

• Chronic for 4 years approx (not as serious)
• Accelerated phase
• Then acute stage after 4 years and more mutations, very few functional white blood cells

Question 9

What are symptoms of leukaemia?

Answer 9

Fatigue, Anaemia, swelling of spleen and liver

Question 10

In Chronic Myeloid Leukaemia what occurs in 95% of cases?

Answer 10

Reciprocal translocation between the chromosome 9 and 22

Question 11

Why is the reciprocal translocation an issue causing cancer?

Answer 11

As different chromosomes have different gene transcriptions, therefore if you move an oncogene to a chromosome with high transcription from low transcription this will contribute to cancer

Question 12

What is the example of reciprocal translocation which occurs in CML?

Answer 12

• Part of the gene for BCL-2 moved from a region of low to high transcription. This is an antiapoptotic pro-survival molecule.
• Placed next to a promotor region
• Formed a fusion protein BRC-ABL

Question 13

Why does the fusion protein BCR-ABL in CML cause cancer?

Answer 13

• swapping of the genes means the kinase domain is constantly active as protein cannot fold over
• this constantly signals for cell growth

Question 14

What class is c-ABL and its normal function?

Answer 14

Protein tyrosine kinase in the Src family
Normally has a cytoplasmic localisation and is protective

Question 15

Upon stress what occurs to c-Abl?

Answer 15

• translocates into the nucleus and engages with tumour suppressor proteins
• Complexes with P53 to induce cell cycle arrest and DNA repair
• complexes with P73 to induce apoptosis
• bound by pRb inducing cell cycle arrest

Question 16

How does the fusion BCR-Abl protein stop the normal function of c-Abl?

Answer 16

• Locks in the cytoplasm so cannot go to the nucleus to interact with tumour suppressors
• In cytoplasm activates signalling pathways such as Ras-MAPK which signals for cell growth and is pro survival

Question 17

Small molecule inhibitors for leukaemias with BCR-Abl

Answer 17

Imatinib
Dasatinib
Nilotinib

Question 18

What are the 4 names of Imatinib?

Answer 18

Imatinib, STI 571, Glivec, Gleevec

Question 19

What does imatinib work for?

Answer 19

• Inhibits proliferation of human CML-derived cell lines
• Blocks activity of Abl tyrosine kinase
• Also blocks PDGFbR and c-kit tyrosine kinases

Question 20

What were the responses like for CML patients receiving imatinib?

Answer 20

Nearly all had normal white cell count within 4 weeks
Over half had cytogenic response

Question 21

How does resistance build against Glivec in CML?

Answer 21

• Loss of sensitivity maybe due to BCR-ABL gene being amplified
• Mutations acquired over time
• clone population from the start with these resistant mutations

Question 22

What pathway can imatinib upregulate which causes resistance?

Answer 22

Imatinib upregulates Wnt pathway increasing
activation of pro-tumorigenic TCF7 TF which confers resistance

Question 23

Names of next generation 2nd and 3rd drugs made after imatinib for treatment of leukemia

Answer 23

2nd Dasatinib, Nilotinib

3rd Tozasertib

Question 24

Fusion protein associated with chronic myelomonocytic leukaemia (CMML)?

Answer 24

TEL-PDGFbeta receptor fusion

Question 25

What occurs to form TEL-PDGFbeta receptor fusion protein?

Answer 25

Translocation between chromosomes 5 and 12:
- amino terminal region of TEL (transcription factor)
- tyrosine kinase domain of the PDGF receptor
- Tyrosine kinase always active

Question 26

Which drug can be used to target TEL-PDGFbeta receptor fusion in chronic myelomonocytic leukaemia (CMML)?

Answer 26

Imatinib also

Question 27

What is the most important thing to do before treating cancers to know what to treat with?

Answer 27

Profile the patient

Question 28

What is GIST?

Answer 28

Gastro Intestinal Stromal Tumours

Question 29

What can be used to treat GIST?

Answer 29

Glivec (imatinib)

Question 30

Where is c-kit found?

Answer 30

• In gastrointestinal tract peristalsis coordinated by interstitial cells of cajal (ICCs)
• In the ICC membrane is c-kit

Question 31

What is c-kit?

Answer 31

A receptor tyrosine kinase

Question 32

What ligand binds c-kit and under normal conditions what does this cause?

Answer 32

Stem cell factor

• dimerization of c-kit receptor
• phosphorylation of downstream targets
• leading to proliferation and survival signals

Question 33

What happens in mutated c-kit?

Answer 33

• Membrane domain disrupted and constitutive activation of c-kit
• constant signalling for proliferation and cell survival
• formation of GISTs

Question 34

How does Glivec (imatinib) inhibit c-kit?

Answer 34

• GIST have consituitive c-kit activity
• can be inhibited by Glivec (tyrosine kinase inhibitor) which competitively binds to ATP binding pocket of c-Kit in both active and inactive receptors
• inhibiting downstream signalling inducing apoptosis in GIST cells or quiescence