W5L1 thurs Cancer 2

Question 1

Angiogenesis

Answer 1

-Tumors recruit blood vessels
-Signals from the tumor can cause nearby blood vessels to sprout new branches which invade the tissue.
-Once the tumor is vascularised its growth becomes exponential and it is able to metastasis.

Question 2

How can tumor attract blood vessels

Answer 2

-As tumors grow, central cells become starved of oxygen. This condition is sensed by a protein called Hypoxia Inducible Factor 1a (HIF1a).
-At normal oxygen levels, HIF1a is degraded (due to another protein pVHL) but in hypoxic conditions it builds up and is able to activate transcription of Vascular Endothelial Growth Factor (VEGF).
-VEGF diffuses out and activates the VEGF Receptor on nearby endothelial cells triggering the sprouting of new vessels

Question 3

Initial stages of metastasis

Answer 3

-The metastasis of epithelial tumors begins with an epithelial mesenchymal transition (EMT).
-Cells lose their epithelial characteristics and become migratory. This allows them to invade the surrounding tissue and enter the vasculature.
-A process called intravasation.

Question 4

Gene responsible for EMT: finding it

Answer 4

-4 cell line isolated from mammary tumor
-inject these cell back into a naive mice
- All cell line produce tumors but one did not metastasise. Further analysis by microarray
- Twist was found as the odd one out from the arry. Reducing twist level prevent metastasis

Question 5

Role of TWIST

Answer 5

-Twist represses E-Casherin
-When cells undergo an EMT, they stop expressing epithelial genes like E-Cadherin and b-catenin and start expressing mesenchymal genes needed for migration like the intermediate filament protein, Vimentin.

Question 6

Twist as a prognostic marker for cancer

Answer 6

-Twist is upregulated in many cancers and predicts poorer survival
e.g. Types of cancers include breast, ovarian, cervical, prostate, bladder, hepatocellular, gastric, colorectal, and Chronic Myeloid Leukemia.

Question 7

Origin of TWIST discovery

Answer 7

During early Drosophila development, Twist is expressed in cells on the ventral side of the embryo.
-These cells are initially epithelial but Twist expression causes them to break free of their neighbours and become migratory. In twist mutants this process fails.
-TWIST is a conserved driver of EMT

Question 8

E-Catherine and cancer

Answer 8

-E-Cadherin is a tumor suppressor
-Loss of E-Cadherin is a common feature of cancer cells
-Hereditary loss-of-function mutations in CDH1 (i.e. E-Cadherin) greatly increase risk of diffuse gastric cancer.

Question 9

Circulating Tumor Cells

Answer 9

-The tumor cells that enter the blood stream are then called Circulating Tumor Cells or CTCs
-The formation of the secondary tumors (also called metastases) around the body involves CTCs leaving the blood vessel (a process called extravasation) to seed the new tumor.
-The growth of the secondary tumors is thought to involve a mesenchymal to epithelial transition (MET).
-Circulating tumor cells (CTCs) can also circulate as clusters, also called Circulating Tumor Microemboli (CTMs)

Question 9

What is more dangerous? CTCs or CTMs Experiment

Answer 9

1. inject mix of red and green tumor cells.
2. look in the blood for CTCs (i.e. single colour) and CTMs (single colouror multi-coloured).
3. Look at metastases. Are they single or multi-coloured.
   > CTMs are much better at metastasisng but higher discovery of CTC in the bloods

Question 10

The cell search system

Answer 10

-The CellSearch system is designed to detect Circulating Tumor Cells.
-Blood samples from a patient are submitted to the machine, it immunomagnetically sorts the cells, immunostains them, and then presents them for inspection

Question 11

Principle of Cellsearch system

Answer 11

-anti-EpCam recognises the Epithelial Cell Adhesion Molecule which is strongly expressed in human adenocarcinoma cells
-anti-CD45-APC identifies leukocytes, (regarded as a contaminant)
-anti-CK-PE identifies intracellular cytokeratins 8, 18, and/or 19

Question 12

Counting CTCs has prognostic value

Answer 12

• 430 metastatic colorectal cancer patients were analysed with the CellSearch® System before and during treatment.
• patients were classified into favourable (<3 CTC/ 7.5 mL) and unfavourable (≥3 CTC/7.5 mL) groups.
• favorable patients had longer survival time (18.5 v 9.4 months).
• Conversion of unfavorable to favorable at 3-5 weeks of treatment was associated with longer survival time (compared to patients with unfavorable CTCs at both time points) (11.0 v 3.7 months).

Question 13

Cancer stem cell hypothesis

Answer 13

-like normal tissue, cancer is comprised of different cell type
-experiments show that typically only 1% of tumor cell can initiate a new tumor
-Think that there is a cancer stem cell that can under goes self-renewal and give rise to the many cell types

Question 14

cancer heterogeneity and clonal evolution

Answer 14

-Cancer cells can be thought of as being in competition. If a cell acquires a mutation that makes it proliferate more quickly, for example, its progeny will become more prevalent in the tumor. —Thus a tumor is actually genetically heterogenous (i.e. made up of cells with different mutations) and this genetic makeup evolves over time.

Question 15

Evolutionary tree of mutations in a cancer

Answer 15

-By sequencing the genomes of 100 cells from different regions of a tumor researchers were able to piece together a family tree of the cell lineages.
-The tumor cells had branched into three main lineages. One of these had acquired mutations that resulted in increased levels of K-Ras which would have taken over the tumor with time

Question 16

Cancer evolution and genetic variation

Answer 16

• The more genetic variation there is in a tumor, the more opportunity there is for evolution. Therefore, mutations that lead to more variation will help a cancer to evolve.
• Hence, cancers often acquire mutations in genes that are involved in the DNA damage response:
  – i.e. p53 is a good example, but there are many other genes involved.
• Loss of such proteins leads to increased mutation rate and chromosomal instability (CIN)

Question 17

Chromosomal instability

Answer 17

-Can lead to aneuploidy
chromosomal instability (CIN) = gain or loss of whole chromosomes or fractions of chromosomes.
aneuploidy = an abnormal number of chromosomes

Question 18

How CIN lead to aneuploidy

Answer 18

-Normally microtubules from each spindle pole attach to one of the two chromatids via the kinetochores. Mitotic checkpoints delay mitosis until all chromosomes are correctly attached. This ensures each cell gets a full copy of the chromosomes.
-Otherwise, anaphase could occur when some kinetochores weren’t yet attached, or were attached to both spindle poles. Sometimes there might be multiple spindle poles. The result is chaos (aneuploidy, breakages, etc)

Question 19

Colorectal carcinoma and APC

Answer 19

People that have inherited one faulty copy of the Adenomatous Polyposis Coli (APC) gene are predisposed to colorectal cancer (through LOH for APC).
APC is part of the complex that targets beta-Catenin for destruction so loss of APC overactivates the Wnt pathway and causes these tumor growths (adenomas)

Question 20

What can cause cancer relapses

Answer 20

-Just as one can have genetic bottlenecks in the evolution of species, events that kill off the majority of cells within a cancer may leave some survivers. These survivers will then come to dominate the new tumors.
-If these bottlenecks are due to chemotherapy, it can mean that one is selecting for cell types that are resistant, and will therefore cause relapse and require different drugs