Tissue Invasion and Metastasis

Question 1

Describe what is meant by metastasis

Answer 1

Cancer cells breaking away from their primary site and invading surrounding tissues and spreading, forming a metastases at a distant site

Question 2

Explain the seed and soil theory

Answer 2

The idea that cancer metastases are the result of interaction between seeds (tumour cells) and congenial soil (a receptive metastatic microenvironment). This implies that certain organs are more likely to have metastases eg. lung, liver, bone. This idea was preceded by the fact that arrested cancer cells form emboli and travel via vasculature.

Question 3

Describe the difference between monoclonal and polyclonal metastasis

Answer 3

A tumour will have different clones in it due to genetic mutations. Genomic instability could result in metastasis. Metastasis can either come from one type of clone, or multiple, or one type that undergoes further mutations in the metastases, becoming phenotypically heterogenously monoclonal

Question 4

Describe the concept of circulating cancer cells

Answer 4

They are cancer cells disseminating through blood/lymphatic vessels. First observed in the peripheral blood of a patient with metastatic disease. The blood showed cells with similar characteristics to those in the metastases.

Question 5

Why are CTCs so hard to detect and how can they be detected?

Answer 5

They are rare compared to other circulating cells, and there is no universal recognition marker. Other circulating cells include: cell-free tumour DNA, ctRNA, proteins, and exosomes coming from the primary tumour.

Question 6

How can liquid biopses be used to detect cancer?

Answer 6

Liquid biopsies are clinical tests that examine the fluid from patients with cancer such as urine, saliva, pleural effusions, CSF in order to detect CTC or other tumour derived material

Question 7

Why is CTC screening not easy?

Answer 7

Because ctDNA is highly fragmented and underrepresented compared to tumour DNA. Also, the abundance of CTCs is super low in the blood and only a limited number of CTCs can be isolated per sample

Question 8

What are the two methods of enriching samples for CTC detection?

Answer 8

Negative enrichment - Removing other blood cells based on shape size, or other biophysical properties
Positive enrichment - Selecting cells expressing specific surface markers eg CAM and not CD45

Question 9

How do proliferating tumour cells gain movement ability?

Answer 9

They overcome their mechanisms of attachment to other cells/ECM, and breakdown tissue architecture. Enzymes are produced that breakdown adhesion molecules and basement membrane, this is the process of EMT.

Question 10

Describe the process of metastasis

Answer 10

The basement membrane is breached and tumour cells intravasate into blood/lymphatic vessels until they reach their new destination, where they extravasate into a new tissue and expand a metastatic colony.

Question 11

Describe what the basement membrane is

Answer 11

The basement membrane is produced by epi and endothelial cells and stroma, to separate the epi/endothelia from stroma and interstitial matrix. It is a specialised type of ECM that is more compact, less porous, and has a distinctive composition. ECM is divided into BM and interstitial matrix. Basal lamina is a layer of BM

Question 12

Outline the role of proteases in the breakdown of the ECM

Answer 12

Tumours and metastatic cancer cells degrade the ECM with proteases that hydrolyse the peptide bonds present in the ECM.

Question 13

Explain the role of cathepsins in the breakdown of ECM

Answer 13

They turnover and degrade the ECM, activating, processing and degrading various GFs, cyto/chemokines and influencing CAMs

Question 14

Explain the role of urokinase in the breakdown of ECM

Answer 14

The urokinase receptor and urokinase-type plasminogen activator receptor regulates urokinase proteolytic activity and ECM component degredation, regulation of cell adhesion, migration, prolif., survival by interaction with other transmembrane receptors eg integrins. uPAR binds vitronectin and triggers the induction of EMT

Question 15

Describe the role of MMPs in the breakdown of ECM

Answer 15

Degredation of collagen and other ECM proteins and regulation of cell behaviour. There are 6 types, collagenases, stromelysins, matrilysins, membrane-type MMPs, non-classified MMPs

Question 16

Describe the role of cell junctions in cell attachment

Answer 16

Adherens junctions are the master junction, involving cadherins attaching one cell to the actin microfilaments of the other cell, controlling the function of all the other junctions.

Question 17

Describe the role of integrins in cell attachment

Answer 17

Integrins interact with ECM ligands, sensing outside-in signals and becoming active. This allows them to respond to ECM alterations, triggering intracellular responses. On the intracellular side, integrins interact with other ligands upon activation, recruiting focal adhesion kinases which are autophosphorylated and complex with Src, activating both kinases

Question 18

Describe the process of EMT

Answer 18

During EMT, cancerous epithelial cells lose their epithelial characteristics and acquire mesenchymal characteristics, becoming an invasive phenotype. Key changes involve a reduction in E-cadherins and a dysmorphic shape. These changes facilitate motility and invasiveness. At the secondary site, the cells undergo MET in order to form a new carcinoma

Question 19

How is F-actin involved in cell migration

Answer 19

Filamentous actin is involved in the production of forward protrusions that allow cells to undergo mesenchymal migration - cells migrating on a basement membrane

Question 20

Describe and explain the difference between the two types of migration

Answer 20

Ameboid migration - A single round or ellipsoid cell without strong adherence to ECM travels in this way. Involves rapid production and retraction of actin-rich front protrusions. Non actinous protrusions (blebs) also present, produced through hydrostatic pressure and cytoplasmic flow
Epithelial migration - Bulk migration of cells connected via CAMs, the cells can move as clusters, sheets, strands, fluid like masses.

Question 21

Which cancers travel via haematogenous and lymphatic routes?

Answer 21

Haematogenous spread is common for sarcomas and some carcinomas
Lymphatic spread is common for carcinomas
Lymphatic spread often leads to haematogenous spread

Question 22

Describe and explain what is meant by transcoelomic spread

Answer 22

Cancer cells spreading through the surfaces of body cavities. Ovarian cancer can spread into the peritoneal cavity and in lung cancer, cells can spread into the pleural cavity. The peritoneal cavity is between visceral peritoneum covering interal organs and peritoneum surrounding abdominal wall

Question 23

Describe and explain what is meant by perineural spread

Answer 23

Cancer cells spreading into the layers of the nerves. This spread is found in highly innervated cancers such as head, neck, prostate, colorectal cancers

Question 24

Describe and explain what is meant by leptomeningeal spread

Answer 24

Cancer cells travelling via the CSF, characteristic of cancers occuring in the brain such as non-Hodgkins lymphoma. They invade the meninges or the dura mater and epidural space via the spinal cord

Question 25

How does the TNM staging system work in describing metastasis

Answer 25

It works by assuming a sequential metastatic journey, going from tumour to lymphnodes to metastases. Lymphnodes can be removed surgically

Question 26

Explain why lymphatic sprad usually leads to haematogenous spread

Answer 26

Because lymphatic vessels drain into blood vessels at thoracic ducts

Question 27

What is the role of the circulation in determining the final location of the metastases

Answer 27

It can promote certain organs to become secondary sites of metastasis. Once in the venous blood, CTCs are circulated via the arterial system, extravasating and colonising a suitable site. In ovarian cancer, metastatic cells can grow in the peritoneal cavity in the ascites fluid or by attaching to the surface of peritoneal organs. They don’t establish metastases in other visceral organs.
Some phenotypes establish regardless of blood flow eg. melanomas form metastases in lungs and pulmonary and ovarian tissue. This highlights the importance of the metastatic niche

Question 28

What are the 4 steps in the formation of the metastatic niche?

Answer 28

Priming, licensing, initiation, and progression.

Question 29

Explain what occurs in the priming stage of metastatic niche formation

Answer 29

Highly proliferative cells in the primary site become hypoxic and inflammatory - releasing cytokines, ECM remodelling enzymes, and exosomes (cell-cell communication). Exosomes are used to transport metastasis suppressing or encouraging factors between different cells which reach the premetastatic niche via circulation to prepare it for invasion - forming an immature premetastatic niche

Question 30

Explain what occurs during the licensing stage of metastatic niche formation

Answer 30

The ECM is remodelled by bone marrow derived cells and immune cells. They’re attracted by a variety of chemokines. These recruited cells and the host stroma remodel the local microenvironment and provide metastasis promoting cyto/chemokines - forming a mature premetastatic niche

Question 31

Explain what occurs during the initiation stage of metastatic niche formation

Answer 31

Circulating cancer cells arrive at the new site and begin extensive remodelling of the microenvironment to promote their growth. After braking down the existing ECM, cancer cells begin to produce their own ECM components. They also send signals to stromal cells to produce more matrix proteins. Cancer cells also secrete crosslinking enzymes to stiffen the ECM and increase integrin/focal adhesion signalling to favour metastasis. Leaving and reentering a new site involves changes in expression of different integrins depending on the environment. This forms a micrometastases

Question 32

Explain what occurs during the progression stage of metastasis

Answer 32

The pre-metastatic niche can host more cancer cells and result in increased tumour growth - forming a macrometastases

Question 33

Explain the concept of dormancy

Answer 33

Dormancy refers to the fact that metastasis can occur at any stage during the primary tumour development. This process can also pause at any point. Dormant tumour cells can reside in any tissue at any time, becoming reactivated to form a metastases. They’re called disseminated tumour cells. Dormancy is clinically the period of time between removal of tumour and relapse in a clinically disease free patient. It provides a therapeutic window to target metastatic cells

Question 34

Describe the two theories as to how dormancy arises

Answer 34

The disseminated tumour cells can be dormant simply due to being in a quiescent proliferative state.
Micrometastases can become dormant due to insufficient vascularisation or as a response to immune defences

Question 35

Explain how in vitro models are used to study metastasis

Answer 35

In vitro studies allow investigation of progression and initiation of metastasis, also responses to treatment. Wound healing assays, Boyden chambers, microfluidic devices, micropatterning and performing rings/paths. The cells response to different chemokines and organ derived proteins is assessed

Question 36

Explain how in vivo models are used to study metastasis

Answer 36

Amoebae, worms, flies, fish, mice. We use the genetic code in order to scale up the simple systems to complex human models.

Question 37

How is it possible for cells to cross basement membranes?

Answer 37

Using a nematode worm model, scientists discovered that the cell is arrested in G1 phase, and chromatin is modified and certain TFs are upregulated. One of these is FOS-1A (an orthologue to oncogene FOS) which is strongly associated with cancer cell invasion.

Question 38

Describe and explain the stages involved in cells crossing the basement membrane

Answer 38

Invasive machinery becomes polarised
Invadopodia breach the basmement membrane
The invasive protrusion displaces the basement membrane
The basement membrane shifts past the anchor cell

Question 39

What is meant by an orthotopic injection?

Answer 39

When cancer cells are injected into the site from which they’re derived, allowing to study early metastasis and primary tumour growth in mouse models.

Question 40

What is meant by a xenograft model?

Answer 40

When primary tumour models are investigated by injection the cancer cells subcutaneously into another area

Question 41

How can cancer cell circulation be studied?

Answer 41

Via an IV injection, allowing for the study of colonisation

Question 42

How can final stage colonisation and metastases be studied?

Answer 42

By injection the cancer cells into another area of the mouse