Cancer and the Immune System

Question 1

The role of the immune system is what?

Answer 1

Maintain tissue homeostasis

Question 2

What resides within all healthy tissue ?

Answer 2

Many immunocytes particularly t cells along with sentinel myeloid dendritic cells and macrophages. Their role is to be conscious to damage to the tissue either through infection with bacteria, viruses but also in terms of transformation.

Question 3

Explain the schreiber immunosurveillance immunoediting model of tumourigenesis?

Answer 3

Normal cells subject to common oncogenic stimuli ultimately undergo transformation and become tumour cells

At early stages of tumourigenesis, these cells may express distinct tumour-specific markers and generate pro-inflammatory “danger” signals that initiate the cancer immunoediting process

Cancer immunoediting is an extrinsic tumor suppressor mechanism that engages only after cellular transformation has occurred and intrinsic tumor suppressor mechanisms have failed. In its most complex form, cancer immunoediting consists of three sequential phases: elimination, equilibrium, and escape.

Elimination: innate and adaptive immunity work together to destroy developing tumors long before they become clinically apparent.If this phase goes to completion, then the host remains free of cancer, and elimination thus represents the full extent of the process.

If, however, a rare cancer cell variant is not destroyed in the elimination phase, it may then enter the equilibrium

Equilibrium: Tumour outgrowth is prevented by immunologic mechanisms. T cells, IL-12, and IFN-γ are required to maintain tumor cells in a state of functional dormancy, whereas NK cells and molecules that participate in the recognition or effector function of cells of innate immunity are not required; this indicates that equilibrium is a function of adaptive immunity only. Editing of tumor immunogenicity occurs in the equilibrium phase. Equilibrium may also represent an end stage of the cancer immunoediting process and may restrain outgrowth of occult cancers for the lifetime of the host. However, as a consequence of constant immune selection pressure placed on genetically unstable tumor cells held in equilibrium, tumor cell variants may emerge that (i) are no longer recognized by adaptive immunity (antigen loss variants or tumors cells that develop defects in antigen processing or presentation), (ii) become insensitive to immune effector mechanisms, or (iii) induce an immunosuppressive state within the tumor microenvironment. These tumor cells may then enter the escape phase, in which their outgrowth is no longer blocked by immunity. These tumor cells emerge to cause clinically apparent disease.

Question 4

What is the immunotherapy cure rate for metastatic melonoma?

Answer 4

50%

Question 5

How is the immune system classified?

Answer 5

Humoral and cellular

Further divided into innate and adaptive

Innate: “Hard coded”, No memory and rapid

Adaptive: Somatic recombination, memory, can take time

Humoral Innate: C reactive protein, Complement Cytokines

Humoral Adaptive: Antibodies, Cytokines

Cellular innate: Neutrophils, Eosinophills, Basophils, Monocyte/macrophages, dendritic cells and natural killer cells

Cellular adaptive:
B cells, Alpha beta T cells (CD4 & CD8 T cells)

Question 6

What bridges humoral innate and adaptive immunity?

Answer 6

Natural antibodies

Question 7

What bridges cellular innate and adaptive immunity?

Answer 7

NK T cells

yo T cells

Question 8

What are T Cells

Answer 8

Adaptive immune cells

Capacity to see virtually any organism

Use the T cell receptor (TCR) to see things

See things presented to them on MHC

CD8 T cells can directly kill cells that do not look healthy (e.g. virus infected, cancerous)

Question 9

Explain the major histocompatibility complex class 1

Answer 9

Self protein synthesis

Some proteins degraded by proteosome, this chops protein to short fragments which then are loaded onto surface of MHC class 1 molecule

Which is then exported onto surface of cells and provides immune system ability to look into healthy of this cell.

“Endogenous” peptide allows CD8 T cells to see what’s going
on inside

Question 10

Explain the major histocompatibility complex class 1 when a virus is present

Answer 10

Viruses are obligate intracellular parasites because they absolutely depend on host cell for protein translation as do not have any organelles.

Virus enters nucleus, transcribes into mRNA and this leads to viral protein synthesis.

Most of viral protein will be incorporated into viral membrane, some however will be degraded by proteosome.

In virally infected cell, the cell is aware there is a viral infection and is able to change this proteosome to a stress proteosome and this produces more peptides.

Peptides that are chopped are then loaded onto MHC class 1 which is exported onto surface of cell and is now presenting a foreign peptide. If CD8 is activated to see foreign peptide, is then aware that this exists and then will kill this cell.

Question 11

Explain the major histocompatibility complex in cancer

Answer 11

In cancer mutant/ abnormal protein typically arise, some of which are degraded by proteosome and loaded on MHC class 1 and others are not.

MHC class 1 alerts CD8 cell which then kills cell

Question 12

T cell receptor diversity exceeds the human genome coding capacity

How is this possible?

Answer 12

To generate diversity, RAG enzyme is expressed

RAG- Recombination activated gene

RAG leads to somatic recombination of each Variable, Diversity, Junction gene from B chain locus and this somatic combination attains combinatorial diversity. This will not attain 10 to the 8 diversity

This diversity achieved because as part of RAG complex one of the enzymes within complex Terminal deoxynucleotidyl transferase(TdT), transfers extra nucleotides into junctions between V, D and J and this leads to junctional diversity, this process is random (junctions can be very long or short). This results in random diversity of T cells and thus may generate T cell receptors that are entirely ineffective or some that react again self which leads to autoimmunity and ultimately tissue damage- opposite of role of immune system.

Development within the thymus deletes many but not all- thymus important for tolerising against cells and developing t cells but this process is not 100%

Question 13

RAG is only switched on in what?

Answer 13

Thymus: T cells

Bone marrow: B cells

Question 14

What is peripheral intolerance?

What does this highlight?

Answer 14

Process whereby a mature naïve T cell that only receives one signal via T cell receptor becomes anergic. Important as if this t cell were to observe an antigen, the most likely antigen it will observe is self antigen, likely a healthy one and so the safest response is to become anergic.

Immune system is built to tolerate self peptides.

Question 15

Mature naïve T cell encounters a dendritic cells (professional antigen presenting cells, role is to present antigens to T cells)

Here dendritic cells in tissue has transported bacteria in tissue, digests bacteria, and presents bacterial peptide on MHC

In this case here T cell is a perfect match for bacterial antigen in the context of MHC with this T cell receptor

What happens to CD8 T cell?

Answer 15

A naïve T cell that only receives TCR signal (signal 1) becomes anergic

Important as within gut billions of bacteria exist which may be pathogenic, however in gut are perfectly symbiotic and responding against them would lead to colitis

Question 16

Dendritic cell resides within tissues that display tissue damage – inflammation and inflammatory cytokines.

Stressed cells present which may be releasing extra signals such damage associated molecular patterns.

Inflammation, stressed cells, damage associated molecular patterns inform dendritic cell that tissue in which it resides in is not healthy and that it is inflamed.

Dendritic cell then transfers damage to MHC and signals to T cell that the tissue is unhealthy.

Thus now bacterial antigen is presented with a second signal of danger/inflammation/unhealthy tissue.

Second signal is CD80 and CD86, once both presented they engage a molecule on the t cell called cd28 which delivers a second signal to the t cell

What happens to CD8 T cell?

Answer 16

A naïve T cell that receives TCR signal (signal 1) AND ”co- stimulation” (signal 2) becomes activated and becomes a “licensed” memory cell (no longer naïve)

Question 17

Once a naive t cell becomes a licensed memory t cell what occurs?

Answer 17

Co stimulation is not obligatory to kill infected/cancer cell

Question 18

How is autoimmunity prevented

Answer 18

If have a stressed cell for example after an infection/chemical damage/transformation, the stressed cell may be releasing its own peptides and these peptides will be presented on MHC.

Thus now self peptide presented in context of MHC along with a second signal here. Danger here is activation of self reactive CD8 t cells.

Many autoimmune diseases that are clinically relevant are probably triggered by initial infectious or inflammatory insults within the tissues

There is a further, potent regulation of this process to try and prevent autoimmunity – presence of T suppressor t regulatory cells (Treg), this recognises self peptide but it expresses on it several inhibitory molecules –CTLA4 and this molecule binds CD80 and CD86 with a much higher affinity than CD28 and this means if there is a Treg present and this is specific to the self peptide then this will sequester this signal 2 and remove signal 2 from CD8 t cells. Thus one signal left and t cell is anergised.

T cell also excrete large amount of interleukin 10, ( immunosuppressive cytokine that prevents activation and proliferation of other T cells)

Finally T-regs are full of interleukin 2 receptors and thereby it acts as a IL-2 sink and all of this occurs locally to suppress any t cells sat around this dendritic cell expressing self peptide

Question 19

What do all t cells need to group and therefore is expressed by all t cells?

Answer 19

Cytokine interleukin 2

Question 20

Stressed cell (red) e.g through prolonged inflammation, bystander damage, metabolic cost or generally bad

If CD8 t cell activated and specific for this cell will secrete cytokines and try to kill stressed cell

Issue is that cytokines are not hugely specific, most will be focused on this cell but some of them will leak and affect surrounding healthy cells

The purpose of the immune system is to maintain tissue homeostasis – thus do want to rid of stressed cell but do not want to kill other normal cells

How can this be overcome?

Answer 20

Thus part of an inbuilt physiological mechanism to protect tissues against excessive CDH killing or excessive inflammation several mechanisms happen.

1. Cytokines that the CD8 t cell secrete also activate negative regulator of CD8 t cells, thus all cd8 t cells will express receptor PD1 and in epithelial tissues when CD8 t cells activated is that a negative feedback mechanism (upregulation of PDL1) occurs. Binding of PDL1 to pd1 suppresses CD8 cell, makes it less likely to secrete cytokines and less likely to divide
2. Other mechanism is several tregs arise and suppress t cells and this is a physiological mechanism to prevent excessive inflammation within tissues

Question 21

Stressed cell (red) e.g through prolonged inflammation, bystander damage, metabolic cost or generally bad

If CD8 t cell activated and specific for this cell will secrete cytokines and try to kill stressed cell

Issue is that cytokines are not hugely specific, most will be focused on this cell but some of them will leak and affect surrounding healthy cells

The purpose of the immune system is to maintain tissue homeostasis – thus do want to rid of stressed cell but do not want to kill other normal cells

How can this be overcome?

Answer 21

Thus part of an inbuilt physiological mechanism to protect tissues against excessive CDH killing or excessive inflammation several mechanisms happen.

1. Cytokines that the CD8 t cell secrete also activate negative regulator of CD8 t cells, thus all cd8 t cells will express receptor PD1 and in epithelial tissues when CD8 t cells activated is that a negative feedback mechanism (upregulation of PDL1) occurs. Binding of PDL1 to pd1 suppresses CD8 cell, makes it less likely to secrete cytokines and less likely to divide
2. Other mechanism is several CTLA-4 + tregs arise and suppress t cells and this is a physiological mechanism to prevent excessive inflammation within tissues

PD-L1 and CTLA-4 + Treg infiltration are PHYSIOLOGICAL responses to inflammation to limit excessive tissue damage (not cancer specific)

Question 22

What checkpoints exist in the physiological setting?

Answer 22

CTLA-4 is important peripheral tolerance mechanism

PD1/PDL1 axis is important in protecting tissues from excessive inflammation

Question 23

Self proteins are more likely to what? tolerize immune system

Answer 23

Tolerize immune system

Question 24

Likelihood of responding to antigens is dependent upon how similar they are to self, thus if very similar to healthy human self, peripheral tolerance mechanism is much more likely to suppress any immune response generated

Whereas if different i.e. from a different species, more likely to elicit immune response as tregs will not be specific to staphylococcus antigen for instance

Issue is that cancer is mutant self and closer to healthy human cell

How can this be overcome?

Answer 24

However now understand how immune checkpoint works, can try to block it, in particular CTLA4 which inhibits CD80 and CD86 simulation and can use drug or antibody (Iipulimubab) which binds to CTLA4 and prevents association with CD80 and CD86 thus restoring co-stimulation to t cells which may be searching for a peptide.

Question 25

Why is chronic inflammation an issue in cancer?

Knowing this what has developed?

Answer 25

Cancer develops over many years, thus within cancerous tissues T cells entering in trying to kill cancer over many years or decades leads to chronic inflammation

In the setting of chronic inflammation and release of these cytokines negative regulatory factors such as PDL 1 will be induced, and eventually these may well overcome the positive immune surveillance from T cells and these T cells are suppressed and cancer eventually escapes

Drugs that specifically bind either PDL1 or PD1 (pembrolizumab, nivolumab) and block this interaction

These drugs bind to PD1 and block association of PDL1 with PD1 thus restoring some of the function of these CD8 t cells

Question 26

Why is neoantigen burden instead of mutational burden used to examine effect of patients with melanoma treated with Anti CTLA4?

Answer 26

Patients with much higher mutational burdens survived longer than patients with low mutants upon treatment

Overall mutational burden is a crude measure of differences as can have synonymous or non synonymous mutants

Depending on where mutant resides it may or not may be presented on the appropriate MHC

e.g mutant in anchor amino acids means may not be able to bind to MHC class 1 .And if you cant do this you cant be seen by t cell and killed

Such crude segregations just regarding mutational burden are not used and what is used instead is neoantigen burden, to be a neoantigen must be a non synonymous mutation and must be presented on MHC molecule

If now only examine neoantigens, find that those with neoantigen burden survive longer than those without

More accurate than just a crude mutational burden

Thus must look very different to self and be presented to t cells for this anti ctla4 treatment to work

Question 27

1. Sequencing across what can give us a unique molecular fingerprint?
2. What can this track?

Answer 27

TCR

Track where these cells go, how they expand. Powerful scientific and clinical tool

Question 28

1. What was found when patients treated with anti PD1

2. Why was this found?

Answer 28

1. Patients with more oligoclonal, presumably t cell repertoire dominated by anti tumour t cells respond and progress better
2. Thus those that respond have a much more oligoclonal t cell repertoire than those who do not and this is determined by sequence across that variable region

Polyclonal- not dominated by any single clone. Might expand that they are not as successful as
less of their repertoire is composed of tumour reactive cells t cells

Oligoclonal- Dominated by certain clones

Question 29

What are T Cell Lymphomas?

Answer 29

Malignant CLONAL proliferation

TCR is a passenger molecular fingerprint

Can track disease relapse using presence of clonal T lymphoma TCR sequence

Question 30

1. When attempting to track T cell lymphomas the gold standard is flow cytometry. What steps are involved in this?

Answer 30

1. Usually use induction chemotherapy to reduce lymphoma burden below limit of detection

When lymphoma burden is below limit of detection typically pursue stem cell transplant

Because it is an allogenic transplant would usually follow with immunosuppression, and these stem cells will repopulate bone marrow and from this will produce new immune cells, allogenic immune cells important as they will be able to fight off any residual lymphoma cells called t graft verses lymphoma effect which is effective for preventing relapse

2. This is all below the limit of detection (behind black box) thus when run flow cytometry difficult to understand when to start and stop immune suppression

Question 31

What is a limitation of using flow cytometry to track t cell lymphoma and what are the consequences of this?

Answer 31

This is all below the limit of detection (behind black box) thus when run flow cytometry difficult to understand when to start and stop immune suppression:

1. Late detection of relapse,
   late withdrawal of immunosuppression and disease takes off
2. Sensitivity- Early detection of relapse and withdrawal of immunosuppression
3. Specificity- False positive relapse, early withdrawal of immunosuppression and GVHD

Question 32

What is an alternative to using flow cytometry to detect t cell lymphoma?

Why is this advantageous?

Answer 32

TCR sequencing

2. Molecular remission more sensitive than flow cytometry

Molecular remission predicts prolonged remission

TCR sequence - more specific for relapse