Cytotoxic T and Natural Killer Cells

Question 1

What is the first step in the activation of the adaptive immune system, and how are cytotoxic T cells activated?

Answer 1

-Activation of T helper cells by Antigen presenting MHC-II DC
-PAMP-PRR-induced cytokines secreted by APC to stimulate T helper differentiation to direct the immune response
-T helper cells proliferate and secrete cytokines to stimulate antigen-specific Tc and B cells

Question 2

What are the two ways how an APC activates a cytotoxic T-cell?

Answer 2

1. Sequential: APC presents MHC-II antigens to T helper cells -> license to present it through MHC-I to naive T-cells -> Naive cell develops to a CTL
2. Simultaneous: Antigen presentation through MHC-I and MHC-II to T helper and naive T cell at the same time
   -> this way is more effective

-CTL dont require signal 2 (CD28 - CD80) or signal 3 (cytokines) anymore

Question 3

What are the two pathways used by cytotoxic effector T cells to kill their target cells and how are cytotoxic effector T cells different from neutrophils in terms of cell killing?

Answer 3

-Perforin/granzyme pathway and Fas/FasL pathway

-One difference is that Neutrophils secrete their granules by spreading them, and also causing damage to adjacent tissues, effector T cells get close to their target and release granules right upon the target cell

Question 4

What is the Perforin/granzyme pathway?

Answer 4

-Perforin monomers are exocytosed on the target cell to build Pores in the membrane -> now granzyme B (protease) can get in and activates caspases-3 by cleaving it -> APOPTOSIS

Question 5

IN CLASS QUIZ: Which PRR is involved in caspase-mediated pyroptosis with the help of inflammasomes?

Answer 5

NLRs -> activates inflammosomes -> activate caspase -> APOPTOSIS

-happens when a cell decides to undergo apoptosis bc infected

Question 6

Explain the Fas-FasL pathway?

Answer 6

-cytotoxic effector cells create vesicles with FasLigand
-Vesicle fuse with cytotoxic effector membrane and shift FasL to the cell surface

-Cell-Cell interaction to Fas receptor on the target cell (all cells should have that unless downregulated by a pathogen)
-Caspase cascade: Caspase 8 -> Caspase 3 -> death signal -> APOPTOSIS

Question 7

What are some T cell effector subset types?

Answer 7

Tc1 for intracellular pathogens: (similar to TH1) secrete IFN-γ but no IL-4
-> use perforin and Fas pathway to kill

Tc2 for extracellular worms: (similar to TH2) secrete IL-4 and IL-5, much less IFN-γ
-> use perforin to kill

If they use perforin and Fas why are they secreting IFN g and ILs for?? -> answered: for communication with other cells, maybe telling them which path to differentiate

-> maybe similar to T helper to activate macrophage killing (IFN-g) and B cell antibody production and Neutrophile activation (IL-4,5)

Question 8

What is the difference between a NK cell and a cytotoxic T c ell (CTL)

Answer 8

-NK cells are part of the innate immune system, they look like CTLs but they dont have TCR

-CTLs have fully recombined TCRs

-NKT cells are inbetween they look like NK cells but they have TCR (but not as variavle as CTLs)

Question 9

What is the function of NKT cells?

Answer 9

-They recognize glycolipids presented by CD1

-can act as a helper or killer cells (Fas-FasL)

-No memory cells
-have NK surface proteins, rather than T-cell varieties

Question 10

How do NK cells work?

Answer 10

-No TCR, no receptor gene arrangement
-NK cells help regulate innate/adaptive immunity with cytokines
-they recognize and destroy infected and cancer cells

How do NK cells recognize their targets, since they don’t have TCRs??? -> with activating receptors (no MHC as a signal to KILL) inhibitor receptors

Question 11

How does the NK cell response look like?

Answer 11

-They are innate, so they always circulate
-They can be upregulated with IFN-γ

-They recognize the balance of activating and inhibiting NK receptors, which tell to kill or not to kill a cell
-Lack of MHC-I is a major signal to kill -> bc viruses f.e. like to prevent MHC-I presentation to not expose themselves

-Once activating cell signal overweigh they kill by perforins/granzymes

Question 12

What is the purpose of the activating and inhibitng signal balance?

Answer 12

Every cell has activating signal (KILL) and inhibiting signal (NOT KILL) one reason is bc they undergo stress or something is going wrong (cancer, cellular damage) and in case they need to get killed they switch the balance and upregulate the activating signal

Question 13

How do NK cells “receive” their license to kill?

Answer 13

-By prior interaction with a healthy cell through MHC-I (inhibitory signal) and not killing it -> to see if it has the ability to not kill healthy cells

Question 14

Why do NK cells upregulate faster than CTLs

Answer 14

-Because CTLs have TCR specific for pathogens, the number of CTLs with specific TCR are fewer that’s why it takes time to build up the number; but NK cells don’t have TCR so there are more NK cells ready to be upregulated

Question 15

ORDER:

Question 16

How do cells fight mutations and why do they still turn into tumors?

Answer 16

-Cells undergo checkpoints throughout their cell replication cycle, whereby they are screened for mutations and repaired

-Mutations on checkpoint proteins disorders the checkpoint process, leading to the accumulation of mutations

Question 17

What is the difference between benign and malignant tumors?

Answer 17

-Benign: unable to invade surrounding tissues, and no indefinite grwoth

-Malignant: becomes more invasive, may metastasize and invade other distant tissues

Question 18

How does the immune system detect tumor cells, since they are self and immune cells should not recognize them? (bc of central tolerance (bone marrow and thymus) and peripheral tolerance)

Answer 18

4 groups of antigens recognized by T cells:
-Ag expressed only by tumors (f.e. viral proteins HPV)
-Ag of the mutated form of a normal gene
-Ag expressed at a certain stage of development
-Ag overexpressed in certain tumor

Question 19

IN CLASS QUIZ 1
If cancer is an intracellular disease, which T-helper response would be the best for it?

Answer 19

TH1

Question 20

What are tumor-specific antigens (TSA) and why are they not detected easily by t cells, since they are not self anymore?

Answer 20

-They are unique sequences leading to altered proteins (nonself),
-bc developing T cells are presented with thymus self cells (negative selection) and TSA are non-thymus cells

-TSA: best case bc now they are specific (f.e. prostate-specific antigen) and look like a different protein shown at MHC-I to activate T -cells by the cross-presenting process

How are they recognized since the self has no CD80 -> T effector doesn’t need signal 2 -> and now they recognize altered self-antigen as foreign -> KILL

Question 21

What are tumor-associated antigens?

Answer 21

TAAs are normal cellular proteins with unique expression patterns (escape peripheral tolerance bc they are not exposed a lot to T cells with CD28 - CD80 check)

->not exposed because: fetal cell and only expressed in fetal stage or antigens expressed at very low level or their expression is dysregulated

Question 22

How does the body deal with mutated cells?

Answer 22

1. intrinsic suppression: repair of DNA mutations or APOPTOSIS through inflammasomes
2. cytotoxic cells:
   -NK cells sense increased expression of activating ligands (signal to KILL) due to stress caused by f.e. unfolded proteins due to mutations

-IFN-γ and IL-12 encourage DC to activate strong TH1 and CTL response
(remember: IL-12 -> TH1 -> IFN-γ -> macrophage killing)

-T helper and cytotoxic can detect tumor antigens

Question 23

How do cancerous cells escape the host immune response?

Answer 23

-After multiple mutations, they get less responsive to immune clearance

-Downregulation of tumor antigens or NK activating ligands (stress) -> poor target for NK cells
-Shutting down the Fas-FasL pathway (don’t express Fas anymore) -> poor target for CTLs
-reduced MHC expression -> poor target for CTLs and NK cells

Question 24

Further strategies of cancer cells to escape immunity:

Answer 24

-Loss of adhesins -> cells use adhesins to stick to each other -> without adhesins tumor cells can migrate to blood vessels and metastasize

-Expression of anti-inflammatory markers (IL-10 and TGF-ß) to show that there is no infection going on -> sensed by Treg

Question 25

How do Immunotherapies fight against tumor cells?

Answer 25

-Chemotherapy: blocks DNA synthesis and cell division -> but you also stop the proliferation of healthy cells (f.e. immune cells)

-Hormonal therapies: interfere with tumor cell growth

-Targeted therapies: small molecule inhibitors -shutting down the expression of specific proteins

-Immunotherapies: enhance the antitumor immune response

Question 26

How are TAA-loaded DCs used against specific tumor cells?

Answer 26

TAAs are the proteins usually expressed on a low level (f.e. prostate cancer Ag or PAP), signifying it is a human peptide -> so you could do a biopsy and take PAPs and culture them with DCs (important to take the patients DC bc compatibility with MHC)

So the DC will now take up PAPs antigen on MHC-II, and if we add the cytokines it gets licensed to present it through MHC-I (cross-presentation)-> so we skipped the part whereby T helper would have licensed

The DCs are replicated to a high number and reinfused into the patient to induce the immune response against prostate tumor cells

Question 27

How are Antitumor cells used against tumor cells?

Answer 27

IL-12 is responsible for the proliferation of T-cells

If there are some tumor-specific T-cells in the body, we can take them out and expose them to IL-2, so that they can build up the number, since some T-cells don’t sufficiently proliferate bc being exposed to IL-10 (anti-inflammatory signal stating that everything is fine) -> Reinfuse into the patient

-Downside: it is patient-specific (everyone has their own MHC, and own T-cells) -> COSTLY

Question 28

How can we fight tumors by blocking Anti-inflammation?

How can we use APC properties against tumor cells?

Answer 28

-
-Normal cells which are not antigen-presenting don’t have the co-stimulatory signal B7 (CD80/86) and cant bind to T helper cells -> so we can take tumor cells and transfect them with the CD80 gene -> so that they start expressing CD80 and get recognized by CTLs and their CD28 -> so the tumor cell has been turned into an APC

Question 29

How can we fight tumors by blocking Anti-inflammation?

Answer 29

CTLA-4 is expressed after a few days to stop the inflammatory response and T-cell activation

-this 2-day limit provided by CTLA-4 is not enough to fight tumor cells -> so we can block CTLA-4 with antibodies

-APCs express inhibitory receptors to prevent CTL killing during cross-presentation -> tumor cells express these inhibitory receptors to prevent CTL killing through mutation -> so we use antibodies to block them -> CTL killing

What if these Ab also block DCs expressed inhibitory receptors??

Question 30

How are chimeric antigen receptors used?

Answer 30

-In the body Abs are generated with light and heavy chains with their variant domains (heavy and light) on the ligand binding site

-we can build these Abs in the Lab -> to create an Ab against a tumor-specific antigen and put them into T cells to target these particular tumor cells

we activate the T cells by mimicking Signal 1, signal 2, and signal 3
->modified antigen-binding sites
->modified signal domains for a specific signaling pathway

create some that don’t have MHC ???

Question 31

What is the first step in the activation of the adaptive immune system, and how are cytotoxic T cells activated?

Answer 31

Antigenpresentation MHC-II DC