Immuno 20

Question 1

T or F. Cancer cells are typically very similar to normal cells, so the immune system does not efficiently “see” them as foreign

Answer 1

T.

Question 2

What is the major way the immune system can attack tumors?

Answer 2

CTLs that have specificity for tumor antigens(s) can recognize and kill tumor cells, sometimes.

Question 3

Cancers arise due to mutations of normal cellular ____.

Answer 3

proteins. These mutations can result in alteration of normal host determinants (to which the host is immunologically tolerant) to create new peptide determinants. Since these determinants were not available during T cell development and negative selection in the thymus, the host may have CD8 T cells that have specificity for the new “mutant” determinant.

The mutated protein will certainly be a source of peptides for loading onto newly synthesized MHC class I molecules by the tumor cells and will be available for immune surveillance by CTLs. This type of antigen is what is known of as a tumor antigen.

Question 4

How else can tumors arise?

Answer 4

When embryonic genes are reactivated during post-natal life and are over-expressed. When this occurs, the embryonic protein that is expressed will serve as a source of peptides for loading onto MHC class I molecules produced by the tumor cell, and those peptides will be presented to naïve T cells. Because the embryonic genes were not being expressed during T cell development and negative thymic selection, it is likely that there will be T cells that have specificity for peptides derived from the embryonic gene(s). This is an example of a tumor-associated antigen.

Question 5

T or F. A tumor-associated antigen is an un-mutated protein that is encoded on the germ-line DNA of a cell whose expression level has been dramatically altered by a neoplastic event/ process.

Answer 5

T.

Question 6

What is another example of a tumor-associated antigen?

Answer 6

In some cases, the mutation(s) that result in tumor formation result in increased expression of a normal self protein by the tumor cells. The increase in density of the normal self determinant can sometimes lead to recognition by effector CTLs. This is another example of a tumor- associated antigen.

A point to remember: tumor-specific antigens are expressed only by tumor cells; no non-tumor cells express a tumor antigen.

Question 7

What is an oncogene?

Answer 7

an oncogene is a mutated form of a proto-oncogene. A proto-oncogene is a gene that when mutated and activated can transform a normal cell into a tumor cell.

Question 8

What are some examples of oncogenes?

Answer 8

1) HER2 (growth factor receptor; breast cancer),
2) B-RAF (intracellular signaling; melanoma),
3) MYC (transcription factor; neuroblastoma),
4) RAS (intracellular signaling; colorectal cancer),
5) beta-catenin (intracellular signaling; array of tumor types), and
6) VEGF (angiogenesis; metastatic colorectal cancer)

Question 9

What are some examples of tumor-suppressor genes?

Answer 9

APC (scaffolding protein; mutated in colorectal cancers),

TP53 (regulates cell division and apoptosis; mutated in lung cancers),

RB (regulates cell division; mutated in retinoblastomas),

CDKN2A (regulates cell division; mutated in melanomas), and

CDK4 (regulates cell division; mutated in melanomas).

Question 10

Will MOST people form an immune response to mutated proto-oncogenes and tumor suppressor genes?

Answer 10

Yes. Mutated forms of these genes can serve as unique tumor antigens because the host’s T cell repertoire was generated before the mutant form of the protein was available for presentation during negative thymic selection.

Therefore, most people are not tolerized against peptides derived from the mutant form of these host proteins.

Question 11

Because tumor cells are rapidly dividing and genetically unstable cells, many mutations occur as the cells proliferate. Some of the mutations are in genes that have nothing to do with cell division or regulation of cell division. Any of these mutations could give rise to unique host determinants that could potentially be recognized by some of the CD8 T cells in the repertoire.

Answer 11

Because tumor cells are rapidly dividing and genetically unstable cells, many mutations occur as the cells proliferate. Some of the mutations are in genes that have nothing to do with cell division or regulation of cell division. Any of these mutations could give rise to unique host determinants that could potentially be recognized by some of the CD8 T cells in the repertoire.

Question 12

What is the significance of overexpressed or aberrantly expressed cellular proteins in tumors?

Answer 12

such tumors express antigens that would not normally be expressed by that particular tissue. These antigens can serve as tumor antigens in some cases.

Question 13

What is an example of overexpressed or aberrantly expressed cellular proteins in tumors?

Answer 13

Melanoma-antigen E (MAGE) family proteins- members of the cancer/testis tumor antigen family.

Question 14

What are the members of the MAGE family?

Answer 14

CAGE, BAGE, and RAGE proteins

Question 15

T or F. MAGE proteins are encoded on genes that are expressed in the testes, but are normally not expressed in any other tissues.

Answer 15

T.

Question 16

What is the significance of MAGE proteins typically being only expressed in the testes?

Answer 16

Because the testes are an immunoprivileged site, MAGE proteins are not available during T cell development and negative selection in the thymus. Therefore, most people will not be tolerized to MAGE proteins. When MAGE expression is upregulated in a tumor cell, it can serve as a target for CTL responses.

Question 17

Sometimes, unmutated oncogenes are over-expressed on tumor cells. Example?

Answer 17

The oncogene HER2/NEU that is expressed at high levels on a subset of human breast cancers. These over- expressed proteins are used as targets for treatment with monoclonal antibody drugs.

Question 18

What are Oncofetal antigens?

Answer 18

some genes that are normally expressed only during embryonic development are de- repressed in a tumor (tumor-associated antigens). Their protein products were not available during negative selection of B cells and T cells, so it is possible to make a potent immune response to these proteins

Question 19

T or F. most tumors express either abnormal forms or elevated levels of surface glycolipids and glycoproteins

Answer 19

T. these surface markers (e.g. blood antigens, mucins, gangliosides, etc.) can be used as diagnostic and sometimes even therapeutic targets of tumors.

Question 20

What are MUC-1 mucins? Associated cancer type?

Answer 20

mucins found on many breast carcinomas that have tumor specific epitopes that can be recognized by both CTLs and antibodies. For this reason, MUC-1 based vaccines are thought to have high potential for efficacy.

Question 21

What are cell type-specific differentiation antigens?

Answer 21

these are molecules that are normally expressed by a cell at different stages of differentiation of that cell type. A tumor cell will typically express antigens that were being expressed by the cell when the neoplastic event occurred.

Question 22

What cell type-specific differentiations antigen do B cell lymphomas express?

Answer 22

CD20 because it is a pan B cell marker.

Question 23

What cell type-specific differentiations antigen do developing lymphocytes express?

Answer 23

A developing thymocyte that undergoes a neoplastic event will express CD1 on its surface (T lymphoblastic leukemia, or T-all).

Question 24

What cell-type specific differentiation antigen would a T cell that undergoes a neoplastic event at any point after reaching the thymic medulla express?

Answer 24

CD3 and either CD4 or CD8 on its surface.

Question 25

Are host immune responses generated against cell-type specific differentiation antigens?

Answer 25

No. Because these are normally expressed proteins that were available during thymic negative selection in the host, the host should not normally have T cells in their repertoire that will recognize determinants derived from these antigens. So, tumor antigen-specific T cell responses will not be produced by the host.

These surface markers can be used as targets for diagnostic or therapeutic antibodies, however.

Question 26

What is Rituximab?

Answer 26

anti-CD20 mAb used as a therapeutic in some B cell lymphoma patients.

Question 27

Can viruses cause cancer?

Answer 27

yes, viruses like EBV, HBV and HPV are associated with some cancer forms

Question 28

What cancer is HPV associated with?

Answer 28

carcinoma of the uterine cervix and oropharyngeal cancer

Question 29

What cancer are HBV and HCV associated with?

Answer 29

hepatocellular carcinoma

Question 30

What cancer is EBV associated with?

Answer 30

Burkitt’s lymphoma, Nasopharyngeal carcinoma

Question 31

What are some mechanisms tumors use to evade/overcome host immune surveillance?

Answer 31

1) Tumors are genetically unstable, resulting in accumulation of mutations that allow then to evade pre-formed immune responses. This is essentially antigenic drift.
2) One-third to one-half of all tumors have defects is expression of MHC class I molecules. This allows them to evade CTL-mediated surveillance/killing. However, it does make them more of a target for NK cell-mediated killing. (NOTE: CTLs AND NKs are involved in tumor immune response)
3) Some tumors produce cytokines that downregulate immune responses (e.g. TGF-beta).
4) Some tumors express Fas-ligand on their surface, enabling them to induce apoptotic death of CTLs that interact closely with them
5) Some tumors express PD-L1. PD-L1 is a ligand for the protein PD-1 that is expressed on effector T cells. PD-1 is a negative regulator of T cell effector function. When PD-L1 binds to PD-1, it causes downregulation of T cell effector function.

Question 32

What antigen does the drug Trastuzumab target? Cancer type?

Answer 32

HER2/NEU protein, a growth factor receptor

Breast cancer

Question 33

What antigen does the drug Alemtuzumab target? Cancer type?

Answer 33

CD52, a differentiation antigen

Chronic lymphocytic leukemia

Question 34

What antigen does the drug Cetruximab target? Cancer type?

Answer 34

EGRF, a growth factor receptor

Colorectal and head/neck cancer

Question 35

What antigen does the drug Panitumumab target? Cancer type?

Answer 35

EGFR for colorectal cancer

Question 36

What antigen does the drug Bevacizumab target? Cancer type?

Answer 36

VEGF, a growth factor promoting angiogenesis

Colorectal and Non-small cell lung cancer

Question 37

What are some examples of drugs that consist of MAb that have specificity for a specific tumor antigen that have been conjugated to either a toxin, a drug, or a radionuclide?

Answer 37

Gemtuzumab, Ibritumomab, and Tositumomab

Question 38

What are the major uses of the conjugated mAbs described in the last slide?

Answer 38

The antibody conjugates that include a toxin or drug are designed to kill the targeted tumor cells. Those conjugated to radionuclides can be used either to kill the targeted tumor cell or diagnostically to “find” the location of tumors throughout the body.

Question 39

What is Gemtuzumab used to treat? What is it conjugated to?

Answer 39

Acute myelogenous leukemia (CD33)

Conjugated to ozogamicin, a cytotoxic antibiotic derivative

Question 40

What is Ibritumomab used to treat? What is it conjugated to?

Answer 40

Non-Hodgkins lymphoma. (CD20)

Conjugated to indium-111 for imaging and yttrium-90 for treatment

Question 41

What is Tositumomab used to treat? What is it conjugated to?

Answer 41

Non-Hodgkins lymphoma. (CD20)

Conjugated to indium-131

Question 42

How are conjugated antibodies used to kill tumor cells?

Answer 42

1) Upon administration of the drug to a patient, the antibodies bind only to the tumor cells, and deliver the toxin into the cell where it can exert its toxicity and kill the tumor cell.
2) A radioactive tumor-specific antibody has been conjugated to a radionuclide that, once delivered to the tumor cell, can actually irradiate and kill the targeted cell via radiation that damages cell DNA

Question 43

Antibody-based cancer treatments can act as antagonists to cell growth. How?

Answer 43

1) by binding to receptors causing steric hindrance of their ligand from binding to them (i.e. IL-6)
2) Other antibody drugs bind to the ligand (such as VEGF), preventing it from binding to its receptor. This is analogous to toxin or pathogen neutralization.
3) Other drugs can bind to an important receptor and cause its depletion by endocytosis (e.g. alpha:beta2 integrin).

Question 44

T or F. Some antibody-based drugs can act as agonists.

Answer 44

T. some mAb drugs bind to a receptor and either mimic signaling through that receptor, or promote signaling through that receptor by recruiting its ligand to the receptor.

Question 45

What are some other ways mAbs can suppress tumor growth?

Answer 45

1) Effectors of cell killing: some mAb drugs recruit host immune cells (NK cells or even CTLs to the tumor cell, resulting in its destruction.
2) If the target of these drugs is expressed at a high enough density, activation of the complement cascade can result in so much deposition of C3b and production of MAC that the host cell can be directly lysed by the MAC. MAC killing of host cells does not usually occur during an infection due to the soluble and surface bound complement control proteins, but these measures can be overcome with high copy numbers of antibodies bound directly to the host cells.
3) Some mAb drugs are conjugated to toxins, radionuclides, or drugs. These mAb-conjugates target the tumor by binding only to tumor antigens, bringing the toxin/radionuclide/drug in close enough proximity to the tumor cell that it can kill that cell.

Question 46

What is an example of an mAb that recruits NKs to cells to kill them via ADCC?

Answer 46

Rutiximab anti-CD20 to cause killing of B cell lymphomas

Question 47

What are some examples of antagonist mAbs?

Answer 47

Traztazumub, Pertuzumab, and T-DM1- all antagonists that cause reduction in HER-2 positive breast cancer tumor growth by preventing dimerization of HER2 and HER3 receptors

Cetuximab and Panitumumab

Question 48

What is T-DM1?

Answer 48

T-DM1 is a unique product in that it combines a MAb drug with a chemotherapy drug, making it a good drug for pre-treated HER2-positive breast cancers.

Question 49

How do Cetuximab and Panitumumab work?

Answer 49

Cetuximab and Panitumumab are mAb drugs that bind to the epidermal growth factor receptor (EGFR), preventing it from being activated by EGF binding. These drugs are antagonist antibodies as well.

Question 50

What is CTLA-4?

Answer 50

an important surface marker of Treg cells. When it binds to B7, Treg cells begin to secrete TGF-B, a very potent anti-inflammatory cytokine that can prevent naïve T cell activation.

Question 51

Where else is CTLA-4 found besides on T-regs?

Answer 51

CTLA-4 is also a surface marker of T cells that, when engaged with its ligand (B7.1 or B7.2), increases the threshold of signaling required for activation of naïve T cells and interfering with the ability of CD28 on the T cell from binding to B7. However, its exact role as a surface protein of naïve T cells is not clearly understood.

Question 52

What does Lpilimumab do?

Answer 52

mAb drugs (e.g. Lpilimumab) that have specificity for CTLA-4 serve as antagonists that prevent CTLA-4 from binding to B7 molecules. This prevents Tregs from performing their effector function, which is to prevent T cell activation on an APC that is presenting the Tregs cognate determinant.

Question 53

What is PD-L1?

Answer 53

A ligand that binds to PD-1 on the surface of effector T cells. PD-1 is a member of the CD28/CTLA-4 protein family that is expressed on the surface of effector T cells. When it is bound by is ligand (PD-L1 or PD-L2), it interferes with the effector function of the T cell. The PD-1 ligands are normally expressed on APCs in secondary lymphoid tissues, and I assume that their role is to help prevent CTLs from performing their effector functions within secondary lymphoid tissues, between the time that they are differentiating until they leave to enter the circulation.

Thus, Tumor cells that express PD-L1 are protected from CTL-mediated killing.

Question 54

mAb that is specific for either PD-1 or PD-L1 can prevent them from interacting. This prevents a tumor cell from down-regulating the effector function of CTLs. Treatment with these drugs causes rapid reduction in tumor size, although it is not yet known if patients treated with these drugs live any longer.

Answer 54

mAb that is specific for either PD-1 or PD-L1 can prevent them from interacting. This prevents a tumor cell from down-regulating the effector function of CTLs. Treatment with these drugs causes rapid reduction in tumor size, although it is not yet known if patients treated with these drugs live any longer.

Question 55

NOTE: In June of 2013, researchers reported that combining ipilimumab (anti-CTLA-4) and anti–PD-1 MAb drugs led to “deep and rapid tumor regression” in almost a third of melanoma patients.

Answer 55

In June of 2013, researchers reported that combining ipilimumab (anti-CTLA-4) and anti–PD-1 MAb drugs led to “deep and rapid tumor regression” in almost a third of melanoma patients.

Question 56

What are Bispecific antibodies?

Answer 56

recognize both CD3 and a surface target of tumor cells recruit cytotoxic T cells to tumor cells.

Blinatumomab is a single-chain bispecific antibody that is composed of an anti-CD3 arm that recognizes the T cell receptor and an anti-CD19 arm that recognizes a surface antigen that is found on B cell lymphomas. Recruitment of the T cells to the tumor cells in this way results in efficient tumor cell killing, as if the T cell had recognized its cognate peptide-MHC on the tumor cell target.

Question 57

What is CAR (chimeric antigen receptor)?

Answer 57

a novel personalized medical approach to tumor therapy. This therapy depends on the creation of chimeric CD8 T cells whose specificity is determined by an antibody specificity, instead of a T cell receptor specificity.

If a unique tumor antigen can be identified on a patient’s tumor, monoclonal antibodies specific for that determinant can be generated, and then the rearranged heavy and light chain coding regions can be cloned from those cells and used to produce a synthetic gene that encodes a single chimeric chain that includes the CD3 zeta-chain. T cells that have been genetically altered to express this construct can then be generated, amplified, and infused into the patient.

These CD8 T cells can now recognize the unique tumor antigen using their chimeric surface antibody, and because the CD3 zeta chain is part of the construct, intracellular signaling will occur. These CD8 T cells can be activated in secondary lymphoid tissues and the resulting CTLs can recognize the tumor antigen via their chimeric surface antibody and can be induced to degranulate on target cells that bear the unique tumor antigen.