Tumor Immunology

Question 1

Describe the thoery of immune surveillance.

Answer 1

• Immune surveillance is the theory that the immune system scans the body to not only recognize and destroy invading pathogens, but also to recognize and destroy host cells that become cancerous
• Perhaps potential cancer cells arise frequently throughout life, but the immune system usually destroys them as fast as they appear
  
  • Compared to the traditional modalities of cancer treatment—radiation, chemotherapy, and surgery—immunotherapy promises the new concept of specificity
• Despite tumor immune surveillance, tumors do develop in the presence of a functioning immune system, and therefore the updated concept of tumor immuno-editing was developed, and is a more complete explanation for the role of the immune system in tumor development and has led to therapeutic strategies

Question 2

What is cancer immunoediting, and what are the main steps?

Answer 2

• Cancer immuno-editing 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 immuno-editing consists of three sequential phases:
    
    • elimination
    • equilibrium
    • escape
• Elimination:
  
  • The elimination phase is best described as an updated version of cancer immuno-surveillance, in which the innate and adaptive immune systems work together to detect the presence of a developing tumor and destroy it before it becomes clinically apparent
  • If tumor cell destruction goes to completion, the elimination phase represents an endpoint of the cancer immuno-editing process
• Equilibrium
  
  • ​Rare tumor cell variants may survive the elimination phase and enter the equilibrium phase, in which the adaptive immune system prevents tumor cell outgrowth and also sculpts the immunogenicity of the tumor cells
  • We envisage equilibrium to be the longest phase of the cancer immuno-editing process—perhaps extending throughout the life of the host
  • As such, it may represent a second stable endpoint of cancer immuno-editing
  • In equilibrium, the immune system maintains residual tumor cells in a functional state of dormancy, a term used to describe latent tumor cells that may reside in patients for decades before eventually resuming growth as either recurrent primary tumors or distant metastases
  • Equilibrium thus represents a type of tumor dormancy in which outgrowth of occult tumors is specifically controlled by immunity
• Escape
  
  • ​In the escape phase, tumor cells that have acquired the ability to circumvent immune recognition and/or destruction emerge as progressively growing, visible tumors
  • Progression from equilibrium to the escape phase can occur because the tumor cell population changes in response to the immune system’s editing functions and/or because the host immune system changes in response to increased cancer-induced immunosuppression or immune system deterioration

Question 3

What are some potential immune escape mechanisms that cancers have?

Answer 3

• At the tumor cell level, alterations leading to reduced immune recognition (such as a loss of antigens) or increased resistance to the cytotoxic effects of immunity (for example, through induction of anti-apoptotic mechanisms involving persistent activation of pro-oncogenic transcription factors such as STAT3 or expression of anti-apoptotic effector molecules such as BCL-2) promote tumor outgrowth
• Loss of tumor antigen expression is one of the best-studied escape mechanisms, and it can occur in at least three ways:
  
  • through emergence of tumor cells that lack expression of strong rejection antigens
  • through loss of major histocompatibility complex (MHC) class I proteins that present these antigens to tumor-specific T cells
  • through loss of antigen processing function within the tumor cell that is needed to produce the antigenic peptide epitope and load it onto the MHC class I molecule
• All of these alterations are probably driven by a combination of genetic instability inherent in all tumor cells and the process of immunoselection
  
  • The end result is the generation via a Darwinian selection process of poorly immunogenic tumor cell variants that become “invisible” to the immune system and thus acquire the capacity to grow progressively
• Cancer escape may result from the establishment of an immunosuppressive state within the tumor microenvironment
  
  • Tumor cells can promote the development of such a state by producing immunosuppressive cytokines such as:
    
    • vascular endothelial growth factor (VEGF)
    • transforming growth factor–β (TGF-β)
    • galectin
    • indoleamine 2,3-dioxygenase (IDO)
  • Can also recruit regulatory immune cells that function as the effectors of immunosuppression
    
    • Regulatory T cells (Treg cells) and myeloid-derived suppressor cells (MDSCs) are two major types of immunosuppressive leukocyte populations that play key roles in inhibiting host-protective antitumor responses
    • Treg cells are CD4+ T cells that constitutively express CD25 and the transcription factor Foxp3
    • When stimulated, they inhibit the function of tumor-specific T lymphocytes by producing the immunosuppressive cytokines IL-10 and TGF-β; by expressing the negative co-stimulatory molecules CTLA-4, PD-1, and PD-L1; and by consuming IL-2, a cytokine that is critical for maintaining CTL function
    • MDSCs are a heterogeneous group of myeloid progenitor cells and immature myeloid cells that inhibit lymphocyte function by inducing Treg cells; producing TGF-β; depleting or sequestering the amino acids arginine, tryptophan, or cysteine required for T cell function; or nitrating T cell receptors or chemokine receptors on tumor-specific T cells

Question 4

What is a tumor antigen and what are the different types?

Answer 4

• Tumor cells can be shown to have antigens that are not readily found on the corresponding normal cell
  
  • ​Often they are found on normal cells, but in much lower quantities
    
    • they are overexpressed or abnormally expressed by the tumor
  • Such antigens are called tumor-associated antigens (TAA)
    
    • A subclass of TAA are those that can be recognized by the immune system, in a way that leads to the destruction of the tumor
    • Such antigens are often called tumor rejection antigens
• Viral gene products
  
  • Many tumors are known to be caused by tumor viruses; in humans about 20% of tumors are caused directly or indirectly by viruses
  • Especially noteworthy are HTLV-1 and -2 that have been strongly implicated in Acute T-cell lymphoma/leukemia, an epidemic lymphoma in Japan and the Caribbean
  • Cervical cancer (human papilloma virus) is currently the best-known virally-induced tumor in humans; one hopes the HPV vaccine will make it less well known
  • Similarly liver cancer in the developing world follows a hepatitis virus infection
  • Epstein Barr Virus can induce Burkitt lymphoma and nasopharyngeal carcinoma
  • The presence of the bacterium Helicobacter pylori is associated with gastric carcinomas
• Mutant gene products
  
  • Chemical and physical mutagens can lead to cellular transformation
  • Mutated proteins will be processed and presented to the immune system
  • Since the mutations contributing to the development of tumors are not always identical from patient to patient, immunotherapy designed against these antigens may not be as generalizable as might be with viral or normal gene products
  • These antigens are called tumor-specific antigens
• Normal gene products (described in a separate card)

Question 5

What are the normal gene products that can become tumor antigens?

Answer 5

• Oncofetal antigens are made in normal fetal tissues
  
  • They are not found in the normal tissues of adults, but can be re-expressed in the tumor
  • The most familiar is carcinoembryonic antigen (CEA), found in the blood of patients with colon carcinoma and other cancers
    
    • There are commercially available kits to detect CEA in blood. They should not be used as a routine screening test. Why not? Too many false positives
  • The proper use of CEA measurement comes when you have a high index of suspicion of colon cancer; or, when such a cancer has been removed, to confirm complete excision (levels fall to 0 and remain there) or to warn of recurrence.
• Differentiation antigen
  
  • ​​These lineage-specific antigens can be greatly overexpressed in tumors, and they represent the most frequently identified TAA
  • The best studied are those from malignant melanoma (tyrosinase, gp100, MelanA/MART-1)
  • In 30% of breast and ovarian cancers overexpression of the human EGFR-2 gene product (HER- 2/neu) is observed
    
    • Therapeutic antibody and T cell responses to HER-2/neu can be induced
  • Prostate-specific antigen (PSA) appears in the blood of many men with prostate cancer, and its detection is used in screening programs, though its utility as a guide for treatment has recently come into question
• Clonal antigens
  
  • Expressed uniquely on the malignant clone
  • The most familiar example would be the idiotype of the surface immunoglobulin in monoclonal B cell malignancies, or of the TCR in T cell malignancies

Question 6

What are the ways that the immune system can kill tumor cells?

Answer 6

• Cytotoxic T cells
  
  • ​These CD8+ T cells (CTL) are probably the most important cells in tumor resistance, and they’re every right-thinking immunologist’s favorite cell
  • CTL can recognize TAA presented by MHC class I
  • Naive T cells are activated in the lymph nodes, not at the tumor site, via interactions with antigen-presenting dendritic cells
  • Following the initial activating event, the CD8+ T cells undergo clonal expansion and acquire lytic function
  • Activated TAA- specific T cells leave the lymph node and migrate to the tumor
  • CTL can kill tumor cells by inducing apoptosis via either perforin or Fas-mediated pathways
  • The CTL also secrete IFN-Gamma upon engagement of their TCR, which attracts and stimulates macrophages
• Th1 cells
  
  • These CD4+ T cells:
    
    • recognize the tumor antigens
    • make lymphokines
    • attract angry M1 macrophages
  • How can we make people with tumors get this system going better than it is?
    
    • Th1 stimulation may be part of the goal for the development of therapeutic cancer vaccines (remember, they also help CTL get activated.)
• Natural Killer (NK) cells
  
  • ​NK cells look like large lymphocytes, but have peculiar granules in their cytoplasm, so they are usually called LGLs (large granular lymphocytes)
  • They do not need to come from an immunized host to recognize and destroy quite a wide range of tumors, mostly of hematopoietic origin
  • NK cells provide a link between the innate and adaptive immune systems since they have the lethal tendencies of CTL, and the pattern recognition of innate immunity
  • NK cells have receptors of broad specificity for “stress” markers such as might be expressed on a growth-dysregulated cell
  • They also have receptors for MHC class I (unlike TCR, the receptors are not clonally variable; they bind just about any MHC Class I, with or without a peptide in it)
  • Binding of MHC class I suppresses NK cells, so they don’t waste effort trying to kill cells with a lot of MHC Class I; that, after all, is a job for CTL
  • What cells might have low MHC expression? Many tumor cells downregulate MHC to avoid CTL
    
    • But that makes them into NK targets! Between CTL and NK cells the immune system gets tumors coming and going
  • NK cells also have Fc receptors for IgG, and so can target antibody-bound cells even if they are not stressed NK targets
  • When this happens the phenomenon is called antibody-dependent cell-mediated cytotoxicity, ADCC
    
    • This is a very effective way of killing tumor cells
• Macrophages and neutrophils:
  
  • Can be activated in vitro with foreign products (e.g., bacteria) to kill tumor cells
  • Much of this antitumor activity can be attributed to TNF
  • However, tumors frequently learn to subvert macrophages and even recruit them to support tumor growth (turning M1 into M2, N1 into N2 for example)
    
    • Tumors frequently protect themselves by creating an environment in which M2 and N2, not M1 and N1, macrophages are favored
  • M2/N2 seem to protect tumors, even encourage their growth
• Antibody and complement
  
  • ​An antibody response is commonly made in tumor-bearing hosts, but it is not commonly effective
  • Opsonization of tumor cells by antibody and complement can kill some leukemias in vitro, but a strong B cell response to tumor antigens does not seem to correlate with resistance to the tumor
  • We see tumors that have survived immunoediting; the cells of the tumor that survive are likely to have downregulated antigen expression as much as they can
  • Others have become resistant to complement, or can inactivate it

Question 7

What is active immunotherapy?

Answer 7

• The host actively participates in mounting an immune response
  
  • Specific activation using vaccines:
    
    • Hepatitis B vaccine useful against development of hepatocellular cancer.
    • Human Papillomavirus (HPV) vaccine (Gardasil) has been successfully used to prevent cervical cancers
  • Nonspecific activation which results in stimulation of generalized immune response is achieved by immunization with:
    
    • Bacillus Calmette-Guerin (BCG) mycobacteria.
    • Corynebacteriumparvum
    • These microbes lead to activation of macrophages which are tumoricidal

Question 8

What is passive immunotherapy?

Answer 8

• This involves transfer of preformed Abs, immune cells and other factors into the hosts
• Specific: Preformed Abs or CTL directed against tumor Ags are used in the treatment of tumors
  
  • Antibodies against tumor Ags (e.g. Abs against Her2/Neu for treatment of breast cancer)
  • Agents directed against the interleukin-2 receptor (IL-2R) are used in the treatment of Human T lymphotropic virus 
(HTLV-1)-induced adult T cell leukemia as this virus infects T cells and leads to production of IL-2 
that binds to IL-2R and induces the T cell proliferation as well as other forms of cutaneous T-cell lymphomas
  • Abs against CD20 (rituximab) expressed on all B cells are used in the treatment of non Hodgkin’s B cell lymphoma
    
    • These Abs bind to tumor Ags on the cell surface and activate complement (C’) to mediate tumor cell lysis
    • In addition, Fc receptor bearing cells such as NK cells, macrophages and granulocytes may bind to the Ag-Ab complexes on tumor cell surface and mediate tumor cell killing through Ab-dependent cell-mediated cytotoxicity
  • Abs conjugated to toxins, radioisotopes and anti-cancer drugs have also been used. These enter the cells and inhibit protein synthesis because of the toxin. e.g. anti-CD20 conjugated to Pseudomonas toxin or ricin toxin has been used in the treatment of B cell tumors.
There are several problems with the use of Abs:
    
    • Abs are not efficient because the tumor Ags are associated with class I MHC Ags
    • The tumors may shed Ag or Ag-Ab complexes. Thus, immune cells cannot mediate tumor 
destruction
    • Some Abs may not be cytotoxic
    • Abs may bind nonspecifically to immune cells expressing the Fc receptors which include NK 
cells, B cells, macrophages and granulocytes without binding to tumor cells
  • Dendritic cells pulsed with tumor Ags may be administered which can present tumor Ags in the context of class II MHC to tumor-specific Th cells
    
    • As tumor Ags are usually not known, tumor lysates are used
    • The Th cells may in turn produce cytokines which lead to development of CTL activity
    • On the other hand, the dendritic cells may be transfected with the gene for tumor Ags, in which case, the Ags will associate with the Class I MHC and elicit a CTL response
• Nonspecific:
  
  • Adoptive Transfer of lymphocytes
    
    • Lymphokine-activated killer (LAK) cells are IL-2 activated T cells and NK cells can be used in the treatment of melanoma and renal cell carcinoma although studies of unselected LAK cells proved ineffective in the clinic.
    • Tumor-infiltrating lymphocytes (TIL) include T cells and NK cells. While the infiltrating NK cells will kill tumors nonspecifically, the CTL will be able to kill specific tumor targets. Unselected TILs have little utility in the clinic.
  • Cytokines
    
    • Interleukin-2 (IL-2): Activates T cells/NK cells which express IL-2 receptors and leads to their 
proliferation. Used in the treatment of renal cell carcinoma and melanoma and has proven clinical benefit.
    • Interferon-alfa (IFNα): Activates NK cell activity against tumors, but also has anti-viral and anti-proliferative capabilities and also used in the treatment 
of Kaposi sarcoma, renal cell carcinoma and melanomas.
    • IFN-γ: Ιncreases class II MHC expression; .
    • Tumor necrosis factor (TNF)-α: Kills tumor cells.
    • Granulocyte-macrophage colony stimulating factor (GM-CSF): Useful in overcoming 
neutropenia due to chemo- or radiation therapy
  • Cytokine gene transfected tumor cells may also be used which can activate T or LAK cells that can mediate anti-tumor immunity

Question 9

Describe the process of immune checkpoint blockade in treating cancer.

Answer 9

• This approach to immunotherapy is exemplified by antibodies directed against CTLA-4 (ipilimumab, tremilimumab), which block the immunosupression mediated by the interaction between B7 family members (on antigen-presenting cells) and CTLA-4 (on CD8+ and CD4+ T cells)
• Anti-CTLA4 therapy was the first agent to demonstrate a survival benefit in patients with advanced melanoma and was approved by the US Food and Drug Administration (FDA) in 2010 and now shows activity in renal cell cancer as well in combination with a PD-1 antibody (see below)
• A second major checkpoint, mediated by the interaction between PD-1 on T cells and its ligand PD-L1 on either antigen-presenting cells or tumour cells, has been the subject of several recent clinical trials, and has shown evidence of efficacy in both non-small-cell lung cancer and renal cell carcinoma and is now several are approved for the treatment of malignant melanoma
• Multiple additional immune-checkpoint receptors and ligands, some of which are selectively upregulated in various types of tumour cells, are prime targets for blockade, particularly in combination with approaches that enhance the activation of antitumour immune responses, such as vaccines
• Strategies to combine these drugs and identify predictive biomarkers are underway

Question 10

Describe CAR T-Cell Therapy.

Answer 10

• Although adoptive cell transfer has been restricted to small clinical trials so far, treatments using these engineered immune cells have generated some remarkable responses in patients with advanced cancer
• After collection, the T cells are genetically engineered to produce special receptors on their surface called chimeric antigen receptors (CARs)
• CARs are proteins that allow the T cells to recognize a specific antigen on tumor cells
• These engineered CAR T cells are then grown in the laboratory until they number in the billions
• The expanded population of CAR T cells are then infused into the patient
• After the infusion, if all goes as planned, the T cells multiply in the patient’s body and, with guidance from their engineered receptor, recognize and kill cancer cells that harbor the antigen on their surfaces
• CAR T-cell therapy eventually may “become a standard therapy for some B-cell malignancies” like ALL and chronic lymphocytic leukemia