week 7

PHASES OF ADAPTIVE IMMUNE RESPONSES

Adaptive immune responses consist of distinct phases, the first three being the recognition of antigen, the activation of lymphocytes, and the elimination of antigen (the effector phase). The response contracts (declines) as antigen-stimulated lymphocytes die by apoptosis, restoring homeostasis, and the antigen-specific cells that survive are responsible for memory. The duration of each phase may vary in different immune responses. The y-axis represents an arbitrary measure of the magnitude of the response. These principles apply to humoral immunity (mediated by B lymphocytes) and cell- mediated immunity (mediated by T lymphocytes).

MDSC use a variety of immune and nonimmune mechanisms to promote tumor progression, but have beneficial effects in other settings

In individuals with cancer, MDSC inhibit adaptive antitumor immunity by suppressing CD4 and CD8 T cell activation and function, and by driving and recruiting T regulatory cells. They inhibit innate immunity by polarizing macrophages toward a type 2 tumor-promoting phenotype and by inhibiting NK-mediated cytotoxicity. MDSC also promote cancer cell stemness, facilitate angiogenesis, and drive tumor invasion and metastasis. Beneficial effects of MDSC include their lowering of blood glucose levels and reduction of insulin tolerance in obese individuals, and their maintenance of maternal- fetal tolerance and embryo implantation during pregnancy.

ADOPTIVE CELL THERAPY

Lymphocytes isolated from the blood or tumor infiltrate of a patient may be expanded by culture in IL-2 and infused back into the patient (A) . The lymphocytes may be transfected with CAR genes (B) . This treatment, often combined with systemic IL- 2 administration, leads to tumor regression in some patients. In some cases, the patient's T cells may be genetically transduced ex vivo to express recombinant chimeric antigen receptors (CARs) before transfer back into the patient. CARs (B) are composed of receptor domains specific for tumor antigens, and signaling domains, such as ITAMs and cytosolic motifs of CD28, which promote robust T cell activation.

Exposed from the beginning

The newborn's gut microbiota trigger development and maturation of the newborn's immune system. Although there is still a great deal of research needed to understand precisely what happens in this developmental process, it appears the maturing immune system relies on the presence of microbial communities, and especially the presence of these early microbes, to distinguish "self" from "non-self." It is these particular microbes that shape our immune systems. Once the immune system has matured, it will consult its "memory banks" if another microbe is encountered in order to determine if this microbe is considered "self" or "non-self" and to mount defenses against the microbe if it is recognized as a pathogen

• Tumor-associated macrophages may promote tumor growth and invasiveness by altering the tissue microenvironment and by suppressing T cell responses.

These macrophages have an M2 phenotype, as discussed briefly earlier, and they secrete mediators, such as IL-10 and prostaglandin E 2 , that impair T cell activation and effector functions. Conversely, tumor-associated macrophages also secrete factors that promote angiogenesis, such as TGF-β and VEGF, which may enhance tumor growth.

T CELL INHIBITOR BLOCKADE

Tumor patients often mount ineffective T cell responses to their tumors because of the upregulation of inhibitory receptors such as PD-1 on the tumor-specific T cells and expression of the ligand PD-L1 on the tumor cells. Clinical trials using blocking anti-PD-1 or anti-PD-L1 antibodies have shown efficacy in treating several types of advanced tumors. A similar strategy using anti-CTLA-4 has been approved for treatment of melanomas, which may work by blocking CTLA-4 on effector T cells or Treg

Features that determine antitumor T cell reactivity

Two major types of Ags, tumor-associated and tumor-specific, can be recognized by endogenous T cell responses. The ability for epitopes derived from these Ags to be detected by responding T cells is modulated by the host's HLA type and the epitope' processing and presentation efficiency. Intratumoral heterogeneity may also allow individual tumor cells to escape recognition. On the T cell side, immunodominance hierarchies can be generated leading to an individual Ag being the major target of the response. Additionally, holes in the TCR repertoire and T cell tolerization and exhaustion can limit response efficacy.

Antibiotic Resistance

• Antibiotic resistance develops when bacteria are exposed to sub-lethal doses of an antibiotic that do not kill them but, instead, allow them to develop genetic resistance against the antibiotic. • Exposure to sub-lethal doses of antibiotics occurs when patients don't take the full regimen of prescribed antibiotics or when the bacteria are exposed to antibiotic doses that aren't high enough to cause complete bacterial mortality. • The development of antibiotic resistance means that microbes aren't eradicated when exposed to the same antibiotic at therapeutic doses during subsequent infections. • Moreover, antibiotics can have unintended consequences and kill off beneficial bacteria in our microbiomes that are not the original target of the antibiotic—so-called "non-target bacteria."

Good Bugs Gone Bad? Or, What Upset the Microbial "Apple Cart?"

• At the same time we are beginning to appreciate the microbiome, scientists are also growing concerned about things we are doing that may disturb this delicate system. • Antibiotic use is just one example of a common medical practice that may be altering the human microbiome by reducing, removing, or changing fundamental elements. • Antibiotics have been in broad use for treating infectious diseases in humans for over 70 years and are also used at sub-therapeutic levels to stimulate meat production in livestock. • As with vaccines, antibiotics have proven to be a very important medical advance, effectively eliminating many infectious diseases that have plagued human history. • Today, as a result of antibiotics and vaccines, children do not die of the infectious diseases that killed them even 50 years ago. • However, routine use of antibiotics may cause collateral damage to our microbial flora in two ways: through the unintended death of non-targeted bacteria and through the emergence of antibiotic-resistant bacteria

Tumours stimulate specific adaptive immune responses

• Clinical observations and animal experiments have established that although tumour cells are derived from host cells, the tumours elicit immune responses. • Histopathologic studies show that many tumours are surrounded by mononuclear cell infiltrates composed of T lymphocytes, natural killer (NK) cells, and macrophages, and that activated lymphocytes and macrophages are present in lymph nodes draining the sites of tumour growth. • The presence of lymphocytic infiltrates in some types of melanoma and carcinomas of the colon and breast is predictive of a better prognosis.

CD8 Cytotoxic T cells (ii)

• Furthermore, mononuclear cells derived from the inflammatory infiltrate in human solid tumors, called tumor-infiltrating lymphocytes (TILs), contain CTLs with the capacity to kill the tumor from which they were derived. • Importantly, the inability to detect tumor-specific CTLs in some patients may be because of regulatory mechanisms exploited by the tumor. • One of the most impressive results of recent clinical trials is that blocking these inhibitory pathways, and thus removing the brakes on immune responses, leads to the development of strong T cell responses against the tumor.

A special relationship

• If microbes are germs then how do we benefit from them? In fact, most of the microbes we come in contact with are not germs but beneficial microbes that • digest many things in our diet—like vegetables—that we could not digest without microbial enzymes, • provide energy for our metabolism • make essential vitamins • act as a first line of defense against potential pathogens (i.e., germs). Although we use the general term "microbe," which is often thought to be synonymous with bacteria, we now know the human microbiome is primarily composed of bacteria, but also includes numerous and diverse viruses, fungi and protozoa.

EVASION OF IMMUNE RESPONSES BY TUMORS

• Immune responses frequently fail to prevent the growth of tumors. There may be several reasons that anti-tumor immunity is unable to eradicate transformed cells. First, many tumors have specialized mechanisms for evading host immune responses. Second, tumor cells are derived from host cells and resemble normal cells in many respects. Therefore, many tumors tend to be weakly immunogenic. Tumors that elicit strong immune responses include those induced by oncogenic viruses, in which the viral proteins are foreign antigens. Many spontaneous tumors induce weak or even undetectable immunity. This may be because the tumors that grow have undergone mutations that reduce their ability to stimulate strong immune responses. Thus, the importance of immune surveillance and tumor immunity varies with the type of tumor. Third, the rapid growth and spread of a tumor may overwhelm the capacity of the immune system to effectively control the tumor, which requires that all the malignant cells be eliminated.

NK CELLS (ii)

• In addition, NK cells can be targeted to IgG antibody-coated tumor cells by Fc receptors (FcγRIII or CD16). The tumoricidal capacity of NK cells is increased by cytokines, including interferon-γ (IFN-γ), IL-15, and IL-12, and the anti-tumor effects of these cytokines are partly attributable to stimulation of NK cell activity. • IL-2-activated NK cells, called lymphokine-activated killer (LAK) cells, are derived by culture of peripheral blood cells or tumor-infiltrating lymphocytes from tumor patients with high doses of IL-2. These cells are more potent killers of tumors than are unactivated NK cells.

MACROPHAGES (ii)

• M1 macrophages can kill tumour cells by mechanisms that they also use to kill infectious organisms. Prominent among these is production of nitric oxide (NO), which has been shown to kill tumours in vitro and in mouse models in vivo. • There is evidence that some macrophages in tumours contribute to tumour progression and have an M2 phenotype. These cells secrete vascular endothelial growth factor (VEGF), transforming growth factor-β (TGF-β), and other soluble factors that promote tumor angiogenesis.

MACROPHAGES

• Macrophages are capable of both inhibiting and promoting the growth and spread of cancers, depending on their activation state. Classically activated M1 macrophages can kill many tumor cells. • How macrophages are activated by tumors is not known. Possible mechanisms include recognition of damage-associated molecular patterns from dying tumor cells by macrophage TLRs and other innate immune receptors, and activation of macrophages by IFN-γ produced by tumor-specific T cells.

MICROBIOME IMPACT

• Many doctors are learning that patients recover faster if their microbiomes are protected. • An excellent example of this comes from research involving the treatment of cancer patients. • Radiation and chemotherapy cause collateral damage because they kill healthy cells in addition to cancer cells; this is why these treatments are so difficult to endure, as they also kill the patients' microbiomes in the process. • As doctors have begun to realize this, some have started saving their patients' microbiomes, a process akin to a patient storing their blood in anticipation of upcoming surgery. • In the case of cancer patients, doctors collect and store the patients' gut microbiomes, again in the form of stool, prior to treatment. • Once the cancer treatments are stopped, doctors re-inoculate the patients with their own microbiomes via a procedure similar to the fecal transplants, as described above. • In this way, cancer patients receive a fresh inoculum of their own microbiota, allowing their microbiomes to quickly become re-established and shortening their recovery time.

WHERE?

• Most of these microbes are growing in our large intestine, but each region of our body has its own distinct community of microbes living in or on it. • We have a particular kind of microbial community that prefers to grow on our skin or in our nose. Our mouths have a rich mixture of microbes, with specific microbes that prefer our teeth versus those that prefer our gums. • Even though your tongue is in constant contact with the roof of your mouth, the microbes growing on the roof of your mouth are, in fact, very different from those growing on your tongue. • We're still trying to understand which factors regulate microbial colonization in areas of the body that are just millimeters apart.

NK CELLS

• NK cells kill many types of tumor cells, especially cells that have reduced class I MHC expression and express ligands for NK cell-activating receptors. In vitro, NK cells can kill virally infected cells and certain tumor cell lines, especially hematopoietic tumors. • NK cells also respond to the absence of class I MHC molecules because the recognition of class I MHC molecules delivers inhibitory signals to NK cells. Some tumors lose expression of class I MHC molecules, perhaps as a result of selection against class I MHC-expressing cells by CTLs. • This loss of class I MHC molecules makes the tumors particularly good targets for NK cells. Some tumors also express MIC-A, MIC-B, and ULB, which are ligands for the NKG2D activating receptor on NK cells.

• Secreted products of tumor cells may suppress anti-tumor immune responses. An example of an immunosuppressive tumor product is TGF-β, which is secreted in large quantities by many tumors and inhibits the proliferation and effector functions of lymphocytes and macrophages

• Regulatory T cells may suppress T cell responses to tumors. Evidence from mouse model systems and cancer patients indicates that the numbers of regulatory T cells are increased in tumor-bearing individuals, and these cells can be found in the cellular infiltrates in certain tumors. Depletion of regulatory T cells in tumor-bearing mice enhances anti-tumor immunity and reduces tumor growth.

New concepts arising

• The human body is made up of about 10 times more microbial cells (~1014) than human cells (~1013). Further, there may be millions more microbial genes than human genes in this human+microbiome system (which is often thought of as a human 'superorganism'), and it is the ways in which these microbial genes interact with the human host that describe their ultimate role in our health. • Scientists now believe that infants are sterile (meaning free of microbes) in the womb and receive their first inoculum of microbes from the mother during natural childbirth. This inoculum goes on to colonize the newborn and initiate a succession of events leading to the development of the child's own microbiome. • The newborn relies on this maternal vaginal microbial inoculum and the additional inoculum of microbes from mother's breast milk for microbial colonization of all exposed surfaces in and on the infant's body (e.g., oral, nasal/airways, gut, urogenital, skin). • This is a dynamic process in which microbial abundances increase from effectively zero at birth to over six orders of magnitude (that's more than a million times!) within just the first few weeks of life, with wide swings in the microbial membership of these communities until the microbiota largely stabilize in composition and numbers after approximately three years of life

IMMUNITY TO TUMORS

• The possibility that cancers can be eradicated by specific immune responses has been the impetus for a large body of work in the field of tumor immunology. • The concept of immune surveillance of cancer, which was proposed by Macfarlane Burnet in the 1950s, states that a physiologic function of the immune system is to recognize and destroy clones of transformed cells before they grow into tumors and to kill tumors after they are formed.

CD8 Cytotoxic T Cells

• The principal mechanism of adaptive immune protection against tumors is killing of tumor cells by CD8 + CTLs. The ability of CTLs to provide effective anti-tumor immunity in vivo is clearly seen in animal experiments using carcinogen-induced and DNA virus-induced tumors. • CTLs may perform a surveillance function by recognizing and killing potentially malignant cells that express peptides that are derived from tumor antigens and are presented in association with class I MHC molecules. • Tumor-specific CTLs can be isolated from animals and humans with established tumors, and there is evidence that the prognosis of human tumors, including common tumors such as colonic carcinomas, is more favorable when more CTLs are present within the tumor.

Antibiotic Resistance (ii)

• There is thought to be a relationship between the theorized disturbance of the human microbiome through antibiotic use and the unexpected rise in autoimmune diseases and allergies, particularly in Western countries. • Autoimmunity is the failure of our own immune systems to distinguish "self" from "non self." • This failure can lead to an immune response being mounted against our own cells and tissues. Examples of autoimmune diseases include rheumatoid arthritis, lupus, diabetes and celiac disease. • The current line of thinking is that loss of normal microbiome constituents through antibiotic use may remove the necessary trigger for normal immune system development. • As a result, an underdeveloped immune system might possibly allow autoimmune diseases to develop. • Currently, much research is being conducted to better understand the relationship between the human microbiome and autoimmune diseases, and to find better treatments and cures

ACTIVE INHIBITION OF IMMUNE RESPONSES

• Tumors may engage inhibitory mechanisms that suppress immune responses. There is strong experimental and clinical evidence that T cell responses to some tumors are inhibited by the involvement of CTLA-4 or PD-1, two of the best-defined inhibitory pathways in T cells. • A possible reason for this role of CTLA-4 is that tumor antigens are presented by APCs in the absence of strong innate immunity and thus with low levels of B7 costimulators. These low levels may be enough to engage the high-affinity receptor CTLA-4. • PD-L1, a B7 family protein that is a ligand for the T cell inhibitory receptor PD-1 is expressed on many human tumors, and animal studies indicate that anti-tumor T cell responses are compromised by PD-L1 expression. PD-L1 on APCs may also be involved in inhibiting the activation of tumor-specific T cells. • As we will discuss later, blockade of the CTLA-4 and PD-L1/PD-1 pathways is now being used in the clinic to enhance tumor immunity.

WHERE? (ii)

• What we eat, combined with our hormones, bodily fluids, skin oils, genetic makeup, where we live, and many other factors, contribute to the colonization and growth of these microbes. • Bathing, washing your hair, washing your hands, and brushing your teeth remove some microbes, but they eventually grow back. And it's thought that each of us has our own personal group of microbial species and strains (meaning microbial subspecies) that make our bodies their only homes. • In other words, each of us supports a unique group of microbes that are ours and ours alone. How do you feel about having your own "personal" microbes?

CAR T cells

• the first chimeric antigen receptor T cell (CAR-T) therapy, for the treatment of patients up to 25 years of age with B-cell precursor acute lymphoblastic leukemia (ALL) that is refractory or in second or later relapse. Kymriah is a novel immunocellular therapy and a one- time treatment that uses a patient's own T cells to fight cancer. Kymriah is the first therapy based on gene transfer approved by the FDA.