6.0 SPECIFIC IMMUNOTHERAPY II: VACCINATION
The aim of cancer vaccination is to place an antigen within the body of a cancer patient so that the immune system can be provoked to unleash the wrath of the killer T cells on the patient’s tumor cells. Armed with the knowledge of how T cells interact with antigens at the atomic level, scientists are able to design antigens that can selectively activate specific T cells to eradicate cancer cells. In general, the success of vaccine strategies depends on the mode of antigen delivery, the choice of adjuvant, and the particular antigen being used. For the rest of this section, let us examine a few examples of anticancer vaccination therapies including tumor-based, virus-based, peptide-based, and professional APC-based vaccinations.
6.1 Specific Immunotherapy II: Tumor-based vaccines
A relatively ancient but still useful form of anticancer vaccination strategy is to extract whole tumor cells, mash them up, and inject the crude extract back into the patient. These days, this method has been refined somewhat: whole tumor cells are extracted from the patient, blasted with radiation to weaken them substantially before they are transferred into the patient in the presence of an adjuvant such as BCG. Although the exact mechanism of action remains unknown, scientists think that the BCG supplement creates a suitable environment within the patient so that the antigens associated with the tumor cells can be properly presented to the immune system for the subsequent generation of T cells that can seek and destroy tumor cells. In a study conducted in 1999, patients with stage II cancer of the colon who had undergone surgery to remove part of the tumor were given a tumor-BCG mixture to fight against the reappearance of the tumor and it was observed that this approach reduced the risk of recurrence by about 61 percent. Other scientists have shown via studies in mice that the use of cytokines, such as GM-CSF, or co-stimulatory molecules, such as B7, can dramatically improve the efficacy of tumor-based vaccinations. Adjuvants, cytokines and co-stimulators are all believed to improve this anticancer therapy by creating an optimal in vivo environment for the presentation of tumor-associated antigens to the immune system. The main advantage of this method is that scientists do not have to isolate a specific antigen: the use of a crude cancer cell extract is good enough. A disadvantage is that it is difficult to measure specific immune responses without knowing the stimulating antigen within the crude mixture. This limitation, therefore, makes it more difficult for researchers to learn from these tests if the vaccine fails to generate a tumor regression. Also, isolating and persuading cancer cells to grow in a test-tube is a tedious and costly job whose application is limited to the single patient from which the tumor cells were originally isolated. As we shall see in section 6.3, modern techniques have rendered the use of crude cell extracts almost obsolete in the fight against cancer.
6.2 Specific Immunotherapy II: Virus-based vaccines
In 1910, physicians observed that the tumor cells of a woman suffering from a cancer of the cervix went into remission while she was receiving a rabies vaccine. This has become one of those serendipitous discoveries in medical science that provided the impetus for a form of anticancer strategy called in vivo viral oncosylate vaccination that involves the direct injection of viruses into tumor sites. Scientists rationalized the success of this technique as follows: the viral proteins are foreign to the body and as such, they are highly immunogenic whereas the tumor proteins that arose from the body’s own cells are weakly immunogenic. The association of the two types of protein makes the tumor proteins immunogenic enough for them to elicit a tumor-specific immune response. Although earlier clinical trials of this technique were encouraging, the results were deemed too inconsistent. This led to a change in the strategy whereby the tumor cells were infected with the virus in vitro. After the virus had successfully infected and broken up the tumor cells, scientists isolate parts of the mixture that lack nuclei and use that to vaccinate cancer patients. This approach has led to results that are more consistent and its efficacy has been demonstrated in patients suffering from melanoma, colon and ovarian cancers.
These days, scientists are attempting to further improve this technique with the help of genetic engineering techniques. This involves the isolation of the human genes that code for tumor antigens and then genetically engineering them into viral vectors. The latter is a gene courier that delivers the gene to a particular address within the body. Infecting a patient with such an engineered virus will hopefully, lead to an immune response against both the virus and the tumor antigen. In one example of this approach, scientists equipped the vaccinia virus with the gene of a human cancer antigen called carcinoembryonic antigen (CEA). After injecting this into patients with CEA-expressing tumors, the scientists observed that an effective immune response was generated and this led to the production of CEA-specific T cells that fought off the CEA-expressing tumors.
6.3 Specific Immunotherapy II: Protein and Peptide-based vaccines
If we cast our minds back to how the immune system works (chapter 2), we will recall that during the activator phase of both the humoral and cellular responses, antigens need to be processed into peptides before they are presented to the immune system as MHC-peptide complexes. Recently, one of the most remarkable advances in immunology has been the understanding at the atomic level of how T cells actually recognize and dock onto processed peptides sitting in the groove of MHC molecules. Armed with this knowledge, scientists think that for a particular tumor antigen, they can home in on just the specific region called the epitope that is presented via the MHC molecule. By supplementing this epitope with the appropriate co-stimulatory molecule(s), they can activate tumor-specific T cells into action. This technique is essentially peptide-based vaccination.
A cancer antigen can be defined as an antigen that is selectively or abundantly expressed in cancer cells. Human cancer antigens that are recognized by our T cells include cancer testis (CT) antigens such as MAGE-3, BAGE, GAGE, and NY-ESO-1; melanocyte differentiation antigens such as Melan-A/MART-1, tyrosinase, and gp100; protein products of point mutant genes like beta-catenin, MUM-1, CDK4, p53, and ras; overexpressed “self” antigens such as Her-2/neu, p53, and MUC-1; and viral antigens such as the Epstein-Barr virus (EBV), hepatitis B virus (HBV), hepatitis C virus (HCV), and the human papilloma virus (HPV). Researchers have amassed a wealth of evidence that shows that some human tumors tend to overproduce certain proteins in either their normal or mutated forms. Of all the examples mentioned above, NY-ESO-1 represents one of the most potent naturally occurring cancer antigens. With the exception of the testis, this protein is completely absent from normal tissues hence its categorization as a CT antigen. NY-ESO-1 is found in about 30 percent of breast, prostate and ovarian cancers as well as melanoma. These desirable features—that is, rarity in normal tissues, high immunogenecity, and significant presence in a relatively broad range of cancers—have made NY-ESO-1 a highly attractive target for specific immunotherapy in certain cancer patients. The identification of antigens such as NY-ESO-1 that are selectively or abundantly expressed in cancer cells has set the basis for the design of a large number of cancer vaccine trials around the world. The CRI and the Ludwig Institute for Cancer Research (LICR) have created a new partnership with the objective of developing a cancer vaccine collaborative (CVC) program involving a number of internationally recognized medical centers. Specific information on this program and participating institutions can be obtained from the website of the CRI.
The main advantage of a peptide-based vaccine is that it provides a method for monitoring a specific immune response for a particular antigen. This affords researchers the opportunity to obtain important information for evaluating the efficacy of other tumor antigens. Other advantages are as follows: first, it bypasses the need for antigen-presenting cells to process a whole cell before presenting the antigen to the immune system. Second, administration of a peptide antigen does not carry the risk of introducing dangerous substances into the patient—unlike other vaccines that rely on weakened pathogens or crushed tumor cells. The potential of peptide vaccination as an anticancer therapy will be brighter when current loose ends (such as peptide dose, adjuvant, cytokine combination, method of delivery, optimized peptide sequences and maybe the synergistic use of MHC class I and Class II peptide combinations) are tied up by scientists.
6.4 Specific Immunotherapy II: Antigen-enhanced, APC-based vaccines
As mentioned in chapter 2, section 2.5 of this book, T cells only recognize antigens that have been properly processed and presented on the surface of the APC bound to a protein called MHC. Recently, scientists have demonstrated that they can exploit the unique skills of APCs for vaccination against cancer. This can be achieved by isolating a tumor antigen from a cancer patient and then loading or pulsing APCs (also isolated from the patient) with the tumor antigen ex vivo (outside the patient). The transfer of these pulsed APCs into the cancer patient elicits a significant tumor-specific immune response that attacks the tumor cells. In essence, the tumor antigen is hitching a ride into the patient’s immune system from the APCs. Currently, there are three different methods for letting tumor antigens piggyback on APCs into the human immune system. First, growing APCs in the presence of a tumor-associated protein; second, using genetic engineering techniques to introduce the gene that codes for a tumor-associated protein into APCs, and third, pulsing APCs with fragments (peptides) isolated from a tumor antigen.
The main advantage of APC-based vaccination is that DCs produce all the molecules required for eliciting an immune response, unlike other forms of cancer immunotherapy where adjuvants and co-stimulatory molecules are required to boost the ensuing immune response. The potency of DCs as vehicles for delivering antigen and achieving a tumor-specific immune response has been demonstrated in a number of clinical trials around the world. For example, in a recent clinical study, involving 16 patients, DCs pulsed with tumor-associated peptide or lysate were shown to be effective in treating metastatic melanoma among five of the patients. However, a recent study has reminded us of the need for caution in utilizing DCs for clinical trials.
A group of Rockefeller University scientists, including CRI-funded researchers Drs. Madhav Dhodapkar, Ralph Steinman, and Nina Bhardwaj has undertaken a clinical study testing the capacity of mature versus immature dendritic cells (DCs) to stimulate immune responses. Immature and mature dendritic cells simply refers to a state of development just like a teenager (with certain unique traits) on the way to becoming an adult with a different set of unique characteristics. As described in a report published in the January 14, 2001, issue of the Journal of Experimental Medicine, this group found that in contrast to prior findings using mature DCs, injection of immature DCs into healthy subjects led to the specific inhibition of antigen-specific T-cell function. They found that immature DCs were not simply weaker adjuvants, but led instead to silencing of preexisting immune effectors. When the T cells elicited by immature DCs were boosted in culture, they were dysfunctional as they lacked the ability to kill target cells and exhibited a reduced level of interferon-gamma production. Additionally, the use of immature DCs stimulated a population of antigen-specific regulatory T cells that produced a cytokine called IL-10, which suppresses the immune system. While DCs are currently under active investigation, mostly for their immunostimulatory properties in cancer and viral infection, this study suggests that caution must be utilized in the use of immature DCs when trying to enhance tumor and microbial immunity. The suppressive properties of immature DCs observed in this report suggest that these DCs may also be valuable for antigen-specific inhibition of T-cell function in the setting of autoimmune diseases and organ transplantation in humans.
6.5 The future of cancer vaccination
As we saw in chapter 2 of this booklet, the immune response can be broadly divided into a cellular and a humoral response. The latter relies on T cells whereas the former involves antibodies. Currently, the vast majority of cancer vaccines are directed at achieving a cellular immune response. Scientists are now working on anticancer strategies that will result in a humoral immune response because recent studies have demonstrated that the presence of tumor-specific antibodies can lead to tumor regression in some patients. Some of the potential antigens that researchers have their eyes on include molecules like p53 and gangliosides. The gangliosides are carbohydrate molecules that are normally present on cell membranes. In one clinical study, patients with metastatic melanoma who were vaccinated with a ganglioside called GM2 followed by BCG treatment showed a 14 percent increase in overall survival rate compared to those who received BCG alone. None of the patients had antibodies to GM2 before the trial began. Scientists are conducting trials in which they are experimenting with other adjuvants such as keyhole limpet haemocyanin. Also, the search is on for alternative ganglioside targets such as GD2 and GD3.
Although cancer vaccines that activate the humoral immune response remain a vital strategy in the toolkit of cancer immunotherapists, the current method of choice for practitioners involves the activation of the cellular immune response. Some of the strategies that researchers are using to achieve this effect involve vaccines based on genetically engineered bacteria and DNA. Scientists have known for a while that bacteria such as salmonella, BCG, and listeria are very good at infecting professional APCs. We can turn this to our advantage by genetically altering these bacteria so that they can carry tumor antigens directly to the sites within a patient where an optimal immune response can be guaranteed.
A new and exciting anticancer vaccination strategy involves DNA vaccines. The efficacy of this approach was first demonstrated in animal studies where the administration of DNA effectively protected the mice against the influenza virus. This approach involves using genetic engineering techniques to put the DNA for a tumor antigen into a plasmid (see chapter 8). By disabling the ability of the plasmid to replicate, it can be safely injected into animals and it has been demonstrated that this results in the expression of the plasmid-encoded tumor antigens. This means that physicians will have the ability to direct the DNA vaccines to the exact location that they think will lead to the best immune response within the patient. DNA vaccination against cancer is a very promising field in immunotherapy because it is relatively safe, practical, and affordable.
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