Dermatology Online Journal

Melanoma vaccines
Mark F. Naylor, MD¹
Dermatology Online Journal 6(1): 5

1. Department of Dermatology, University of Oklahoma Health Sciences Center

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Abstract

Melanoma vaccines are the best near term hope for improving mortality in patients with advanced disease. Although these vaccines are being developed for treatment of patients with advanced disease, their relatively low toxicity make them attractive for adjuvant therapy in stage I patients at high risk for recurrence. Recent developments in melanoma vaccines are reviewed and their importance to dermatologists is emphasized. (transaction of the RxDerm-L Meeting, November 1999, San Antonio)

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Introduction

Development of melanoma vaccines is occurring at an accelerated pace, a topic of interest to dermatologists for a number of reasons. Managing advanced melanoma usually involves more than one physician. Dermatologists are frequently the first or second physician consulted, and we should be adept at initiating and orchestrating the further management of patients with advanced disease even if we are not going to be the primary care givers. When these patients ask for advice about their treatment options, we need to be prepared to discuss these issues knowledgeably. Melanoma is and should be our disease. The fact that other physicians such as surgical specialists, oncologists and radiologists are needed to properly manage patients with advanced disease does not divorce us from our responsibility to help in any way we can. At a minimum, these patients should be monitored by a dermatologist for the occurrence of new primaries. Primary prevention efforts should also be made to reduce new occurrences in the patient and their immediate family. Melanoma vaccines will enhance our ability to intervene in the future and we should be familiar with developments in this area.

Melanoma vaccines as they are currently used are not intended to prevent melanoma; rather they are used to boost immune responses to preexisting tumors. Unlike traditional chemotherapy or biological response modifiers, melanoma vaccines have relatively low toxicity and potentially, a high degree of efficacy. Because of this, they are likely to be adopted as adjuvant therapies for stage I patients at high risk for recurrence. This makes dermatologists the most logical physicians to administer vaccines, since we are diagnosing and treating these patients most often.

Immunotherapy includes techniques to boost natural Immune resistance to tumors with both vaccines and biologic response modifiers, primarily cytokines involved in modulating immune responses. The occasional dramatic disappearance of widespread metastases either spontaneously or more commonly with a palliative course of dacarbazine is probably attributable in most cases to a brisk and successful immunologic response. The occasional patient who survives in the face of known metastatic disease for a decade or more is probably also a testament to an effective immunologic host response. These occasional examples demonstrate the power of the immune system to arrest or even cure what appears to be a hopeless case. Clearly, we would like to be able to achieve this result deliberately in all melanoma victims. The key to achieving predictable and significant immune responses in cancer patients lies in a better understanding of the nature of tumor immunity.

Tumor Immunity

Generally speaking there are two broad types of anti-tumor immune responses. One involves the humoral arm of the immune system and the other involves the cellular arm of the immune system. An important aspect of either is the ability of antigen presenting cells to process and present tumor-related peptide antigens that are the primary basis for immune recognition of tumor cells. Tumor antigens that have been phagocytosed and partially digested by antigen presenting cells are presented as peptides bound to MHC type II receptors on the surface of antigen presenting cells (

Figure 1). Examples of such antigen-presenting cells include macrophages, epidermal Langerhans cells, other types of dendritic cells and B-cells. The MHC class I cell surface receptors that are the basis for HLA tissue typing are present on all nucleated cells in the body including tumor cells. These receptors semi-randomly present examples of peptides present within the cell. MHC class I receptors also present tumor-specific peptide antigens on the tumor cell surface, giving the opportunity for the properly sensitized immune system to react to the tumor.

The Antibody-Mediated Arm of Tumor Immunity

Antibody-dependent mechanisms of tumor immunity include antibody dependent cell-mediated cytotoxicity (ADCC), complement dependent cytotoxicity (CDC) and opsonization. These mechanisms depend on the ability of the immune system to create antibodies to tumor cell surface antigens that in this case do not have to be presented on class I MHC receptors as with the T-cell-mediated responses to be discussed later.

Antibody-Dependent Cell Medicate Cytotoxicity (ADCC)

ADCC involves the attachment of tumor-specific antibodies to tumor cells and the subsequent destruction of the tumor cell by immunocompetent cells. Fc receptors on immunocompetent cells recognize the Fc portion of antibodies adhering to surface tumor antigens (

Figure 2). Most commonly the effector cell of ADCC is a natural killer (NK) cell. Following recognition and attachment via its Fc receptors, the NK cell can destroy the target tumor cell through release of granules containing perforin and granzymin B and/or activation of the FAS/FAS ligand apoptosis system in the target cell. Perforin molecules make holes or pores in the cell membrane, disrupting the osmotic barrier and killing the cell via osmotic lysis.

Complement-dependent cell-mediated cytotoxicity (CDC)

Complement-dependent cell-mediated cytotoxicity involves the recognition and attachment of complement-fixing antibodies to tumor specific surface antigens followed by complement activation (

Figure 3). Sequential activation of the components of the complement system ultimately lead to the formation of the membrane attack complex (MAC) which forms transmembrane pores that disrupt the osmotic barrier of the membrane and lead to osmotic lysis. The MACs function similarly to the perforin molecules released by cytolytic T cells and NK cells, killing cells by osmotic lysis.

Opsonization

Opsonization is the process in which tumor-specific antibodies attach to their target antigens on tumor cell surfaces, thus marking them for engulfment by macrophages (

Figure 4). This can also lead to processing and presentation of new tumor-specific antigens by the macrophage in addition to direct destruction of the tumor cells.

The Cell-Medicated Arm of Tumor Immunity

Cell-mediated tumor defenses include cytolytic T-lymphocytes, NK cells and macrophages. Cytolytic (CD8 positive) T-cells destroy tumor cells via T-cell receptor recognition of tumor-specific antigen presented on MHC type I receptors at the tumor cell surface. Tumor antigen-specific T-cells bind to the MHC I receptor-tumor antigen complex and destroy the tumor cell via the release of granules containing granzyme B, perforin and via induction of FAS pathway apoptosis (

Figure 5).

In addition to being the principal effector cell for ADCC, NK cells participate in tumor immune responses in another way. All nucleated cells express MHC I receptors on their surface (

Figure 6). The primary purpose of the MHC I receptor is to present endogenous peptide (self) antigens on a cell's surface. When a foreign peptide antigen such as a tumor-specific antigen is presented by the MHC I surface complex, cytolytic T-cells reactive to the antigen can recognize the foreign peptide with their T-cell receptor and kill the cell. Another function of MHC I receptors is to inhibit the innate tendency of NK cells to bind to and kill cells. Tumors may attempt to avoid immunosurveillance by downregulating expression of MHC I receptors, thus avoiding T-cell recognition. However, because MCH I expression is necessary to inhibit natural killer function, tumor cells that downregulate expression of MHC I receptors target themselves for natural killer attack.

Phases of Clinical Trials

The FDA divides clinical trials into 3 and sometimes 4 types or phases; studies in each phase have a particular intent.. Knowing the phase of the trial therefore gives you clues as to the primary intent of the study, which should be taken into consideration when recommending patients for clinical trials. Phase I trials are small scale trials of a few to a few dozen patients that are primarily dose-ranging studies intended to reveal any toxicity of the treatment under study. Efficacy is a secondary issue in Phase I trials, although this information will typically be sought in addition to toxicity and side effect data. Although in most cases animal studies of safety will have preceded the phase I trial, this is typically the first time that the drug or device has been used in humans and is therefore still highly experimental.

Phase II clinical trials are usually dose ranging studies in a larger number of patients (a few dozen or more) and are usually intended to identify the optimum dose of a drug. Toxicity and efficacy issues are of approximately equal importance in Phase II trials. If a treatment still looks promising after Phase II trials, a larger scale Phase III will typically be conducted next.

Phase III clinical trials are much larger scale trials of many dozens or hundreds of patients intended primarily to demonstrate the efficacy of the drug or device in a significant number of subjects. Efficacy, safety and toxicity are all of approximately equal importance in Phase III trials, although the safety of the drug projected into the wider public will be a very important concern of these studies. This is usually the last chance for the FDA to stop a drug that has unacceptable toxicity before it is introduced into the American market.

Phase IV studies have been used to describe post-marketing efforts to gather safety data or other information in much larger numbers of people once a drug has been released into the market. Phase II or III studies tend to have less risk to volunteers in terms of treatment toxicity. However, the fact that melanoma is a fatal disease with no clearly effective treatment of choice means that a highly experimental treatment in a Phase I study may offer the best hope in an individual patient.

Melanoma Vaccines

The melanoma vaccine types that will be discussed here are multivalent cell culture-derived vaccines, autologous melanoma cell vaccines, peptide, DNA and dendritic cell vaccines. These are the major types of vaccines currently in use or under study today.

Multivalent Cell-Culture Derived Vaccines<</H4>

Multivalent cell-culture derived vaccines are created by processing a number of different melanoma cell lines grown in vitro. These are non-autologous cell vaccines and the use of multiple cell lines helps ensure that at least some of the antigens in the vaccines are shared by the patient's own tumor (

Figure 7).

The advantage of these vaccines is that they do not require harvesting and processing autologous melanoma cells from the patient. Due to the use of a number of different cell lines, they are likely to have at least some antigens relevant to the patient's tumor. A disadvantage is that these vaccines are inefficient, since most of the immunologic response is probably wasted on irrelevant antigens that are not present in the patient's tumor.

Most clinical trials of melanoma vaccines in the last two decades used multivalent melanoma cell vaccines. These vaccines have not had a major impact overall on long-term survival in advanced cases of melanoma, although important responses have been seen in individual cases [

1]. Better use of adjuvants or concomitant administration of biologic response modifiers may improve the responses seen with these vaccines, although the issues of expense, lot to lot quality control problems and efficacy may limit the future of vaccines of this type.

Autologous Cell Vaccines

Autologous cell vaccines are prepared by harvesting melanoma cells from the individual patient to be treated (

Figure 8). Autologous cell vaccines come in at least two general variants, killed cell vaccines and recombinant autologous cell vaccines.

Live or Killed Cell Vaccines

Killed cell vaccines require less in vitro manipulation than recombinant autologous cell vaccines. Melanoma tissue is harvested from the patient, processed, and then re-injected with some type of adjuvant. The cells can be living but lethally irradiated so they cannot propagate after reinjection. An advantage of living, lethally irradiated cell vaccines is that they are highly analogous to the unaltered tumor cells remaining in the patient. Immunologic responses that prove lethal to the vaccinated cells are more likely to be lethal for the remaining wild type tumor cells.

A good example of a killed cell vaccine is the AVAX vaccine in which the patient's cells are processed and dinitrophenylated. This haptenizes tumor proteins and helps stimulate immune responses in a manner analogous to topical immunotherapy for warts [

2]. Since this is a product of a commercial enterprise, it may eventually be available in more locations than many vaccines offered through one or two academic centers (http://www.avax-tech.com/).

Recombinant DNA Cell Vaccines

Recombinant DNA techniques can be used to alter autologous melanoma cell vaccines in ways that boost immune responses (

Figure 9). For example, IL-2, GM-CSF, or other cytokines can be transfected into melanoma cells to make them ectopic producers of these potent immunomodulating substances. The combination of tumor-specific antigens presented in the form of living but lethally irradiated tumor cells in the presence of high local concentrations of immune enhancing biologic response modifiers can substantially boost the immune response from immunocompetent cells infiltrating the vaccination site.

The advantage of autologous cells is that since the vaccine is derived from the patient's own tumor, most of the antigens presented will be relevant. A principal disadvantage of the autologous cell approach is the difficulty of obtaining viable malignant cells in sufficient numbers. This is usually possible only when the tumor is quite advanced and the patient near death. The expense of harvesting and processing the cells is considerable, and recombinant manipulation requires advanced laboratory facilities geared to produce such vaccines.

Phase I trial results of engineered autologous tumor cells administered to humans have been published [

3, 4]. Although newly positive ANAs were reported in some of these patients, no frank lupus has been reported so far as a result of such vaccinations. Other than this, toxicity has been reasonably mild, limited to vitiligo, pain, itching at injection sites, fever, and malaise. Although these early results do not show dramatic improvement in every patient, they do show that this approach can be safe and effective and should encourage further work.

Peptide Vaccines

Peptide vaccines are based on peptide epitopes (

Figure 10). The peptide response epitopes used in melanoma vaccines are amino acid subsequences from tumor-derived proteins that have been the target of successful immune responses in other patients. These epitopes have been determined through tedious efforts to sequence peptides displayed on MHC I receptors mediating responses in tumor infiltrating lymphocytes (see Figure 5) for a schematic of a peptide response epitope presented on the MHC I receptors of tumor cells). Peptide response epitopes are potentially a more efficient means of immunization, since only the most immunogenic portions of tumor marker proteins are used for immunization. A comprehensive list of such peptide response epitopes presumably would represent the most immunogenic targets for melanoma vaccines.

The advantage of vaccines based on peptide response epitopes is that they lend themselves to mass production. Peptide vaccines can be made commercially at reasonable cost, and have minimal technical requirements for administration. Thus, multivalent vaccines of this type potentially can reach a much greater number of melanoma victims with a better response than might be anticipated from cellular or crude protein vaccines.

The disadvantages of peptide vaccines are several-fold. A multivalent peptide vaccine that includes the best antigenic peptides to the most common tumor targets is highly desirable but currently unavailable. At present relatively few epitope targets are known, limiting the value of contemporary peptide vaccines.

Another major drawback of peptide vaccines is a problem called HLA restriction (

Figure 11). This derives from the fact that MHC I receptors, the cellular determinants of HLA tissue types, can only bind to peptides within certain size, shape and charge limits. Different HLA (MHC I) receptors have different size, shape and charge to their antigen binding pockets, meaning that each HLA receptor has a limited repertoire of peptides that it can present at the cell's surface. Thus if a vaccine peptide cannot bind to the antigen binding pocket of a given MCH I receptor, using a vaccine based on this peptide will be futile, at least in theory. This means that peptide epitopes are HLA tissue type-specific since different MCH I receptors will have different optimum peptide response epitopes that they characteristically bind to. Thus the best use of peptide vaccines will probably require knowledge of the HLA tissue type of the vaccine recipient to determine the peptide epitopes compatible with the MHC I receptors on the patient's tumor cells.

"Naked" DNA Vaccines

Naked DNA vaccines are one of the newest category of melanoma vaccines (

Figure 12). These differ from recombinant DNA vaccines in that the transfection of exogenous DNA occurs in vivo rather than in vitro, and no autologous tumor cells are required. The biologic response modifiers or other proteins to be expressed are inserted in a plasmid vector capable of expressing exogenous proteins in mammalian cells. These plasmids are introduced into a skeletal muscle bed or subcutaneous tissue with the intent of transfecting local host cells with the recombinant plasmid, thus creating a local deposit of ectopically expressed protein. The adjuvants introduced with the plasmid will then help stimulate an immune response to the components of the vaccine. The protein coded in the plasmid could be an antigenic response epitope or a series of them. Alternatively, this could be a way to administer sustained release of the ectopically expressed biologic response modifier known to be useful in stimulating innate immune responses to the tumor, such as IL-2. In the latter role they could be used as an adjunct to other vaccines or treatment strategies. Conceivably they can be used for both, either in different plasmids injected into different sites, in different plasmids co-injected into the same sites, or even in one plasmid encoding one or more proteins encoded from the same plasmid.

A major advantage of this type of vaccine is that it does not require autologous tumor cells or individualized manipulations of DNA. As such they would also be amenable to mass production with minimal technical requirements for administration.

Although they have great potential, DNA vaccines are the most experimental vaccine strategy of those discussed here. A major potential drawback of this approach that has been encountered in other gene therapy studies using exogenous DNA vectors of various types is the difficulty maintaining tissue expression levels of the plasmid-encoded protein(s) over time. This may or may not be a significant problem for antigen (peptide) delivery, although it may be for the continuous delivery of ectopic cytokines and biologic response modifiers.

Dendritic Cell Vaccines

Another new approach to melanoma vaccines that has a great deal of promise is the dendritic cell vaccine (

Figure 13). This approach has been made feasible by the observation that the peripheral blood lymphocyte fraction can be stimulated in vitro to obtain functional antigen presenting cells. Thus, peripheral blood can serve as the source of antigen presenting cells that can be primed in vitro with a relevant peptide antigen and then re-injected into the patient.

The advantage of this approach is that, while it does require autologous cells, no tumor tissue is required, and peripheral blood is the only autologous tissue involved. Because the peptide antigen is incubated with the most important cells for initiating an immunologic response, the hope is that this approach will produce optimum immunologic responses to relevant antigens.

The disadvantage of this approach is that it is highly experimental and suffers from the same drawbacks as peptide vaccines, since peptide response epitopes are most commonly the tumor antigen that is incubated with the dendritic cells. This approach also requires a sophisticated immunology laboratory equipped to properly process and handle the dendritic cells.

Biologic Response Modifiers

Biologic response modifiers are frequently involved with vaccine technology, since these signaling molecules are usually mediators of immune responses. Biologic response modifiers are often used as an integral part of the primary vaccine strategy or as important vaccine adjuvants. Some of the ones commonly associated with vaccine therapy are covered here.

Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF)

Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) induces dendritic cells to develop into mature antigen-presenting cells. Thus, it significantly enhances presentation of antigens that are present either endogenously or as part of the vaccine strategy. Early studies indicate that this may be one of the most potent adjuvant agents yet discovered. Thus, GM-CSF is commonly encountered as a primary strategy (e.g. as an ectopically expressed protein in recombinant vaccines) or as a pharmacological adjuvant that is injected with other types of vaccines.

Interleukins

Interleukins are the generic name given to the intracellular signaling molecules that lymphocytes use to communicate with each other. As such they are important mediators of immunologic responses and can be used to enhance responses to vaccine antigens.

IL-2 is one of the most important early signaling molecules in an immune response. IL-2 has been used to stimulate peripheral blood lymphocytes in vitro (LAK therapy) and to stimulate tumor infiltrating lymphocytes in vitro (TIL therapy) with varying success. IL-2 is currently one of the few biologic response modifiers approved by the FDA for exogenous administration as an antitumor agent. IL-2 has also been used as an ectopically expressed protein in recombinant DNA cell vaccines so that the immunizing tumor cells are locally stimulating immune responses.

IL-12 is involved in stimulating the differentiation of helper lymphocytes into Th1 type cells, which are believed to be more important in cell mediated defense against tumors than are Th2 type helper lymphocytes, which represent the other major differentiation pathway for helper cells.

IFN-g

IFN-g is the interferon that is most often used in conjunction with vaccine therapy. Major roles of IFN-g are to activate macrophages and stimulate antibody production by B-cells. IFN-g has been used as an independent therapy for tumors and as an adjuvant.

Finding Melanoma Vaccine Trials

There are over 100 trials sponsored or sanctioned by the

National Cancer Institute (NCI) for the treatment of melanoma by means of vaccines or biologic response modifiers or both. The NCI website is the best source to find these trials. Using the keywords "melanoma" and "vaccine", melanoma vaccine trials can be identified nationally or regionally.

There are also commercially sponsored vaccine trials that can be found on the Internet.

Genzyme Corporation is sponsoring a phase I peptide vaccine trial. Progenics Corporation has a phase III trial of the ganglioside antigen GM2 . Avax Technologies has a phase III trial for an autologous cell vaccine utilizing a hapten adjuvant . Regional availability of these trials will vary, so you will have to check the web site to see if a participating center is in your area.

Other sources for vaccine trials include clinical trial organizations such as the

Eastern Cooperative Oncology Group (ECOG, ) and the Southwestern Oncology Group (SWOG). A particularly good web resource (although oriented primarily for melanoma patients) is the Melanoma Patient's Information Page. This resource has information about staging and other general educational materials for melanoma patients in addition to information about available clinical trials.

Your local oncologist is also potentially a good source of information on the trials available in your area. However, you should not let your search for clinical trials start and stop with this single resource, since it is difficult for one physician to keep track of all the clinical trials for which melanoma patients are potentially eligible. The best information about trials available for an individual patient is usually obtained by concerted effort on the part of a knowledgeable and interested physician.

Currently, most of the melanoma vaccine trials are oriented toward patients with advanced disease, because these individuals have the least to lose and the most to gain by volunteering for experimental therapies. However due to the generally low toxicity of melanoma vaccines compared with other adjuvant regimens such as high dose interferon alpha, it is only matter of time before these therapies are applied on a routine basis for stage I patients at high risk for relapse. As the efficacy issues are worked out in patients with more advanced disease, the case for offering the same benefits to high-risk stage I patients will become compellingÝ. Thus, it is important for dermatologists to become familiar with melanoma vaccines since it is likely that they will play in increasingly important role in the management of stage I patients. The skin specialist who makes the diagnosis should be prepared to offer appropriate treatment and/or referral.

References

1. Ollila DW, Kelley MC, Gammon G, Morton DL. Overview of melanoma vaccines: active specific immunotherapy for melanoma patients. Semin Surg Oncol 1998;14(4):328-36. PubMed

2. Berd D, Kairys J, Dunton C, Mastrangelo MJ, Sato T, Maguire HC Jr. Autologous, hapten-modified vaccine as a treatment for human cancers. Semin Oncol 1998;25(6):646-53. PubMed

3. Soiffer R, Lynch T, Mihm M, Jung K, Rhuda C, Schmollinger JC, Hodi FS, Liebster L, Lam P, Mentzer S, Singer S, Tanabe KK, Cosimi AB, Duda R, Sober A, Bhan A, Daley J, Neuberg D, Parry G, Rokovich J, Richards L, Drayer J, Berns A, Clift S, Dranoff G, et al. Vaccination with irradiated autologous melanoma cells engineered to secrete human granulocyte-macrophage colony-stimulating factor generates potent antitumor immunity in patients with metastatic melanoma. Proc Natl Acad Sci U S A 1998;95(22):13141-6. PubMed

4. Schreiber S, Kampgen E, Wagner E, Pirkhammer D, Trcka J, Korschan H, Lindemann A, Dorffner R, Kittler H, Kasteliz F, Kupcu Z, Sinski A, Zatloukal K, Buschle M, Schmidt W, Birnstiel M, Kempe RE, Voigt T, Weber HA, Pehamberger H, Mertelsmann R, Brocker EB, Wolff K, Stingl G. Immunotherapy of metastatic malignant melanoma by a vaccine consisting of autologous interleukin 2-transfected cancer cells: outcome of a phase I study. Hum Gene Ther 1999;10(6):983-93. PubMed



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