Dendritic-cell- and peptide-based vaccination strategies for glioma
- First Online:
- Cite this article as:
- Yamanaka, R. Neurosurg Rev (2009) 32: 265. doi:10.1007/s10143-009-0189-1
- 250 Views
Despite advances in radiation and chemotherapy along with surgical resectioning, the prognosis of patients with malignant glioma is poor. Therefore, the development of a new treatment modality is extremely important. There are increasing reports demonstrating that systemic immunotherapy using dendritic cells and peptide is capable of inducing an antiglioma response. This review highlights dendritic-cell- and peptide-based immunotherapy for glioma patients. Dendritic-cell- and peptide-based immunotherapy strategies appear promising as an approach to successfully induce an antitumor immune response and increase survival in patients with glioma. Dendritic cell- and peptide-based therapy of glioma seems to be safe and without major side effects. There are several types of glioma; so to achieve effective therapy, it may be necessary to evaluate the molecular genetic abnormalities in individual patient tumors and design novel immunotherapeutic strategies based on the pharmacogenomic findings. Here, in this review, recent advances in dendritic-cell- and peptide-based immunotherapy approaches for patients with gliomas are discussed.
KeywordsDendritic cellsGliomaPeptide vaccinationImmunotherapy
Malignant gliomas are the most prevalent type of primary brain tumor in adults. Despite progress in brain tumor therapy, the prognosis of malignant glioma patients remains dismal. The median survival of patients with glioblastoma multiforme, the most common grade of malignant glioma, is around 15–20 months [48, 49]. Conventional therapy of surgery, radiation, and chemotherapy is largely palliative. Essentially, tumor recurrence is inevitable. Salvage treatments upon recurrence are palliative at best and rarely provide significant survival benefit. Basic research is providing novel insights into the complex molecular pathways involved in the pathogenesis of malignant glioma transformation and progression. By unraveling the intricate signaling cascades responsible for sustained proliferation, angiogenesis, invasion, and resistance to apoptosis in glioma, we are now confronted with an ever-expanding list of molecular targets. Technologic advances in oncogenomics, proteomics, and functional genomic screens are providing mechanisms to rapidly identify the critical targets whose inactivation will lead to a substantive tumor growth arrest. Tumor tissue biomarkers that identify those tumors most likely to respond to a specific inhibitor are needed as a mechanism toward tailoring therapy to the individual patient with malignant glioma. Therapies targeting the underlying molecular pathogenesis of brain tumors are urgently required. Common genetic abnormalities in malignant glioma specimens are associated with aberrant activation or suppression of cellular signal transduction pathways and resistance to radiation and chemotherapy. Additional clinical studies will combine novel targeted therapies with radiation, chemotherapies, and immunotherapies.
Cellular and molecular biology of gliomas
Recent scientific advances have enhanced our understanding of the biology of gliomas and the role of tyrosine kinase receptors and signal transduction pathways in tumor initiation and maintenance, such as the epidermal growth factor receptors, platelet-derived growth factor receptors, vascular endothelial growth factor receptors, and the Ras/Raf/mitogen-activated protein kinase and phosphatidylinositol-3 kinase/Akt/mammalian target of rapamycin pathways. Novel targeted drugs such as small molecular inhibitors of these receptors and signaling pathways are showing some activity in initial studies . As we learn more about these drugs and how to optimize their use as single agents and in combination with radiation, chemotherapy, and other targeted molecular agents, they will likely play an increasing role in the management of this devastating disease.
These include the importance of O6-methylguanine-DNA-methyltransferase in glioblastoma sensitivity to the DNA alkylating chemotherapy temozolomide . In addition, coexpression of phosphatase tensin homolog on chromosome ten and a mutant variant of the epidermal growth factor receptor (EGFRvIII) appears to predict sensitivity of recurrent glioblastomas to EGFR inhibitors . Finally, loss of heterozygosity for chromosomes 1p and 19q correlates with both response to therapy and improved prognosis for patients with oligodendrogliomas . The best way to incorporate these findings into office practice, however, remains unclear, especially in the absence of effective alternatives to currently available treatments. While subdividing gliomas molecularly may allow tailoring of therapy to individual patients based on individual tumor biology, a superior strategy may be to identify common molecular abnormalities that are targets of more universally applicable therapies.
Recently, a population of glioma stem cells has been isolated. This subpopulation of stem-like cells plays an important role in the tumorigenic process [20, 46, 51, 63]. Since glioma stem cells can self-propagate, it may also be important to specifically target glioma stem cells, to avoid recurrence of the glioma. The identification of glioma stem cells should provide new molecular targets for treating gliomas.
Therapeutic cancer immunotherapy has become a therapeutic option because tumor-associated antigens recognized by tumor-reactive cytotoxic T lymphocytes (CTL) have been identified during the last two decades . In fact, a large number of clinical trials for cancer immunotherapy have been carried out worldwide . Among the vaccines tested, cell-based and peptide vaccines have an advantage for the treatment of cancers.
Although tumor-specific endogenous immunity can be elicited in cancer patients by overexpressing tumor antigens, the generated immune response is not capable of preventing tumor growth. Several factors may play a role in inhibiting the antitumor efficacy of the immune response, including the immunosuppressive environment of the tumor, the low avidity of the T cells for the antigen expressed on the tumor, and the low magnitude of the endogenous immune response.
Glioma cells are considered to be poor antigen-presenting cells (APC) because of secretion of immunosuppressive cytokines and factors such as transforming growth factor beta  and vascular endothelial growth factor . An increased regulatory T cell fraction appears to play a critical role in tumor tolerance [9, 40]. The high expression of HLA-E on gliomas, upon binding to NKG2A/B, has a negative effect on T cell and NK cell activity . Cancer vaccines designed to augment tumor-specific cellular immunity could partially overcome these defects by boosting low-level immunity and stimulating the proliferation of higher-avidity T cells that are capable of homing to a tumor [34, 36, 58, 61]. Recent reports demonstrate that systemic immunotherapy using dendritic cells (DCs) and peptide antigens is capable of inducing an antitumor response within the immunologically privileged brain, confirming that the central nervous system (CNS) is accessible to systemic immunotherapy [17, 18, 38, 58–62].
Van der Bruggen et al.  first reported a cDNA expression cloning technique to identify genes and peptides of tumor-associated antigens in 1991. Subsequently, a reverse immunology technique using autologous antibody was introduced for the identification of genes and peptides recognized by the host immune system . These advanced techniques have identified a large number of tumor antigens and peptides with potential applications as cancer vaccines. Peptides used as cancer vaccines usually consist of nine amino acids capable of binding to a particular major histocompatibility complex (MHC) class I antigen with the ability to activate CTL reactive to tumor cells.
Tumor antigens expressed by human malignant neoplasms can be classified into three different groups: (1) the differentiation antigens; (2) the products of viral, mutated, differentially spliced, overexpressed genes; (3) housekeeping and/or metabolic pathway antigens. Both melanocytes and glial cells are derived from the neural ectoderm , and their malignant transformed counterparts, melanoma and glioma cells, may share common biological properties. Melanoma antigen-encoding genes (MAGE)-1 , MAGE-E1 , and MAGE-3 were expressed in different types of gliomas but never in normal brain tissue. MAGE-1, MAGE-E1, and MAGE-3 might be potential targets for active specific immunotherapy. Other possible glioma-associated antigens have also been described. These include altered EGFRvIII, tenascin, and GP 240 which are extracellular matrix-associated molecules, primarily overexpressed in malignant glioma . The expression of Homo sapiens testis (HOM-TES)-14/stromal-cell-derived protein 1, HOM-TES-85, synovial sarcoma X breakpoint (SSX-1), SSX-2 , GAGE-1 , Sry-related high-mobility group box-containing gene 5,6 cancer testis antigen [52, 53], interleukin (IL)-13 receptor alpha 2 , epherin A2 , antigen isolated from immunoselected melanoma 2 , squamous cell carcinoma antigen recognized by T cells (SART) 1 , SART 3 , kinesin superfamily protein (KIF) 1C, and KIF3C  were confirmed in glioma.
Dendritic cells in tumor immunology
DCs originate from CD34+ bone marrow stem cells. Human DC precursors are found in the bone marrow as well as peripheral blood and, in a more mature form, in lymphoid and nonlymphoid tissues. Three different subtypes of DC have been defined: Langerhans cells, interstitial DCs, and plasmacytoid DCs. Human skin contains the first two types of DCs, which are generally referred to as myeloid DCs. The plasmacytoid cells are derived from the lymphoid lineage and are found in the T cell areas of lymphoid organs, the thymus, and in the peripheral circulation . The most widely emphasized functions of DCs are antigen uptake and processing. DCs are considered to be the nature’s best APCs. Immature DCs have several features that allow them to capture antigens. They can take up antigens via receptor- or nonreceptor-mediated mechanisms. Upon being internalized, tumor antigens are processed and split into peptides in the cytosol or endocytic vesicles in DC, which are then reexpressed on the cell surface in association with MHC molecules. To accomplish this complex series of events, DCs are equipped with a sophisticated molecular array of cell components representing the antigen-processing machinery (APM). The APM is essential for the uptake and processing by DCs of tumor-derived antigens, so that tumor-derived epitopes can be cross-presented to T cells. Immature DCs are characterized by their ability to induce a tolerizing immune phenotype through activation of regulatory T cells and inability to stimulate naïve or antigen-specific memory T cells [6, 55]. The presence of toll-like receptor (TLR) ligands provided by microorganisms or stimulating cytokines induces a transformation into fully activated mature DCs, with resulting presentation of the internalized entities on appropriate MHC molecules. Mature DCs express high levels of class I and class II molecules, altered chemokine expression, and co-stimulatory molecules [2, 7]. They migrate to draining lymph nodes and activate innate and adaptive immune response.
Human tumors express a variety of protein antigens recognizable by the immune system, and these antigens are potential targets for cancer vaccination therapy. Unfortunately, the tumor antigens are self-derived antigens and are generally considered weak antigens. Selection of tumor antigen and appropriate loading of in-vitro-generated DC with the antigen is an initial and crucial step in the development of an efficient DC-based cancer vaccine.
DC-based therapy in patients with cancer is now largely in phase II trials. With the safety of DC transfers established, the challenge of the ongoing clinical studies will be to determine effective therapeutic doses and to obtain evidence for clinical efficacy of this form of immunotherapy. Opportunities will be available in the context of these clinical trials to acquire a better understanding of how DCs mediate antitumor effects. A number of questions that have to be addressed concern, e.g., optimization of culture conditions for DCs, especially if tailored subsets of polarized DCs are to be produced; definition of cytokine/chemokine profiles that characterize different DC subsets; establishment of conditions for DC polarization or repolarization toward clinically beneficial type 1 T cell responses; and, finally, finding the means to sustain DC functions in the hostile tumor microenvironment. Studies are necessary to be able to understand which subsets of DCs exert immunogenic vs. tolerogenic effects in vivo. In a study involving patients with stage IV melanoma, de Vries et al.  directly compared the efficacy of vaccines using immature or mature DCs in inducing an immune response. They reported that delayed-type hypersensitivity reactions (DTH) and humoral responses to keyhole limpet hemocyanin (KLH) were observed in patients receiving mature DCs whereas those receiving immature DCs had no DTH reactions. The plasticity of DCs and the potential for differential regulation of their state of maturation have to be carefully handled to assure that cancer patients receive adoptive transfers of immunogenic DCs, engineered to promote Th1- and Tc1-type (type 1 CD8 T cell) tumor-specific responses. These and other issues are components of future translational research aimed at the understanding of the biology of DC subsets, their mechanism of action, and their utility for immunotherapy not only of cancer but also of other diseases.
Dendritic cell vaccine for glioma patients
There are several reports concerning clinical trials of DC-based vaccine for patients with glioma. Yu et al.  reported phase I clinical trial of peripheral blood DC pulsed with peptides eluted from autologous glioma cells. Two patients had recurrent anaplastic astrocytoma and seven glioblastoma (GBM). Peptide-pulsed DCs were injected intradermally in the deltoid region three times biweekly. DC vaccination elicited systemic cytotoxicity in seven patients, and intratumoral cytotoxic and memory T cell infiltration were detected in two patients. DC vaccination proved to be associated with increased survival: median survival times for the study group and the control group were 455 and 257 days, respectively. This phase I study demonstrated the feasibility, safety, and bioactivity of DC vaccine. Yu et al.  reported another phase I trial of peripheral blood DC pulsed with tumor lysate of autologous glioma cells. Four patients had recurrent anaplastic astrocytoma and ten GBM. Six of ten patients demonstrated robust systemic cytotoxicity as demonstrated by interferon gamma expression by peripheral blood mononuclear cells (PBMCs) in response to tumor lysate after vaccination. A significant CD8+ T cell infiltrate was noted intratumorally in three of six patients who underwent reoperation. The median survival for patients with recurrent GBM in this study was 133 weeks. Vaccination with tumor lysate-pulsed DC was safe and no evidence of autoimmune disease was noted.
Kikuchi et al.  reported the immunotherapy with fusions of DC and glioma cells. Three patients had recurrent anaplastic astrocytoma and five GBM. Clinical results showed that there were no serious adverse effects and two partial responses (PR). Kikuchi et al.  reported another immunotherapy with fusions of DC and glioma cells combined with recombinant IL-12. Nine patients had recurrent anaplastic astrocytoma and six GBM. Clinical results showed that there were no serious adverse effects and four PR and one minor response (MR) in patients with anaplastic astrocytoma.
Rutkowski et al.  reported phase I trial of peripheral blood DC pulsed with tumor lysate of autologous glioma cells. One patient had recurrent anaplastic astrocytoma and 11 GBM. There were no serious adverse effects and clinical or radiological evidence of autoimmune reactions in any of the patients in these studies except one patient who repetitively developed peritumoral edema. Two of six patients with complete resection have continuous complete response (CR) for 3 years.
Yamanaka et al. [59, 60] reported the DC therapy pulsed by tumor lysate. Twenty-four patients with recurrent malignant glioma (six grade III and 18 grade IV patients) were evaluated in a phase I/II clinical study. DCs were injected intradermally or both intratumorally and intradermally every 3 weeks. The protocols were well tolerated with only local redness and swelling at the injection site in several cases. Clinical responses were one patient with PR and three MR. Increased enzyme-linked immunosorbent spot (ELISPOT) and DTH responses after vaccination could provide good laboratory markers to predict the clinical outcome of patients receiving DC vaccination. The overall survival of patients with grade IV glioma was 480 days, which was significantly better than that in the control group.
Dr. Liau et al.  have reported tumor lysate-pulsed DC vaccine in combination with TLR-7 agonist, imiquiod, following radio-chemotherapy for newly diagnosed GBM. Thirteen patients received three immunizations at 2-week intervals, following completion of a 6-week course of radio-chemotherapy. Patients without tumor progression received booster vaccinations combined with topical administration of the TLR-7 agonist imiquiod. All immunization were well tolerated, with only mild side effects. Increased levels of CD8+ T cells reactive against tumor antigens were detected in five patients. The median progression-free survival time (PFS) is 18.1 months and median overall survival is 33.8 months. And she is currently enrolling patients with GBM for a multicenter phase II to test the efficacy of their autologous DC vaccine. The target enrollment for the trial is newly diagnosed GBM patients. Patients will undergo surgery, radiation, and chemotherapy. The primary end point for the trial is patient survival with no disease progression and the second end point is overall survival.
Although immunological monitoring including tetramer assay and ELISPOT assay may detect specific antiglioma immunity, clinical efficacy has been minimal following immunotherapies. Due to the limited sample population, further evaluation of the role of DC immunotherapy is necessary. The optimum dose of DCs, the appropriate route of vaccinations, the source of tumor antigens, and methods of antigen loading should be determined.
Peptide vaccination clinical trials
Several clinical trials of peptide-based immunotherapy for cancer have been conducted in the past decade, but major clinical responses were rarely obtained . Rosenberg et al.  summarized the clinical responses of peptide-based vaccine therapy. Objective response rates for peptide vaccines administered to patients with metastatic cancers at the National Cancer Institute in the USA were 2.9% (11 of 381 cases). Similarly, no objective response was obtained in peptide vaccine trials for advanced solid cancers [31, 50]. In addition, a large number of clinical trials of peptide-based cancer vaccines are ongoing as translational clinical research.
Peptides used as cancer vaccines usually consist of nine amino acids capable of binding to a particular MHC class I antigen with the ability to activate CTL reactive to tumor cells. A peptide suitable for the individual patient is generally mixed with an adjuvant followed by subcutaneous administration every 7–14 days to form a vaccine. It is hypothesized that the injected peptide is captured by APCs, which in turn move to regional lymph nodes. Soon after, they present the loaded peptides to the circulating CTL, which possess T cell receptors specific to the corresponding peptide. CTL recognizing a peptide on APC become activated in association with clonal expansion in the nodes. These activated CTL come out through lymph nodes or blood circulation, migrate and infiltrate into tumor sites, recognize the corresponding peptide–MHC complex on cancer cells, and eliminate cancer cells, which in turn results in tumor regression.
Dr. Itoh et al. [30, 58] have developed the personalized peptide vaccination, namely patients exhibiting preexisting responses to specific peptides are then vaccinated with those peptides. This strategy is based on the assumption that initiation of immune boosting of CTLs through peptide vaccination could be more effective than immune priming of naïve T cells with regard to induction of prompt and strong immunity. Vaccination with peptides in patients with higher levels of CTL precursors in prevaccination PBMCs might induce stronger and faster activation of CTL compared to that with rare CTL precursors. Consistent with this notion, the objective response rates of classical (nonpersonalized) peptide vaccines were 0%, whereas that of personalized vaccines was 11.1% in the total advanced cancers .
Yajima et al.  reported the phase I study of personalized peptide vaccination for patients with advanced malignant glioma. Twenty-five patients with advanced malignant glioma (eight grade III and 17 grade IV) were evaluated in a phase I clinical study. For personalized peptide vaccination, prevaccination PBMCs and plasma were provided to examine cellular and humoral responses to 25 or 23 peptides in HLA-A24+ or HLA-A2+ patients, respectively, and then only the reactive peptides (maximum of four) were used for in vivo administration. The peptides derived from SART3, Lck (lymphocyte-specific protein tyrosine kinase), and multidrug resistance-associated protein 3 antigens were most frequently selected for vaccination. The protocols were well tolerated with the major adverse effects being a grade 1 or 2 inflammatory skin reaction at the injection site. Increases in cellular or humoral responses specific to at least one of the vaccinated peptides was observed in the postvaccination (sixth) samples from 14 or 11 of 21 patients, respectively. Clinical responses were five PR, eight stable diseases, and eight progressive diseases. The median PFS was 3 months and the PFS at 6 months was 13%.
Izumoto et al.  reported phase II trial of Wilmus tumor 1 (WT1) peptide vaccination clinical trial. They treated 21 patients with recurrent GBM. They received HLA-A-2402-restricted modified 9-mer WT1 peptide every week for 12 weeks. The protocol was well tolerated. The clinical response was two PR, ten stable diseases, and nine progressive diseases. The median PFS was 20 weeks and the 6-month PFS rate was 33.3%. WT1 peptide vaccinations were recommended for further clinical study to malignant glioma patients.
Dr. Sampson et al.  reported the effect of EGFRvIII-targeted vaccine (CDX-110) on immune response and prolonged PFS when given with simultaneous standard and continuous temozolomide in patients with newly diagnosed GBM. CDX-110 conjugated to KLH was given intradermally to 21 patients until tumor progression or death. The survival of the vaccinated patients is better than a matched historical control group. CDX-110 is under investigation in a phase III randomized clinical trial.
From the above information, it is apparent that new therapeutic strategies for gliomas need to take into consideration the unique molecular abnormalities, glioma stem cells, and glioma-associated antigens. To what extent these immunization protocols could be improved remains debatable. To achieve better immunization efficacy, various strategies have been proposed. Direct manipulation of professional APCs to ensure optimal presentation or indirect antigen presentation through molecules, DC-derived exosomes, or cellular vaccines may equally lead to sufficient immune reaction. Injection of the vaccine into lymph nodes could potentially be more efficient because the antigens become quickly available to professional APCs.
The other possibility is to enhance or modify the APC ability of DCs. This would be achieved by adjuvants to upregulate DC function. It would be also important to develop efficient means to generate and present a large number of stable peptide–MHC complexes for long periods of time. How to modulate DC to obtain the efficient immunostimulatory effect should be established. Many new subsets of DC are being identified. Functional diversity of DC subsets cannot be explained by different lineage origin but depends on the activation signals, maturation stage, and local microenvironment . It may be more clinically relevant to consider DC as a mass consisting of many phenotypically and functionally diverse cells. It will be a new challenge to exploit these differences for the purposes of immunotherapy of glioma. The fundamental issues should be further clarified whether in-vitro-derived DC has the capacity to migrate to the lymphoid organ when administered back to the patients.
Several reports have provided evidence that immunotherapy was clinically effective from the perspective of the overall survival of some grade IV glioma patients by DCs or a virus-modified autologous tumor cell vaccine [17, 18, 38, 47, 59–62]. However, such immunotherapies have several disadvantages: limited materials for vaccination, labor intensity for preparation, and difficulty to find a reliable laboratory marker. Tumor lysates or RNA made from tumor tissue is often in limited quantities, and they are an inconsistent source of antigenic material and can make immune monitoring difficult and they may often be contaminated with nontarget cells such as normal brain. In contrast, peptide vaccination has several advantages including easy supply of good manufacturing practice levels of materials and a reliable laboratory marker for the prediction of clinical outcome. However, peptide has problems of the potential of tumor antigens escape and limited repertoire of using defined antigens and because they are human leukocyte antigen class I and class II restricted and, consequently, restrict patient enrollment into clinical trials and class I peptides are insufficient to generate a CD4+ T helper response which are required for optimal effective antitumor immunity .
The first point to improve immunotherapy is how to eliminate MHC class I loss of cancer cells because a large population (30–60%) of cancer cells do not express MHC class I molecules, which are crucial for CTL-mediated elimination of cancer cells . This problem could be overcome by the combined use of a DC or peptide vaccine and either chemotherapy or biological therapy capable of activating natural killer cells and macrophages that are not affected by MHC expression on cancer cells. There are several reports describing sensitization of malignant glioma to chemotherapy through vaccination [26, 27, 56]. Based on these experiences, we propose a DC or peptide vaccine combined with chemotherapy as a new treatment modality for gliomas. The second point is how to overcome the immunosuppressive state of glioma patients. Patients with recurrent glioma are usually in an immunosuppressive state because of the advanced disease and myelosuppression by anticancer agents . It should be difficult to induce antiglioma immunoresponses in such a condition by immunotherapy. Therefore, prophylactic immunotherapy at initial stage of the disease may have significant merit.
DC- and peptide-based strategy appears promising as an approach to successfully induce antitumor immune responses and prolong survival in patients with glioma. DCs and peptide therapy of glioma seem to be safe and without major side effects. Its efficacy should be determined in randomized and controlled clinical trials. Every patient with gliomas will be evaluated for the molecular genetic abnormalities in their individual tumors, and novel immunotherapeutic strategies based on pharmacogenomics will be offered according to the genetic findings.