Progress in active-specific immunotherapy of brain malignancies.

Despite the significant advances in neurosurgical techniques and oncology treatment regimens, the prognosis of patients with brain malignancies remains dismal. Brain tumours remain as lethal in the beginning of this new millennium as they were 30 years ago. Among the promising treatment modalities being tested are various immunotherapeutic approaches. Development of cancer vaccines, also known as active-specific immunotherapy, for malignant brain tumours is summarised in this review. Understanding the mechanisms behind vaccinations and the initiation of immune response have helped the design and improvement of the efficacy of clinical vaccines. The emergence of the antigen-presenting properties of dendritic cells brings the cancer vaccine field into a new generation. Preclinical work on the use of dendritic cell-based vaccine for malignant brain tumours are encouraging. The move from these preliminary studies to the clinic is anticipated with high hope.

Development of dendritic-cell based prostate cancer vaccine.

Available treatments for metastatic prostate cancer have failed to demonstrate significant curative potential. Current efforts are now directed towards developments of novel strategies for the treatment of metastatic prostate cancer. Cancer immunotherapeutic strategies utilize patient immune system components to kill cancer cells. This review discusses progress in active specific immunotherapeutic approaches as potential alternative methods in the treatment of metastatic prostate cancer. One of the newest advances in cancer immunotherapy is the use of dendritic cells as the vehicle to deliver cancer antigens for an effective in vivo T cell activation. The development of dendritic cell-based prostate cancer vaccine, as well as results of several clinical trials in prostate cancer involving the administration of peptide-pulsed autologous dendritic cell pulsed are discussed.

Higher-dose and less frequent dendritic cell infusions with PSMA peptides in hormone-refractory metastatic prostate cancer patients.

BACKGROUND: Infusion of dendritic cells (DCs) pulsed with PSMA peptides was considered possible in hormone-refractory metastatic prostate cancer patients both with or without prior treatment with a greater number of DCs and for lesser infusions than previously administered. METHODS: DCs + PSMA peptides in patients undergoing leukapheresis were administered monthly 1-4 times, at rates greater than 20 million DCs in 17 patients not previously treated, and in 11 patients previously treated. RESULTS: Three partial responders and one complete responder were noted in the 17 previously untreated persons. DCs + PSMA peptides averaged 28.5 million cells (range in millions, 21.0-42.3). All responders received 3 or 4 infusions of greater than 22 million cells (3-4 times). In the previously treated group of 11 patients, DCs infused averaged 29.3 million cells (range in millions, 20-40.5). One new responder (bone scan) was noted. Two prior responders continued. Observation times were similar. Toxicity was minimal. CONCLUSIONS: These results suggest that DCs + PSMA peptide infusions can be given with greater numbers of DCs with a lesser number of infusions (1-4 monthly) with no loss of response rates compared to those noted previously, and without increased side effects. In previously treated patients (both relapsing and nonrelapsing), adverse effects were not noted, and new responses can be anticipated to be without harmful side effects. However, the follow-up time, and number of patients in this group, were small.

Progress in active specific immunotherapy of prostate cancer.

Treatments available for metastatic prostate cancer have failed to demonstrate significant curative potential. Current efforts are now directed towards developments of novel strategies for the treatment of metastatic prostate cancer. Cancer immunotherapeutic strategies utilize patient immune system components to kill cancer cells. This review discusses progress in active specific immunotherapeutic approaches as potential alternative methods in the treatment of metastatic prostate cancer. Various methods of augmenting the immune response against prostate cancer are discussed including systemic cytokine adjuvant therapy, cytokine gene transduced tumor vaccines, non-antigen specific immunization, DNA and peptide vaccines plus adjuvants, as well as dendritic cell-based cancer vaccines.

Use of artificial neural networks in evaluating prognostic factors determining the response to dendritic cells pulsed with PSMA peptides in prostate cancer patients.

BACKGROUND: Our purpose was to compare the importance of over 22 measurements used in evaluating the clinical responses of patients with metastatic or locally recurrent prostate cancer, treated by dendritic cell (DC) infusions with prostate-specific membrane antigen (PSMA) peptides. METHODS: Artificial neural networks (ANNs) were employed for assessment, as well as the traditional methods of logistic regression. RESULTS: Twenty-six patients with metastatic disease and 37 patients with local recurrence were available for evaluation and comparison. ANN evaluation ranked the collective effects of DC infusion, immune responses (CD3+ cells, CD16+ cells, zeta chain+ cells), and cytokines, e.g., IL-6 and PSMA levels, very highly. Logistic regression identified all of these parameters to some degree, but in a different rank order. Patients with metastases showed a sharp rate of response secondary to the level of DC infusion, in contrast to those patients with local recurrence, in which it was more gradual. CONCLUSIONS: ANN analysis emphasizes the importance of level of DC infusion, immune parameters, cytokines, and markers such as PSMA in determining the response to PSMA peptide immunotherapy. The criteria of response were judged to be correct in 86% of metastatic patients and 83% of locally recurrent patients evaluated in this study.

Follow-up evaluation of a phase II prostate cancer vaccine trial.

BACKGROUND: A phase II trial, involving infusions of autologous dendritic cells (DC) and two human histocompatibility antigen (HLA-A2)-specific prostate-specific membrane antigen (PSMA) peptides, was recently completed. Thirty percent of the participants, including subjects with hormone-refractory metastastic disease, and those with suspected local recurrence of prostate cancer, were identified as clinical responders. This report describes the follow-up evaluation of 19 responders in the two study groups. METHODS: After conclusion of the study, study participants were subjected to follow-up evaluations at 6-8-week intervals. Each responder was reevaluated for response status, and duration of response was determined. RESULTS: Subjects were observed for an average of 291 days (metastastic group, group A-2) and 557 days (local recurrence group, group B), which included the treatment and follow-up periods. The average duration of response was 149 days for group A-2, and 187 days for group B. A majority of responders (11/19; 58%) were still responsive at the end of the current follow-up. CONCLUSIONS: The responses observed may be significant and relatively durable. This study suggests that DC-based cancer vaccines in the future may provide an additional therapy for advanced prostate cancer.

Vaccine therapy for prostate cancer.

Vaccine therapy may provide an alternative for prostate cancer patients whose disease no longer responds to hormone therapy. Administration of dendritic cells pulsed with prostate-specific membrane antigen (PSMA) induces cellular immune responses against the tumor with virtually no adverse effects. About 30% of the evaluable patients were identified as partial responders, based on the National Prostate Cancer Project (NPCP) criteria. In addition, there was a 50% decrease of serum prostate-specific antigen or resolution of previously measurable lesions on imaging. Dendritic cell vaccine therapy may have a synergistic effect, when combined with other therapies.

GM-CSF as a systemic adjuvant in a phase II prostate cancer vaccine trial.

BACKGROUND: Recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF; Leukine [sargramostim], Immunex Corp., Seattle, WA) was administered to a subgroup of 44 patients in a phase II clinical trial for prostate cancer using DC pulsed with HLA-A2-specific prostate-specific membrane antigen (PSMA) peptides. Our purpose was to determine if GM-CSF caused any enhancement of patients' immune responses, including enhancement of clinical response to the DC-peptide treatment. This report compares the clinical responses to DC-peptide infusions with and without systemic GM-CSF treatment. METHODS: GM-CSF was administered by subcutaneous injection at a dose of 75 microg/m2/day for 7 days with each of six infusion cycles. Prefilled syringes were supplied to the patients for self-administration. RESULTS: One complete and 8 partial responders were identified among 44 patients who received GM-CSF, as compared to 2 complete and 17 partial responders among 51 patients who did not receive GM-CSF. For patients who received GM-CSF and were tested by delayed-type hypersensitivity (DTH) skin test, 3 cases of improved immune response were identified, compared to 5 cases of improvement in patients who did not receive GM-CSF. The main GM-CSF side effects reported were local reactions at the site of injection, fatigue, pain, and fever. Most reported side effects were of mild severity, with some cases of moderate severity leading to discontinuation of GM-CSF. CONCLUSIONS: Our results suggest GM-CSF as employed in this trial did not detectably enhance clinical response to DC-peptide infusions, or significantly enhance the measured immune response.

Phase II prostate cancer vaccine trial: report of a study involving 37 patients with disease recurrence following primary treatment.

BACKGROUND: A phase II trial was conducted to assess the efficacy of infusions of dendritic cells (DC) and two HLA-A2-specific prostate-specific membrane antigen (PSMA) peptides (PSM-P1 and -P2). This report describes the evaluation of 37 subjects admitted with presumed local recurrence of prostate cancer after primary treatment failure. METHODS: All subjects received six infusions of DC pulsed with PSM-P1 and -P2 at 6-week intervals. Clinical monitoring was conducted pre-, during, and post-phase II study. Data included: complete blood count, bone and total alkaline phosphatase, prostate markers, physical examination, performance status, bone scan, ProstaScint scan, and chest X-ray, as well as other assays to monitor cellular and humoral immune responses. RESULTS: One complete and 10 partial responders were identified from this group based on National Prostate Cancer Project criteria, or on a 50% reduction of prostate-specific antigen (PSA), or on a significant resolution in lesions (biopsy-proven when possible) on ProstaScint scan. CONCLUSIONS: About 30% of study participants in this group showed a positive response at the conclusion of the trial. This study suggests that DC-based cancer vaccines may provide an alternative therapy for prostate cancer patients whose primary treatment failed.

Infusion of dendritic cells pulsed with HLA-A2-specific prostate-specific membrane antigen peptides: a phase II prostate cancer vaccine trial involving patients with hormone-refractory metastatic disease.

BACKGROUND: A phase II trial was conducted to assess the efficacy of infusions of dendritic cells (DC) and two HLA-A2-specific PSMA peptides (PSM-P1 and -P2). This report describes thirty three subjects with hormone-refractory metastatic prostate cancer without prior vaccine therapy history who were evaluated and reported as a group. METHODS: All subjects received six infusions of DC pulsed with PSM-P1 and -P2 at six week intervals. Clinical monitoring was conducted pre-, during, and post- phase II study. Data collected include: complete blood count, bone and total alkaline phosphatase, prostate markers, physical examination, performance status, bone scan, ProstaScint scan, chest x-ray, as well as assays to monitor cellular immune responses. RESULTS: Six partial and two complete responders were identified in the phase II study based on NPCP criteria, plus 50% reduction of prostate-specific antigen (PSA), or resolution in previously measurable lesions on ProstaScint scan. CONCLUSIONS: Over 30% of study participants in this group showed a positive response at the conclusion of the trial. This study suggested that DC-based cancer vaccines may provide an alternative therapy for prostate cancer patients whose disease no longer responds to hormone therapy.

Dendritic cell-based immunotherapy for prostate cancer.

Dendritic cells are unique in their ability to stimulate naive T cells. These investigators have developed a prostate cancer vaccine using autologous dendritic cells as a vehicle to present prostate antigens to T cells in vivo.

Evaluation of phase I/II clinical trials in prostate cancer with dendritic cells and PSMA peptides.

BACKGROUND: A phase I trial involving patients with advanced prostate cancer was conducted to assess the safe administration of dendritic cells (DC) and HLA-A0201-specific prostate-specific membrane antigen (PSMA) peptides (PSM-P1 or -P2). Thirty-three of the phase I participants were subsequently enrolled in a phase II trial, which involved six infusions of DC pulsed with PSM-P1 and -P2 peptides. METHODS: Clinical monitoring was conducted up to 770 days from the start of the phase I study. Data collected included: complete blood count, bone and total alkaline phosphatase, prostate markers, physical examination, performance status, bone scan, ProstaScint scan, and chest X-ray, as well as assays to monitor cellular immune responses. RESULTS: Nine partial responders were identified in the phase II study based on National Prostate Cancer Project (NPCP) criteria, plus 50% reduction of prostate-specific antigen. Four of the partial responders were also responders in the phase I study, with an average response duration of 225 days. Their combined average total response period was over 370 days. Five other responders were nonresponders in the phase I study. Their average partial response period was 196 days. CONCLUSIONS: The responses observed in the phase I and II clinical trials were significant and of long duration. The partial-responder group included patients who continued to respond from phase I, as well as those who started to respond during the phase II trial.

Report of immune monitoring of prostate cancer patients undergoing T-cell therapy using dendritic cells pulsed with HLA-A2-specific peptides from prostate-specific membrane antigen (PSMA).

BACKGROUND: In this paper we describe our program for the immune monitoring of phase II participants given dendritic cell (DC)/prostate-specific membrane antigen (PSMA)-based immunotherapy, and we also present some initial findings. METHODS: Phase II subjects received six administrations of autologous dendritic cells exogenously pulsed with two peptides derived from PSMA. Prior to the initial infusion, and following each treatment, peripheral blood mononuclear cells (PBMC) were collected for the generation of dendritic cells as well as for comprehensive immune monitoring. RESULTS: Thus far, an increase in PSMA-peptide-specific as well as overall cellular reactivity has been observed in several patients receiving DC plus PSM-P1 and -P2, as measured by delayed-type hypersensitivity (DTH) test and enzyme-linked immunosorbant assay (ELISA). CONCLUSIONS: Our initial observations using an ELISA and DTH test indicate that we are enhancing cellular immunity in prostate cancer patients following infusion with DC plus PSMA-derived peptides. Several methods are underway to comprehensively monitor both cell-mediated and humoral immune responsiveness, including: determining anti-PSMA serum antibody titers, testing immunogen-restricted responder-cell proliferation and cytotoxicity, assessing aberrations in signal transduction, antigen processing, and presentation, and measuring soluble factors that may promote tumor outgrowth.

Dendritic cell-based immunotherapy of prostate cancer.

The immunotherapy of cancer, based on eliciting or enhancing the body's own capacity to mount an effective antitumor response, has produced encouraging early results in the areas of melanoma and renal-cell carcinoma. Such treatments utilizing dendritic cells (DC), immune cells that are excellent antigen presenters, are especially promising. We performed a phase I clinical trial assessing the administration of autologous DC pulsed with HLA-A0201-specific prostate-specific membrane antigen (PSMA) for the treatment of 51 men with hormone-refractory prostate cancer. Participants were divided into five groups receiving four or five infusions of peptides alone (PSM-P1 or PSM-P2; group 1 and 2, respectively), autologous DC (group 3), or DC pulsed with PSM-P1 or P2 (group 4 and 5, respectively). No significant toxicity was observed. Immune reactivity against PSM-P2 was detected in HLA-A2+ patients infused with DC pulsed with PSM-P1 or -P2 (group 4 and 5). An average decrease in PSA was observed only in group 5. Seven partial responders were identified based on NPCP criteria + PSA. The excellent tolerance of this treatment approach, as well as the enhanced cellular responses, decreased PSA levels, and partial clinical responses in some patients suggests that it holds great potential in prostate cancer therapy.

Follow-up evaluation of prostate cancer patients infused with autologous dendritic cells pulsed with PSMA peptides.

BACKGROUND: We recently conducted a phase I clinical trial administering autologous dendritic cells pulsed with prostate-specific membrane antigen (PSMA) peptides to advanced prostate cancer patients. Participants were divided into 5 groups receiving 4 or 5 infusions of peptides alone (PSM-P1 or -P2; groups 1 and 2, respectively), autologous DC (group 3), or DC pulsed with PSM-P1 or -P2 (groups 4 and 5, respectively). Seven partial responders were observed. Follow-up evaluation of these responders is presented in this report. METHODS: Clinical monitoring for hematological studies and prostate markers was conducted up to 370 days from the start of the phase I study. Data collected include: lymphocyte, hematocrit, alkaline phosphatase, prostate-specific antigen (PSA), free PSA, and PSMA levels. RESULTS: Groups 4 and 5 (patients infused with DC pulsed with PSM-P1 or -P2) represented 5/7 responders. The length of response was between 100 days (1 patient) to 200 days or above (6 patients). Four patients still remained responsive at the end of the period of observation. CONCLUSIONS: The responses observed in this phase I clinical trial are significant and of long duration. Most of the responders were in treatment groups infused with DC pulsed with PSM-P1 or -P2, suggesting the requirement of both components for effective immunotherapy.

Cancer immunotherapy for prostate cancer.

Our approach to prostate cancer immunotherapy involves two components dendritic cells as antigen-presenting cells; and the antigen used to target T-cell attack, HLA-A0201-associated peptides from prostate specific membrane antigen (PSMA). We have conducted a phase I dose-ranging study in 51 men with advanced prostate cancer, using dendritic cells pulsed with a PSMA peptide. no significant toxicity was observed. In that study, T-cell response was enhanced, with seven men meeting NCPC and PSA criteria for partial response. We are now conducting a phase II study with 67 men, who will receive 6 infusions of dendritic cells that have been pulsed with 2 PSMA peptides, at 6-week intervals. The phase II study design and rationale is described in this paper.

Measurement of prostate-specific membrane antigen in the serum with a new antibody.

Work to date has identified prostate-specific membrane antigen (PSMA) as a membrane-bound glycoprotein with high specificity for prostatic epithelial cells. PSMA reacts with the monoclonal antibody 7E11.C5, which is present in serum, seminal fluid, and prostatic epithelial cells, and is increased in its expression in the presence of a hormone refractory state associated with prostatic cancer. This report confirms these results and further documents the presence of the monoclonal antibody 3F5.4G6, which reacts with the extracellular domain of PSMA. This region of PSMA is also an element present in a truncated version of the protein, so-called PSM'. Immune precipitation with either 7E11.C5 or 3F5.4G6 yields an isolated protein species that are reactive with the reciprocal antibody in Western blot analysis. Thus, 3F5.4G6 recognizes the same PSMA protein as does 7E11.C5, but at different epitopes on essentially opposite ends of the molecule. These two antibodies are well suited for use in a sandwich immunoassay, either one as a capture or detection antibody. Current work on this is underway. This report also confirms that 7E11.C5 Western blots for PSMA are negative with normal human brain tissue. The monoclonal antibody 9H10 does not react with 3F5.4G6 or with 7E11.C5 in studies conducted herein. Moreover, 3F5.4G6 reacts with PSMA found in the LNCaP cell line, but not DU-145 or PC3, which lack PSMA.

Sequence restrictions in T cell receptor beta-chains that have specificity for a self-peptide/Ld complex.

Cytotoxic T lymphocytes that react with a complex of Ld and a ubiquitous self-peptide derived from the enzyme alpha-ketoglutarate dehydrogenase (p2Ca, LSPFPFDL) can be readily elicited by the addition of synthetic peptide to cultures of BALB/c spleen cells. As with other Ld-restricted CTL, the p2Ca-specific cells use predominantly the V beta 8.3 region. In addition, the p2Ca-specific cells use almost exclusively one of three J beta gene segments. Selection for these J beta regions appears to be related to the presence of a glutamic acid residue that is encoded at the 5' end of the J beta and is present within the CDR3. As p2Ca does not contain a complementary charged residue, this finding may suggest that the beta-chain CDR3 from p2Ca-specific CTL contacts one of the five basic residues located on the Ld helices. Together, the results support the possibility that CDR1 and/or CDR2 (within V beta 8.3) and the CDR3 may each contact the Ld molecule. In contrast to the V beta and J beta regions, the V beta D beta J beta junctions and V alpha J alpha repertoires were diverse. The diversity could explain why p2Ca-specific CTL have relatively high precursor frequencies allowing them to be generated rapidly in primary cultures.

Generation of cytotoxic T-lymphocytes to a self-peptide/class I complex: a model for peptide-mediated tumor rejection.

Cytotoxic T-lymphocytes (CTL) typically recognize foreign peptides bound to class I products of the major histocompatibility complex. A function of CTL is to identify and eliminate tumor cells that bear inappropriately expressed peptide/class I complexes (i.e., mutated self-peptides or self-peptides that are expressed at abnormally high levels). The processes that result in tolerance to self-antigens can undermine the effectiveness of this system by deleting or inactivating T-cells that might potentially be reactive with tumor-associated antigens. To up-regulate the response to tumor antigens it will be useful to develop methods whereby CTL responses to specific self-peptides can be elicited without damage to normal tissue. In this report a CTL response was generated in BALB/c mice against the ubiquitous self-peptide p2Ca (LSPFPFDL), which binds to Ld and is derived from the mitochondrial enzyme alpha-ketoglutarate dehydrogenase. CTL derived in vitro recognize specifically the p2Ca/Ld complex and use V beta 8 regions predominantly. The cultured cells lysed target cells with lower levels of p2Ca than the levels used for induction. This result suggests that it may be possible to use peptides at high concentrations to elicit CTL react with endogenous levels of a peptide/class I complex. The in vivo potential of the response was demonstrated by the observation that BALB/c mice, coinjected with a syngeneic BALB/c myeloma and exogenous p2Ca, are able to reject the tumor. The p2Ca/Ld system may thus provide a model for evaluating the parameters for effective immunotherapy with tumor-associated peptides.

Sequence diversity of T cell receptor alpha chain transcripts from BALB/c thymus.

Most of the diversity in T cell receptor subunits resides in the region that is the equivalent of the CDR3 of immunoglobulins. In order to learn more about the relative contributions of the various mechanisms that generate this diversity we have analyzed the sequences of alpha chain transcripts from BALB/c thymus. The J alpha repertoire of BALB/c mice was examined by comparison of new J alpha sequences and previously published sequences. Among the 41 J alpha genes examined, most of the diversity is located at the 5' end, consistent with the notion that this region contacts the antigen. VJ junctional diversity was examined by sequencing various V alpha J alpha combinations derived from different stages of development. Deletion of bases from the ends of V and J genes does not occur with equal frequency. A greater number of bases were deleted on average from the ends of J genes. Bases were added at junctions frequently in isolates from adult animals, consistent with the presence of terminal deoxynucleotidyl transferase. However, there were short stretches of sequences at junctions which were also present at the 5' end of J genes. These findings extend recent observations that alpha chain genes use multiple mechanisms for generating diversity.