Vincent Brichard

Tumor regressions observed in patients with metastatic melanoma treated with an antigenic peptide encoded by gene MAGE-3 and presented by HLA-A1.

Thirty‐nine tumor‐bearing patients with metastatic melanoma were treated with 3 subcutaneous injections of the MAGE‐3.A1 peptide at monthly intervals. No significant toxicity was observed. Of the 25 patients who received the complete treatment, 7 displayed significant tumor regressions. All but one of these regressions involved cutaneous metastases. Three regressions were complete and 2 of these led to a disease‐free state, which persisted for more than 2 years after the beginning of treatment. No evidence for a cytolytic T lymphocyte (CTL) response was found in the blood of the 4 patients who were analyzed, including 2 who displayed complete tumor regression. Our results suggest that injection of the MAGE‐3.A1 peptide induced tumor regression in a significant number of the patients, even though no massive CTL response was produced. Int. J. Cancer 80:219–230, 1999.

Genes Coding for Tumor Antigens Recognized by Cytolytic T Lymphocytes

Presuming that T lymphocytes might be able to eradicate cancer cells as effectively as they kill virus-infected celts, tumor immunologists have been trying to identify specific target antigens displayed by cancer cells that could make them recognizable to cytolytic T lymphocytes (CTL). During the last few years several mouse and human tumor antigens recognized by CTL have been identified at the molecular level. This review focuses on the tumor antigens identified in our laboratory. Interestingly, none of these antigens arises from the product of known oncogenes or tumor-supressor genes. The antigens fall into three categories: antigens encoded by genes expressed in tumors but not in most normal tissues, differentiation antigens and antigens derived from mutated genes that are expressed ubiquitously.

A monoclonal cytolytic T-lymphocyte response observed in a melanoma patient vaccinated with a tumor-specific antigenic peptide encoded by gene MAGE-3

Vaccination of melanoma patients with tumor-specific antigens recognized by cytolytic T lymphocytes (CTL) produces significant tumor regressions in a minority of patients. These regressions appear to occur in the absence of massive CTL responses. To detect low-level responses, we resorted to antigenic stimulation of blood lymphocyte cultures in limiting dilution conditions, followed by tetramer analysis, cloning of the tetramer-positive cells, and T-cell receptor (TCR) sequence analysis of the CTL clones that showed strict specificity for the tumor antigen. A monoclonal CTL response against a MAGE-3 antigen was observed in a melanoma patient, who showed partial rejection of a large metastasis after treatment with a vaccine containing only the tumor-specific antigenic peptide. Tetramer analysis after in vitro restimulation indicated that about 1/40,000 postimmunization CD8+ blood lymphocytes were directed against the antigen. The same TCR was present in all of the positive microcultures. TCR evaluation carried out directly on blood lymphocytes by PCR amplification led to a similar frequency estimate after immunization, whereas the TCR was not found among 2.5 × 106 CD8+ lymphocytes collected before immunization. Our results prove unambiguously that vaccines containing only a tumor-specific antigenic peptide can elicit a CTL response. Even though they provide no information about the effector mechanisms responsible for the observed reduction in tumor mass in this patient, they would suggest that low-level CTL responses can initiate tumor rejection.

Adjuvant MAGE-A3 Immunotherapy in Resected Non–Small-Cell Lung Cancer: Phase II Randomized Study Results

PURPOSEnThe MAGE-A3 protein is expressed in approximately 35% of patients with resectable non-small-cell lung cancer (NSCLC). Several immunization approaches against the MAGE-A3 antigen have shown few, but often long-lasting, clinical responses in patients with metastatic melanoma.nnnPATIENTS AND METHODSnA double-blind, randomized, placebo-controlled phase II study was performed assessing clinical activity, immunologic response, and safety following immunization with recombinant MAGE-A3 protein combined with an immunostimulant (13 doses over 27 months) in completely resected MAGE-A3-positive stage IB to II NSCLC. The primary end point was disease-free interval (DFI).nnnRESULTSnPatients were randomly assigned to either MAGE-A3 immunotherapeutic (n = 122) or placebo (n = 60). After a median postresection period of 44 months, recurrence was observed in 35% of patients in the MAGE-A3 arm and 43% in the placebo arm. No statistically significant improvement in DFI (hazard ratio [HR], 0.75, 95% CI, 0.46 to 1.23; two-sided P = .254), disease-free survival (DFS; HR, 0.76; 95% CI, 0.48 to 1.21; P = .248), or overall survival (HR, 0.81; 95% CI, 0.47 to 1.40; P = .454) was observed. Corresponding analysis after a median of 70 months of follow-up revealed a similar trend for DFI and DFS. All patients receiving the active treatment showed a humoral immune response to the MAGE-A3 antigen, although no correlation was observed with outcome. No significant toxicity was observed.nnnCONCLUSIONnIn this early development study with a limited number of patients, postoperative MAGE-A3 immunization proved to be feasible with minimal toxicity. These results are being investigated further in a large phase III study.

Predictive Gene Signature in MAGE-A3 Antigen-Specific Cancer Immunotherapy

PURPOSEnTo detect a pretreatment gene expression signature (GS) predictive of response to MAGE-A3 immunotherapeutic in patients with metastatic melanoma and to investigate its applicability in a different cancer setting (adjuvant therapy of resected early-stage non-small-cell lung cancer [NSCLC]).nnnPATIENTS AND METHODSnPatients were participants in two phase II studies of the recombinant MAGE-A3 antigen combined with an immunostimulant (AS15 or AS02B). mRNA from melanoma biopsies was analyzed by microarray analysis and quantitative polymerase chain reaction. These results were used to identify and cross-validate the GS, which was then applied to the NSCLC data.nnnRESULTSnIn the patients with melanoma, 84 genes were identified whose expression was potentially associated with clinical benefit. This effect was strongest when the immunostimulant AS15 was included in the immunotherapy (hazard ratio [HR] for overall survival, 0.37; 95% CI, 0.13 to 1.05; P = .06) and was less strong with the other immunostimulant AS02B (HR, 0.84; 95% CI, 0.36 to 1.97; P = .70). The same GS was then used to predict the outcome for patients with resected NSCLC treated with MAGE-A3 plus AS02B; actively treated GS-positive patients showed a favorable disease-free interval compared with placebo-treated GS-positive patients (HR, 0.42; 95% CI, 0.17 to 1.03; P = .06), whereas among GS-negative patients, no such difference was found (HR, 1.17; 95% CI, 0.59 to 2.31; P = .65). The genes identified were mainly immune related, involving interferon gamma pathways and specific chemokines, suggesting that their pretreatment expression influences the tumors immune microenvironment and the patients clinical response.nnnCONCLUSIONnAn 84-gene GS associated with clinical response for MAGE-A3 immunotherapeutic was identified in metastatic melanoma and confirmed in resected NSCLC.

Gene signature in melanoma associated with clinical activity: a potential clue to unlock cancer immunotherapy.

Immunotherapeutic approaches for melanoma and other cancers can impart profound clinical benefit but only for a subset of patients. Interpatient heterogeneity could, in principle, be due to somatic differences in the tumor between individuals or alternatively be accounted for distinct germline polymorphisms in immunoregulatory genes of the host. Analysis of these possibilities has been initiated by investigating gene expression profiling of the tumor microenvironment in the context of clinical trials of cancer vaccines. Distinct gene expression profiles have been identified on pretreatment biopsies that are associated with a positive or negative clinical outcome. These observations suggest that such profiling might be useful as a predictive biomarker for clinical benefit from vaccines and other immunotherapy approaches, and analysis of specific gene products has begun to suggest new therapeutic interventions to overcome mechanisms of tumor resistance.

Tumoral and Immunologic Response After Vaccination of Melanoma Patients With an ALVAC Virus Encoding MAGE Antigens Recognized by T Cells

PURPOSEnTo evaluate the toxicity, antitumoral effectiveness, and immunogenicity of repeated vaccinations with ALVAC miniMAGE-1/3, a recombinant canarypox virus containing a minigene encoding antigenic peptides MAGE-3(168-176) and MAGE-1(161-169), which are presented by HLA-A1 and B35 on tumor cells and can be recognized by cytolytic T lymphocytes (CTLs).nnnMATERIALS AND METHODSnThe vaccination schedule comprised four sequential injections of the recombinant virus, followed by three booster vaccinations with the MAGE-3(168-176) and MAGE-1(161-169) peptides. The vaccines were administered, both intradermally and subcutaneously, at 3-week intervals.nnnRESULTSnForty patients with advanced cancer were treated, including 37 melanoma patients. The vaccines were generally well tolerated with moderate adverse events, consisting mainly of transient inflammatory reactions at the virus injection sites. Among the 30 melanoma patients assessable for tumor response, a partial response was observed in one patient, and disease stabilization in two others. The remaining patients had progressive disease. Among the patients with stable or progressive disease, five showed evidence of tumor regression. A CTL response against the MAGE-3 vaccine antigen was detected in three of four patients with tumor regression, and in only one of 11 patients without regression.nnnCONCLUSIONnRepeated vaccination with ALVAC miniMAGE-1/3 is associated with tumor regression and with a detectable CTL response in a minority of melanoma patients. There is a significant correlation between tumor regression and CTL response. The contribution of vaccine-induced CTL in the tumor regression process is discussed in view of the immunologic events that could be analyzed in detail in one patient.

Diet- and diabetes-induced changes of ob gene expression in rat adipose tissue

ob gene regulation is as yet unknown. We first examined whether the ob gene is under physiological control by the nutritional state. Fasting produced a sharp (95%) decrease of ob mRNA in epididymal and inguinal fat pads from 24 h onward. Refeeding rapidly (3–6 h) re‐induced ob gene expression and corrected it within 24 h. Similar changes in fatty acid synthase (FAS) and GLUT4 mRNAs were observed, whereas phosphoenolpyruvate car☐ykinase (PEPCK) mRNA showed an opposite evolution. We next examined the potential role of insulin. In adipose tissue of streptozotocin‐diabetic rats, ob mRNA levels were decreased by 80%. Insulin treatment (4 days) only marginally increased ob mRNA, but restored euglycemia and overcorrected FAS, GLUT4 and PEPCK expression. In conclusion, we provide evidence for a physiological regulation of ob gene by variations in the nutritional state. We also show that ob expression is impaired in streptozotocin‐diabetic rats and only slightly restored by insulin treatment, which suggests that ob gene is not or only minimally regulated by the hormone.

Selection of Immunostimulant AS15 for Active Immunization With MAGE-A3 Protein: Results of a Randomized Phase II Study of the European Organisation for Research and Treatment of Cancer Melanoma Group in Metastatic Melanoma

PURPOSEnActive immunization against the tumor-specific MAGE-A3 antigen is followed by a few but impressive and durable clinical responses. This randomized phase II trial evaluated two different immunostimulants combined with the MAGE-A3 protein to investigate whether a more robust and persistent immune response could be associated with increased clinical benefit.nnnPATIENTS AND METHODSnPatients with MAGE-A3-positive stage III or IV M1a melanoma were randomly assigned to receive the MAGE-A3 protein combined either with AS02B or with AS15 immunostimulant. Clinical end points were toxicity and rates of objective clinical responses, progression-free survival (PFS), and overall survival (OS).nnnRESULTSnSeventy-five patients were treated, with 36 eligible patients per arm. Both treatments were well tolerated. In the AS15 arm, four objective responses were observed (three complete responses and one partial response) versus one partial response in the AS02B arm. In the AS15 and AS02B arms, the PFS rates after 6 months were 25% and 14%, respectively; and the median OS times were 33 months and 19.9 months, respectively, with a median observation period of 48 months. Antibodies against MAGE-A3, found in all patients, showed three-fold higher titers in the AS15 arm. The anti-MAGE-A3 cellular response was also more pronounced in the AS15 arm.nnnCONCLUSIONnIn the MAGE-A3+AS15 arm, clinical activity was higher and the immune response more robust. Therefore, the AS15 immunostimulant was selected for combination with the MAGE-A3 protein in phase III trials.

Toll-like receptor 4 stimulation with the detoxified ligand monophosphoryl lipid A improves Alzheimer’s disease-related pathology

Alzheimer’s disease (AD) is the most common cause of dementia worldwide. The pathogenesis of this neurodegenerative disease, currently without curative treatment, is associated with the accumulation of amyloid β (Aβ) in brain parenchyma and cerebral vasculature. AD patients are unable to clear this toxic peptide, leading to Aβ accumulation in their brains and, presumably, the pathology associated with this devastating disease. Compounds that stimulate the immune system to clear Aβ may therefore have great therapeutic potential in AD patients. Monophosphoryl lipid A (MPL) is an LPS-derived Toll-like receptor 4 agonist that exhibits unique immunomodulatory properties at doses that are nonpyrogenic. We show here that repeated systemic injections of MPL, but not LPS, significantly improved AD-related pathology in APPswe/PS1 mice. MPL treatment led to a significant reduction in Aβ load in the brain of these mice, as well as enhanced cognitive function. MPL induced a potent phagocytic response by microglia while triggering a moderate inflammatory reaction. Our data suggest that the Toll-like receptor 4 agonist MPL may be a treatment for AD.