Biomedical subjects

Erwin S Schultz

Publications and source records attributed to Erwin S Schultz.

12 recordsLinked to original sources

PubMed2005

Differences in phenotype and function between spontaneously occurring melan-A-, tyrosinase- and influenza matrix peptide-specific CTL in HLA-A*0201 melanoma patients.

Melanoma-specific T cells can occur spontaneously or in response to vaccination or other therapies, but the frequency is much lower than observed in viral infections. The presence of tumor-specific T cells does not necessarily translate into clinical regressions for a variety of reasons such as an insufficient frequency, activation state or homing capacity of the T cells or escape strategies of the tumor. Having screened melanoma patients prior to inclusion in vaccination trials for spontaneous tumor-specific T cells either by Elispot or tetramer-staining, we have identified 3 patients with sufficient numbers of tumor-reactive T cells to more than 1 TAA and at least 1 virus-antigen to perform phenotypic and functional analysis directly ex vivo. These stage IV melanoma patients showed specific CTL against melan-A.A2, tyrosinase.A2 and influenza matrix peptide (IMP).A2 readily detectable in peripheral blood. T-cell receptor (TCR) staining using the tetramer technology was combined with phenotypic characterization and functional assays. In contrast to IMP-specific CTL, melanoma-specific CTL were predominantly terminally differentiated effector cells. However, analysis of melan-A- and tyrosinase-specific T-cell lines showed that only a part of the melanoma-specific CTL were able to lyse peptide-loaded target cells. Interestingly, the described phenotypic and functional differences of melan-A- and tyrosinase-specific CTL appeared not only between patients but were also evident within patients, suggesting that the immune response against various tumor antigens is regulated independently.

Adult↗

PubMed2005

Optimizing the exogenous antigen loading of monocyte-derived dendritic cells.

Dendritic cell (DC) vaccination, i.e. the adoptive transfer of antigen-loaded DC, is still at an early stage and requires standardization. In this study, we investigated the exogenous loading of monocyte-derived DCs with HLA class I- and II-restricted peptides, as despite widespread use, little effort has been put into its pre-clinical validation. We found that only mature DCs (m-DC) but not immature DCs (im-DC) could be sufficiently loaded with exogenous class I-restricted peptides and were by far superior in expanding CD8(+) primary (Melan-A.A2 peptide-specific) and recall [Influenza matrix peptide (IMP) A2-specific] T cell responses. Primary stimulation with peptide-loaded im-DCs even down-regulated antigen-specific T cell responses. Our results indicate that stimulation with m-DCs is superior in terms of quantity and quality compared with im-DCs, supporting their preferred use in clinical DC trials. Loading of m-DCs with high (10 microM) concentrations generated clearly more Melan-A effectors than loading with 1 or 0.1 microM without any negative effect on the quality (affinity) of the resulting T cells. In contrast to the findings with the Melan-A peptide loading with 10 microM IMP was counter-productive, induced apoptosis and yielded fewer specific T cells of inferior affinity as compared with loading with 1 or 0.1 microM. In sharp contrast to the situation for HLA class I, much higher levels and longer half-lives of peptide-HLA class II complexes were obtainable upon loading of im-DCs with exogenous peptide, but m-DCs were functionally preferable to induce T(h)1 responses in vitro. Another surprising finding was that, while presentation to T cells upon simultaneous loading of several peptides with highly varying affinities and competing for the same class I or II molecule was possible, in priming experiments peptide competition clearly inhibited T cell induction. Although peptides will obviously vary in their individual properties, our study clearly points to some important principles that should be taken into account.

Antibodies↗

PubMed2005

Generation of an optimized polyvalent monocyte-derived dendritic cell vaccine by transfecting defined RNAs after rather than before maturation.

Transfection with RNA is an attractive method of Ag delivery to dendritic cells (DCs), but has not yet been standardized. We describe in this study the methods to efficiently generate an optimized mature monocyte-derived DC vaccine at clinical scale based on the electroporation of several RNAs either into immature DC followed by maturation or, alternatively, directly into mature DCs, which has not been possible so far with such high efficiency. Electroporation of DCs resulted in high yield, high transfection efficiency (>90%), and high migration capacity. Intracellular staining allowed the study of the expression kinetics of Ags encoded by the transfected RNAs (MelanA, MAGE-3, and survivin) and a validation of the vaccine (>/=90% transfection efficiency). Expression of all three Ags peaked 3-4 h after electroporation in DC transfected either before or after maturation, but decreased differently. The DC vaccine can also be cryopreserved and nevertheless retains its viability, stimulatory capacity as well as migratory activity. In addition, we uncover that DC transfected after rather than before maturation appear to be preferable vaccines not only from a production point of view but also because they appear to be immunologically superior for CTL induction in sharp contrast to common belief. DCs transfected after maturation not only more effectively generate and present the Mage-3.A1 and MelanA.A2.1 epitopes to T cell clones, but they even are superior in priming to the standard proteasome-dependent MelanA.A2.1 wild-type prototype tumor epitope, both in terms of T cell expansion and effector function on a per cell basis.

Antigen Presentation↗

PubMed2005

Artesunate in the treatment of metastatic uveal melanoma--first experiences.

Artesunate (ART) is a derivative of artemisinin, the active principle of the Chinese herb Artemisia annua L. Artesunate is approved for the treatment of multidrug-resistant malaria and has an excellent safety profile. It has been shown that Artesunate, apart from its anti-malarial activity, has cytotoxic effects on a number of human cancer cell lines, including leukemia, colon cancer and melanoma. We report on the first long-term treatment of two cancer patients with ART in combination with standard chemotherapy. These patients with metastatic uveal melanoma were treated on a compassionate-use basis, after standard chemotherapy alone was ineffective in stopping tumor growth. The therapy-regimen was well tolerated with no additional side effects other than those caused by standard chemotherapy alone. One patient experienced a temporary response after the addition of ART to Fotemustine while the disease was progressing under therapy with Fotemustine alone. The second patient first experienced a stabilization of the disease after the addition of ART to Dacarbazine, followed by objective regressions of splenic and lung metastases. This patient is still alive 47 months after first diagnosis of stage IV uveal melanoma, a situation with a median survival of 2-5 months. Despite the small number of treated patients, ART might be a promising adjuvant drug for the treatment of melanoma and possibly other tumors in combination with standard chemotherapy. Its good tolerability and lack of serious side effects will facilitate prospective randomized trials in the near future.

Aged↗

PubMed2004

Functional analysis of tumor-specific Th cell responses detected in melanoma patients after dendritic cell-based immunotherapy.

Recently, we have demonstrated that tumor-specific CD4+ Th cell responses can be rapidly induced in advanced melanoma patients by vaccination with peptide-loaded monocyte-derived dendritic cells. Most patients showed a T cell reactivity against a melanoma Ag 3 (MAGE-3) peptide (MAGE-3(243-258)), which has been previously found to be presented by HLA-DP4 molecules. To analyze the functional and specificity profile of this in vivo T cell response in detail, peptide-specific CD4+ T cell clones were established from postvaccination blood samples of two HLA-DP4 patients. These T cell clones recognized not only peptide-loaded stimulator cells but also dendritic cells loaded with a recombinant MAGE-3 protein, demonstrating that these T cells were directed against a naturally processed MAGE-3 epitope. The isolated CD4+ Th cells showed a typical Th1 cytokine profile upon stimulation. From the first patient several CD4+ T cell clones recognizing the antigenic peptide used for vaccination in the context of HLA-DP4 were obtained, whereas we have isolated from the second patient CD4+ T cell clones which were restricted by HLA-DQB1*0604. Analyzing a panel of truncated peptides revealed that the CD4+ T cell clones recognized different core epitopes within the original peptide used for vaccination. Importantly, a DP4-restricted T cell clone was stimulated by dendritic cells loaded with apoptotic or necrotic tumor cells and even directly recognized HLA class II- and MAGE-3-expressing tumor cells. Moreover, these T cells exhibited cytolytic activity involving Fas-Fas ligand interactions. These findings support that vaccination-induced CD4+ Th cells might play an important functional role in antitumor immunity.

Amino Acid Sequence↗

PubMed2003

Antigen loading of dendritic cells with whole tumor cell preparations.

Dendritic cells (DC) based vaccinations have been widely used for the induction of anti-tumoral immunity in clinical studies. Antigen loading of DC with whole tumor cell preparations is an attractive method whenever tumor cell material is available. In order to determine parameters for the loading procedure, we performed dose finding and timing experiments. We found that apoptotic and necrotic melanoma cells up to a ratio of one-to-one, equivalent to 1mg/ml protein per 1 x 10(6) DC, can be added to monocyte derived DC without effecting DC recovery extensively. Using the isolated protein content of tumor cells (lysate) as a parameter, up to 5 mg/ml protein per 1 x 10(6) DC can be added. To achieve significant protein uptake at least 1 mg/ml of protein have to be added for more than 24 h as tested with FITC-labelled ovalbumin. Maturation inducing cytokines can be added simultaneously with the tumor cell preparations to immature DC without affecting the uptake. Furthermore, we tested the feasibility of cryopreservation of loaded and matured DC to facilitate the generation of ready to use aliquots. DC were cryopreserved in a mix of human serum albumin, DMSO and 5% glucose. After thawing, surface expression of molecules indicating the mature status (CD83, costimulatory and MHC molecules), was found to be unaltered. Furthermore, cryopreserved DC kept the capability to stimulate allogenic T-cell proliferation in mixed leukocyte reactions at full level. Loaded and matured DC pulsed with influenza matrix peptide (IMP) retained the capacity to induce the generation of IMP-specific cytotoxic T-lymphocytes after cryopreservation as measured by ELISPOT and tetramer staining. The expression of the chemokine receptor CXCR-4 and CCR-7 remained unaltered during cryopreservation and the migratory responsiveness towards MIP-3beta was unaltered as measured in a migration assay. Thus we conclude that the large scale loading and maturation of DC with whole tumor cell preparations can be performed in a single session. These data will facilitate the clinical application of DC loaded with whole tumor cell preparations.

Antigens, CD↗

PubMed2002

Rapid induction of tumor-specific type 1 T helper cells in metastatic melanoma patients by vaccination with mature, cryopreserved, peptide-loaded monocyte-derived dendritic cells.

There is consensus that an optimized cancer vaccine will have to induce not only CD8+ cytotoxic but also CD4+ T helper (Th) cells, particularly interferon (IFN)-gamma-producing, type 1 Th cells. The induction of strong, ex vivo detectable type 1 Th cell responses has not been reported to date. We demonstrate now that the subcutaneous injection of cryopreserved, mature, antigen-loaded, monocyte-derived dendritic cells (DCs) rapidly induces unequivocal Th1 responses (ex vivo detectable IFN-gamma-producing effectors as well as proliferating precursors) both to the control antigen KLH and to major histocompatibility complex (MHC) class II-restricted tumor peptides (melanoma-antigen [Mage]-3.DP4 and Mage-3.DR13) in the majority of 16 evaluable patients with metastatic melanoma. These Th1 cells recognized not only peptides, but also DCs loaded with Mage-3 protein, and in case of Mage-3DP4-specific Th1 cells IFN-gamma was released even after direct recognition of viable, Mage-3-expressing HLA-DP4+ melanoma cells. The capacity of DCs to rapidly induce Th1 cells should be valuable to evaluate whether Th1 cells are instrumental in targeting human cancer and chronic infections.

Adult↗

PubMed2002

The production of a new MAGE-3 peptide presented to cytolytic T lymphocytes by HLA-B40 requires the immunoproteasome.

By stimulating human CD8(+) T lymphocytes with autologous dendritic cells infected with an adenovirus encoding MAGE-3, we obtained a cytotoxic T lymphocyte (CTL) clone that recognized a new MAGE-3 antigenic peptide, AELVHFLLL, which is presented by HLA-B40. This peptide is also encoded by MAGE-12. The CTL clone recognized MAGE-3--expressing tumor cells only when they were first treated with IFN-gamma. Since this treatment is known to induce the exchange of the three catalytic subunits of the proteasome to form the immunoproteasome, this result suggested that the processing of this MAGE-3 peptide required the immunoproteasome. Transfection experiments showed that the substitution of beta5i (LMP7) for beta5 is necessary and sufficient for producing the peptide, whereas a mutated form of beta5i (LMP7) lacking the catalytically active site was ineffective. Mass spectrometric analyses of in vitro digestions of a long precursor peptide with either proteasome type showed that the immunoproteasome produced the antigenic peptide more efficiently, whereas the standard proteasome more efficiently introduced cleavages destroying the antigenic peptide. This is the first example of a tumor-specific antigen exclusively presented by tumor cells expressing the immunoproteasome.

Adenoviridae↗

PubMed2002

Tumor-specific shared antigenic peptides recognized by human T cells.

The first tumor-specific shared antigens and the cancer-germline genes that code for these antigens were identified with antitumor cytolytic T lymphocytes obtained from cancer patients. A few HLA class I-restricted antigenic peptides were identified by this 'direct approach'. A large set of additional cancer-germline genes have now been identified by purely genetic approaches or by screening tumor cDNA expression libraries with the serum of cancer patients. As a result, a vast number of sequences are known that can code for tumor-specific shared antigens, but most of the encoded antigenic peptides have not yet been identified. We review here recent 'reverse immunology' approaches for the identification of new antigenic peptides. They are based on in vitro stimulation of naive T cells with dendritic cells that have either been loaded with a cancer-germline protein or that have been transduced with viruses carrying cancer-germline coding sequences. These approaches have led to the identification of many new antigenic peptides presented by class I or class II molecules. We also describe some aspects of the processing and presentation of these antigenic peptides.

Amino Acid Sequence↗

PubMed2002

Segmental neurofibromatosis.

Segmental neurofibromatosis is characterised by a limited, segmental distribution of cutaneous neurofibromatosis type 1 (NF1) lesions. It has been suggested that segmental NF results from a postzygotic NF1 gene mutation, and, recently, this hypothesis has been proven in a patient with regionally distributed café-au-lait (CAL) spots and freckles by demonstrating an NF1 microdeletion restricted to fibroblasts cultured from CAL spots. We describe here a patient with segmental NF in which we could not demonstrate any NF1 gene mutation in fibroblasts cultured from neurofibromas by use of the protein truncation test, enzymatic mutation detection and fluorescence in situ hybridisation. These data are in line with the concept that NF1 mutations in Schwann cells, but not in fibroblasts, correlate with neurofibroma formation.

Humans↗

PubMedDate not supplied

Mycosis fungoides palmaris et plantaris--an unusual variant of cutaneous T-cell lymphoma.

Mycosis fungoides (MF) represents a low-risk, cutaneous, non-Hodgkin, T-cell lymphoma with a wide spectrum of clinicopathological manifestations and therefore may mimic a number of other dermatoses. Sometimes the clinical diversity makes the diagnosis of MF, and especially its atypical forms, challenging. We report on an 18-year old male patient, who had been previously diagnosed with palmoplantar eczema. Clinical, histopathological, immunohistochemical and molecular findings revealed an atypical case of MF.

Adolescent↗

PubMedDate not supplied

Late syphilis mimicking a pseudolymphoma of the skin.

After a period of declining incidence of syphilis in most of Western Europe until the late 1990s, reports about an increasing trend have been published recently. In contrast to the rising incidence of early syphilis, cases of late (tertiary) syphilis are rarely seen in developed countries. In this report, we describe a 54-year-old patient with infiltrated erythematous plaques on his forehead and neck that histologically revealed a dense lymphocytic cell infiltrate with numerous plasma cells. The serologic examination was characteristic of syphilis and in conjunction with the clinical presentation and the patient's history led to the diagnosis of tertiary syphilis. The diagnosis of this late stage of syphilis can be difficult as clinical pictures can be misleading. Peculiar skin lesions should always remind clinicians of this infectious disease which has still to be considered in differential diagnoses in dermatology.

Diagnosis, Differential↗