Biomedical subjects
Publications and source records attributed to D Schadendorf.
The protein kinase inhibitor imatinib has been approved as an efficient anticancer drug with common but mild cutaneous toxicities. We here report on two out of four melanoma patients treated with high-dose imatinib presenting with severe and strongly dose-dependent skin eruptions, suggesting a cutaneous reactivity pattern different from allergic hypersensitivity.
BACKGROUND: Cancer-testis antigens exemplify a growing number of tumour antigens which are expressed in a variety of malignancies, but not in normal tissues other than germ cells, primarily those of the testis. OBJECTIVES: To investigate the humoral response to known cancer-testis antigens in melanoma patients. METHODS: We used phage clones coding for seven different melanoma antigens MAGE-A or LAGE-1A proteins. These clones were isolated using the newly developed DNA hybridization analysis of recombinantly expressed cDNA libraries (HYREX) approach. HYREX combines the advantage of a nonradioactive library screening method with the possibility of subsequently analysing the serological response to the recombinant proteins. We isolated clones coding for MAGE-A1, -A3, -A4b, -A6, -A9 and -A12, as well as LAGE-1A. Additionally, we correlated gene expression and seroreactivity. RESULTS: Between 13% and 27% of sera (n = 15) were reactive against individual tumour antigens. We found the presence of specific antibodies was, with only two exceptions, generally correlated with mRNA expression of the antigen within cell lines derived from the same patient. While cross-reactivity of patients' IgG might play a role in these cases, antibodies from patients' sera were able to distinguish even the closely related MAGE-A3 and -A6. In general, the mRNA expression frequency was higher than the detected IgG responses. CONCLUSIONS: Antibody recognition of specific tumour antigens by patients' sera may be used for evaluating the possible immunogenicity of new antigens; serological tests could be used for tumour monitoring purposes.
Malignant melanoma is a tumor of increasing incidence. Patients with early diagnosed and surgically excised primary tumors have a high probability to be completely cured. In contrast, the prognosis of patients with distant organ metastases is still extremely poor despite a variety of therapeutical efforts that have been evaluated in numerous clinical trials. Cytotoxic treatment using single agents as well as polychemotherapy revealed only temporary clinical responses but no improvement of patients' overall survival. Likewise, therapy regimens combining cytostatics with cytokines showed no substantial benefit compared to cytostatics alone. Due to these disappointing results, recent research strategies focused on the elucidation of the mechanisms responsible for the complex therapy resistance of melanoma. New treatment concepts have been developed to overcome some of these mechanisms. Another promising approach is the development of advanced vaccination strategies. This paper reviews a selection of innovative therapeutic strategies that are currently tested in clinical trials; in particular pretherapeutic chemosensitivity testing, chemosensitization by downregulation of bcl-2, vaccination with autologous peptide-pulsed dendritic cells and immunomodulation by histamine.
The main goal of follow-up examinations of patients with cutaneous melanomas is early diagnosis of developing recurrent or second tumors. Timely recognition of recurrence has a relevant effect on the prognosis. In 80% of the cases recurrences are diagnosed for the first time in the course of follow-up when follow-up procedures are carried out systematically. Most recurrences are discovered during a physical examination. Sonographic examination of the lymph nodes is also a very sensitive method which can lead to the diagnosis of locoregional recurrences before they are palpable. Further image-producing techniques are not necessary for surveillance of primary tumors. In contrast to former recommendations, new research indicates that less extensive diagnostic procedures are sufficient in primary tumor stages, especially when the tumor thickness of the melanoma is less than 1 mm. Consequently, costs can be reduced considerably without additional risk to the patient.
Malignant melanoma of the uvea is remarkable for purely haematogenous dissemination and its tendency to metastasise to the liver. Although the liver is involved in up to 95% of patients, 50% of these also develop extrahepatic metastases, most often in the lungs, bone, skin, and brain. The only effective treatments reported to date relied on hepatic arterial chemoembolisation or -perfusion. The objective of this study was to establish a therapy protocol addressing patients with both sole liver involvement and systemic disease. Forty-eight patients with metastatic ocular melanoma received fotemustine 100 mg m(-2) either as 60-min infusion into the hepatic artery or as 15-min infusion via a peripheral vein, depending on the metastatic sites involved, i.e., restriction to the liver or hepatic together with extrahepatic disease. For the first treatment cycle this infusion was repeated after one week. For all cycles, subsequent to a three week resting period, patients received an immunotherapy consisting of subcutaneous interleukin 2 and interferon alpha(2). Although objective responses were more frequent within the cohort receiving intraarterial fotemustine (21.7 vs 8%), this difference did not translate into a significant benefit in overall survival, i.e., 369 and 349 days, respectively. Of note, this overall survival is much longer than that repeatedly reported for stage IV uveal melanoma not treated with fotemustine, suggesting a therapeutic activity of this cytostatic drug even after systemic administration.
In a German cohort of 661 melanoma patients the performance, costs and survival benefits of staging methods (history and physical examination; chest X-ray; ultrasonography of the abdomen; high resolution sonography of the peripheral lymph nodes) were assessed at initial staging and during follow-up of stage I/II+III disease. At initial staging, 74% (23 out of 31) of synchronous metastases were first detected by physical examination followed by sonography of the lymph nodes revealing 16% (5 out of 31). Other imaging methods were less efficient (Chest X-ray: one out of 31; sonography of abdomen: two out of 31). Nearly 24% of all 127 first recurrences and 18% of 73 second recurrences developed in patients not participating in the follow-up programme. In follow-up patients detection of first or second recurrence were attributed to history and physical examination on a routine visit in 47 and 52% recurrences, respectively, and to routine imaging procedures in 21 and 17% of cases, respectively. Lymph node sonography was the most successful technical staging procedure indicating 13% of first relapses, but comprised 24% of total costs of follow-up in stage I/II. Routine imaging comprised nearly 50% of total costs for follow-up in stage I/II and in stage III. The mode of detecting a relapse ('patient vs. doctor-diagnosed' or 'symptomatic vs asymptomatic') did not significantly influence patients overall survival. Taken together, imaging procedures for routine follow-up in stage I/II and stage III melanoma patients were inefficient and not cost-efficient.
The aim of this study was to define prognostic factors for survival, and especially for long-term survival in a mature data-set of patients with stage IV melanoma treated within a randomised trial of cytokine-based protocols. Long-term follow-up data on patients enrolled into a European Organization for Research and Treatment of Cancer (EORTC) trial comparing interferon-alpha (IFNalpha) plus interleukin-2 (IL-2) with or without cisplatin were collected. Univariate and multivariate Cox regression analyses were performed to define prognostic factors for survival. The characteristics of patients alive at 2 and 5 years after randomisation were compared with the entire cohort using the chi(2) test. The minimum potential follow-up of the 131 evaluable patients was 5 years. 18 patients (14%) were alive 2 years after randomisation, and 11 (8%) 5 years after randomisation. Pretreatment performance status (PS), serum lactate dehydrogenase (LDH) and tumour mass were significant predictors for survival, whereas site of metastases and number of sites were non-significant. PS and LDH were the only independent prognostic factors. All except 1 patient alive at 2 and 5 years had a pretreatment PS of 100%, and only three long-term survivors had elevated pretreatment LDH. There was no association between the site of metastases and long-term survival. Response to treatment was a major predictor for long-term survival, whereas addition of cisplatin did not impact upon overall survival probability or on long-term survival. The probability of long-term survival in stage IV melanoma patients after IL-2-based treatments is governed by pretreatment PS, serum LDH and response to treatment. Site of metastases, the basis for the M-subcategories of the new AJCC staging system, was not informative in this study.
We report on a 74-year-old female patient with a primary cutaneous CD20+, diffuse large cell B-cell lymphoma of the lower leg resembling a chronic non-healing leg ulcer. There was no evidence of systemic involvement on computed tomography (CT) scans of the chest, abdomen and pelvis; a slightly enlarged lymph node in the right groin showed dermatopathic lymphadenopathy on histology and immunohistochemistry. Involvement of the bone marrow and peripheral blood was ruled out by punch biopsy and fluorescent activated cell sorter (FACS) analysis of the blood, respectively. Therapeutic anti-CD20 monoclonal antibody rituximab was given at 375 mg m(-2) i.v. once weekly for 7 weeks, without adverse effects, resulting in a minor improvement in the centre of the ulcerated tumour. Unfortunately, the response was not maintained, and after 7 weeks of treatment the patient started to develop new tumour lesions at the border of the ulcer. Local radiotherapy was started and combined photon and electron beam irradiation induced complete remission of the B-cell lymphoma.
We describe a patient with a 9-year history of generalized eruptive keratoacanthoma (KA) of the Grzybowski type whose multiple skin lesions showed steady progression, resulting in a sclerotic, mask-like facial expression and ectropion. Eleven tumour biopsies representing lesions of different stages and localizations (erupting and regressing KAs, biopsies from non-involved light-protected and light-exposed skin, dermatosclerosis and squamous cell carcinomas) were analysed for human papillomavirus (HPV) sequences using a polymerase chain reaction approach capable of detecting the majority of all presently known HPV genotypes. None of the biopsy specimens proved to be HPV-positive, although HPV was detected in weakly and heavily affected control specimens by the method applied. These findings suggest an HPV-independent aetiology of this rare type of multiple KA.
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Advanced malignant melanoma has a poor prognosis since chemotherapy is mostly ineffective due in part to the intrinsic and/or extrinsic resistance of melanoma cells to systemic treatment with anti-neoplastic agents. The reasons for the chemoresistant phenotype are unknown. The relevance of well-analyzed drug-resistance mechanisms, e.g., intracellular/extracellular transport and induction of certain enzyme systems, is reviewed. Most anti-cancer drugs kill susceptible cells through induction of apoptosis. Therefore, it appears that differences in the apoptotic pathways which lead to apoptotic deficiency may account for the ability of some tumor cells to resist drug therapy. Human melanomas, which are characteristically drug-resistant, are more likely to have altered apoptotic pathways and fewer pro-apoptotic molecules. Tumor cells with these characteristics are seldom sensitive to drugs. The complexity of the molecular variants involved in signal transduction along apoptotic pathways suggests that the cell may have a variety of possibilities for regulating apoptosis and generating apoptotic deficiency. Thus, apoptosis and apoptotic deficiency should be analyzed to better clarify the mechanisms of melanoma resistance.
Multidrug resistance (MDR) phenotypes have been associated with the overexpression of various members of the superfamily of ATP binding cassette (ABC) transporters. Here we demonstrate that a member of the ABC-transporter family, the heterodimer 'transporter associated with antigen processing' (TAP), physiologically involved in major histocompatibility complex class I-restricted antigen presentation, is significantly overexpressed in the human gastric carcinoma cell line EPG85-257RNOV exhibiting a mitoxantrone-resistant phenotype. This tumor cell line shows an atypical MDR phenotype in the absence of 'P-glycoprotein' or 'MDR-associated protein' overexpression but with an enforced 'breast cancer resistance protein' expression level. Transfection of both TAP subunits encoding cDNA molecules, TAP1 and TAP2, into the drug-sensitive parental gastric carcinoma cell line EPG85-257P conferred a 3.3-fold resistance to mitoxantrone but not to alternative anti-neoplastic agents. Furthermore, cell clones transfected with both, but not singularly expressed TAP1 or TAP2, reduced cellular mitoxantrone accumulation. Taken together, the data suggest that the heterodimeric TAP complex possesses characteristics of a xenobiotic transporter and that the TAP dimer contributes to the atypical MDR phenotype of human cancer cells.
Resistance to drug treatment is a common observation in malignant melanoma. In order to analyze alterations in mRNA expression profiles associated with drug resistance in melanoma cells we previously established a panel of various drug-resistant cell variants derived from the human melanoma line MeWo and compared the mRNA expression profiles by a differential display technique. By that approach it could be demonstrated that the expression level of a mRNA encoded by a gene found to be mutated in non-syndromic hearing impairment, DFNA5 (ICERE-1), was distinctly decreased in the 33-fold etoposide-resistant melanoma cell line MeWo ETO 1. To evaluate the hypothesis that a decrease in DFNA5 mRNA expression level contributes to the acquired etoposide resistance phenotype exhibited by MeWo ETO 1 cells, this drug-resistant line was stably transfected with the DFNA5-encoding cDNA. Transfected clones showed a 30-35% reduced etoposide susceptibility by comparing the IC(25), IC(50) and IC(75) values of these clones with those displayed by the non-transfected, etoposide-resistant melanoma cell line MeWo ETO 1 and controls. Furthermore, etoposide exposure of stable DFNA5 transfectants resulted in an increase of caspase-3-mediated apoptotic events in DFNA5-transfected clones in comparison to MeWo ETO 1 cells and controls. The data therefore demonstrate that a decrease in DNFA5 mRNA expression level is associated with an increased etoposide resistance in melanoma cells due to an elevated cellular susceptibility to trigger a caspase-3-depending signal pathway leading to programmed cell death.
Acquired resistance to antineoplastic agents is a frequent obstacle in tumor therapy. Malignant melanoma cells are particularly well known for their unresponsiveness to chemotherapy; only about 30% of tumors exhibit a transient clinical response to treatment. In our study, we investigated the molecular mechanism of acquired resistance of melanoma cells (MeWo) to the chloroethylating drug fotemustine. Determination of O(6)-methylguanine-DNA methyltransferase (MGMT) activity showed that MeWo cells that acquired resistance to fotemustine upon repeated treatment with the drug display high MGMT activity, whereas the parental cell line had no detectable MGMT. The resistant cell lines exhibit cross-resistance to other O(6)-alkylating agents, such as N-methyl-N'-nitro-N-nitrosoguanidine. Acquired resistance to fotemustine was alleviated by treatment with the MGMT inhibitor O(6)-benzylguanine demonstrating that reactivation of MGMT is the main underlying cause of acquired alkylating drug resistance. As compared with control cells, both MGMT mRNA and MGMT protein were expressed at a high level in fotemustine resistant cells. Southern blot analysis proved that the MGMT gene was not amplified. There was also only an insignificant difference in the CpG methylation pattern of the MGMT promoter whereas a clear hypermethylation in the body of the gene was observed in fotemustine resistant cells. The conclusion that hypermethylation is responsible for reactivation of the MGMT gene gained support by the finding that MGMT activity significantly declined and cells reverted (partially) to the parental sensitive phenotype upon treatment with 5-azacytidine. This is the first report of acquired resistance to a chloroethylating antineoplastic drug of melanoma cells due to gene hypermethylation.
Cutaneous T-cell lymphomas (CTCL) are a group of skin neoplasms that originate from T lymphocytes and are difficult to treat in advanced stages. The present study is aimed at the identification of tumor-specific antigens from a human testis cDNA library using human sera known as the SEREX (serological identification of recombinantly expressed genes) approach. A cDNA library from normal testicle tissue was prepared and approximately 2 million recombinants were screened with sera from Sézary Syndrome and Mycosis fungoides patients. A total of 28 positive clones belonging to 15 different genes/ORFs were identified, including five hitherto unknown sequences. Whereas control sera did not react with most clones, 11-71% sera from CTCL patients were reactive against the identified clones. Expression analysis on 28 normal control and 17 CTCL tissues by reverse transcription-PCR (RT-PCR) and Northern blotting revealed seven ubiquitously distributed antigens, six differentially expressed antigens (several normal tissues were positive), and two tumor-specific antigens that were expressed only in testis and tumor tissues: (i) A SCP-1-like sequence, which has already been detected in various tumors, has been found in one CTCL tumor and four sera of CTCL patients reacted with various SCP-1-like clones and (ii) a new sequence named cTAGE-1 (CTCL-associated antigen 1) was detected in 35% of CTCL tumor tissues and sera of 6/18 patients reacted with this clone. The present study unravels CTCL-associated antigens independent of the T-cell receptor. The SCP-1-like gene and cTAGE-1 were shown to be immunogenic and immunologically tumor-specific and may therefore be candidates for immunotherapy targeting CTCL.
Dendritic cells (DC) are commonly viewed as the professional antigen-presenting cell. They capture antigens, migrate to appropriate lymphoid organs and initiate an antigen-specific CD4 and CD8 T cell response. Much is known about DC physiology, and it is now possible to culture, maintain and expand DC from different human sources, including hematopoietic progenitors in bone marrow and peripheral blood. Combined with the detection of an increasing number of tumor-associated antigens and T cell-recognized peptide epitopes, this has led to a new enthusiasm in the field of tumor immunotherapy and to various clinical applications in phase I/II studies on the treatment of different malignancies. This chapter will review the latest developments and give a brief update of the results obtained in studies of advanced melanoma, as well as provide a short overview of published results for other tumors.
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In this study we have analyzed CD30-antigen expression in three melanoma-directed cytotoxic T lymphocyte (CTL) clones with a T helper 0 (Th0)-like cytokine secretion profile (i.e. interleukin (IL)-4, IL-5, and interferon (IFN)-gamma). We show that all CTL clones expressed high levels of CD30 upon contact with the autologous tumour cells. One CTL clone, termed A2 with a monoclonal feature was selected for further analyses and found its CD30 expression dependent on the presence of IL-4. Functionally, a CD30-expressing A2 CTL was capable of producing higher amounts of IFN-gamma (up to 1.5-fold) and IL-4 (up to two-fold) than its CD30- counterpart. Furthermore, CD30-positive A2 CTL displayed an at least three-fold greater proliferative response to the tumour cell stimulation, contrasting with CD30- CTL. However, the antitumour cytotoxic activity of A2 CTL was not modulated by the CD30 expression. These results suggest that CD30 antigen can be inducible on a subset of tumour-directed CD8+ CTL, and that this subset of cells may have profound effector functions, such as cytokine secretion, proliferation, and cytotoxicity.