Melanoma: Reinhold U

Garbe C, Hauschild A, Volkenandt M, Schadendorf D, Stolz W, Reinhold U, Kortmann RD, Kettelhack C, Frerich B, Keilholz U, Dummer R, Sebastian G, Tilgen W, Schuler G, Mackensen A, Kaufmann R. · University Department of Dermatology, Tübingen, Germany. · Melanoma Res. · Pubmed #18337653 No free full text.

Abstract: Systemic medical treatment of melanoma is administered in the adjuvant and palliative setting. Adjuvant therapy may be considered in patients with primary melanoma with more than 1.5 mm tumor thickness and with regional node metastasis. Presently no indication for systemic adjuvant chemotherapy or for adjuvant therapy with nonspecific immune-stimulatory agents outside controlled studies is seen. Interferon-alpha is the first substance in the adjuvant therapy of melanoma, which has shown to present a significant advantage to the patients in some prospective randomized studies. Good arguments for using adjuvant interferon-alpha therapy in high-risk melanoma patients exist. Both high-dose and low-dose interferon-alpha show promise. The major indications for systemic chemotherapy and chemoimmunotherapy are inoperable recurrent tumors, inoperable regional metastases and distant metastases (stage IV). As treatment in such situations is primarily palliative, the effect of any regimen on the quality of life must be carefully weighed. As a first line treatment, single agent therapy is recommended, as polychemotherapy or biochemotherapy did not show significant advantages for prolongation of survival; hence they are more toxic. An urgent need for development of new treatment modalities is necessary and general principles of experimental immunotherapy are outlined.

Garbe C, Hauschild A, Volkenandt M, Schadendorf D, Stolz W, Reinhold U, Kortmann RD, Kettelhack C, Frerich B, Keilholz U, Dummer R, Sebastian G, Tilgen W, Schuler G, Mackensen A, Kaufmann R. · University Department of Dermatology, Tübingen, Germany. · Melanoma Res. · Pubmed #18227710 No free full text.

Abstract: The primary treatment of a melanoma is surgical excision. An excisional biopsy is preferred, and safety margins of 1 cm for tumor thickness up to 2 mm and 2 cm for higher tumor thickness should be applied either at primary excision or in a two-step procedure. When dealing with facial, acral or anogenital melanomas, micrographic control of the surgical margins may be preferable to allow reduced safety margins and conservation of tissue. The sentinel lymph node biopsy should be performed in patients whose primary melanoma is thicker than 1.0 mm and this operation should be performed in centers where both the operative and nuclear medicine teams are experienced. In clinically identified lymph node metastases, radical lymph node dissection is considered standard therapy. If distant metastases involve just one internal organ and operative removal is feasible, then surgery should be seen as therapy of choice. Radiation therapy for the primary treatment of melanoma is indicated only in those cases in which surgery is impossible or not reasonable. In regional lymph nodes, radiation therapy is usually recommended when excision is not complete (R1 resection) or if the nodes are inoperable. In distant metastases, radiation therapy is particularly indicated in bone metastases, brain metastases and soft tissue metastases.

Ugurel S, Reinhold U, Tilgen W. · Department of Dermatology, The Saarland University Hospital, Homburg/Saar, Germany. · Onkologie. · Pubmed #12006763 No free full text.

Abstract: The non-classical HLA class-I molecule HLA-G, primarily expressed on fetal cells of the human placenta, has been shown to play a crucial role in maintaining an immuno-privileged environment at the materno-fetal interface. Fetal trophoblast cells are protected from attack by CD8+ cytotoxic T lymphocytes due to their lacking expression of classical HLA class-I molecules, again rendering them susceptible to natural killer (NK) cell lysis. HLA-G has been shown to interact with killing inhibitory receptors, hereby rescuing the fetal placenta cells from NK cell attack. Likewise, classical HLA class-I molecules are known to be frequently downregulated or lost during the development of malignancies. This abnormality is often associated with a poor clinical course of disease, despite of the frequent detection of tumor-infiltrating NK cells. This controversy seemed to be resolved with the detection of HLA-G expression on cells of malignant melanoma and other solid tumors. Since interferon(IFN)s are known for their ability of upregulation or induction of HLA-G expression, the potential function of HLA-G as a new strategy of cancer cells to escape from immunosurveillance might be of particular importance in malignant melanoma. The frequent use of IFN-alpha in the immunotherapy of this malignancy might possibly worsen the already impaired antitumoral immune response state of melanoma patients. However, several studies recently failed to detect any HLA-G protein expression in cancer cell lines and tissues of different origin, particularly in malignant melanoma, yet rendering the expression and function of HLA-G in malignancies as a possibly overrated matter of controversial debate.

Abken H, Hombach A, Heuser C, Reinhold U. · Klinik I für Innere Medizin, Universität zu Köln, Germany. · Recent Results Cancer Res. · Pubmed #11092052 No free full text.

Abstract: The application of immunotherapy to the treatment of micrometastases of melanoma has attracted growing interest in recent years. This trend reflects, at least in part, the disappointing results of conventional chemotherapy, the identification of melanoma-associated antigens suitable to be used as targets for immunotherapy, and the significant progress in our understanding of molecular processes involved in the development of an immune response. Because of the general belief that T cell immunity plays a major part in the control of tumor growth, we have recently applied a novel strategy to target cytolytic T cells to melanoma cells. The strategy bypasses the requirement of presentation of melanoma-associated-antigen-derived peptides by the major histocompatibility complex to the T cell receptor complex by permanent grafting of T cells with a recombinant, chimeric T cell receptor. The extracellular moiety of the grafted receptor contains the antigen-binding domain, consisting of a single-chain antibody fragment (scFv) derived from a monoclonal antibody specific for the high-molecular-weight melanoma-associated antigen (HMW-MAA). The intracellular receptor moiety contains the cellular activation domain, consisting of the gamma-signaling chain derived from the Fc epsilon RI receptor. Cytotoxic T cells grafted with the chimeric anti-HMW-MAA scFv-gamma signaling receptor specifically lyse HMW-MAA-positive melanoma cells in a human leukocyte antigen class I-independent fashion. The chimeric T cell receptor strategy is designed to eliminate disseminated tumor cells by the power of cytolytic T cells that physiologically penetrate tissues and that are specifically activated by the grafted receptor after binding to antigen-positive melanoma cells.

Ugurel S, Schadendorf D, Pföhler C, Neuber K, Thoelke A, Ulrich J, Hauschild A, Spieth K, Kaatz M, Rittgen W, Delorme S, Tilgen W, Reinhold U, Anonymous00032. · Skin Cancer Unit, German Cancer Research Center Heidelberg and Department of Dermatology, University Hospital of Mannheim, Mannheim, Germany. · Clin Cancer Res. · Pubmed #17000680 links to  free full text

Abstract: PURPOSE: In vitro sensitivity assays are promising tools to predict the individual outcome of different chemotherapy regimens. However, a direct association between in vitro and in vivo chemosensitivity has to be shown by clinical studies. This multicenter phase II trial was aimed to investigate the efficacy of a sensitivity-directed, first-line chemotherapy in metastasized melanoma patients, and to prove an association between in vitro sensitivity and therapy outcome. PATIENTS AND METHODS: The primary study end point was objective response; secondary end points were safety, overall survival, and progression-free survival. Viable tumor cells obtained from metastatic lesions were tested for chemosensitivity to seven single drugs and five drug combinations using an ATP-based luminescence viability assay. RESULTS: Out of 82 recruited patients (intention-to-treat), 57 received assay-directed chemotherapy and 53 were evaluable for all study end points (per protocol). The drug combinations used were gemcitabine+treosulfan, paclitaxel+cisplatin, paclitaxel+doxorubicin, and gemcitabine+cisplatin. The per protocol population could be divided into 22 (42%) chemosensitive and 31 (58%) chemoresistant patients by an arbitrary chemosensitivity index. Objective response was 36.4% in chemosensitive patients compared with 16.1% in chemoresistant patients (P=0.114); progression arrest (complete response+partial response+stable disease) was 59.1% versus 22.6% (P=0.01). Chemosensitive patients showed an increased overall survival of 14.6 months compared with 7.4 months in chemoresistant patients (P=0.041). CONCLUSION: In vitro chemosensitivity testing may be worthy of further exploration to see if it could be a useful tool to predict the outcome of melanoma patients treated with a sensitivity-directed chemotherapy. Therefore, these preliminary results will be evaluated by a planned phase III trial using a randomized, standard-regimen controlled setting.

Kurbacher CM, Kurbacher JA, Cramer EM, Rhiem K, Mallman PK, Reichelt R, Reinhold U, Stier U, Cree IA. · Institute of Gynecologic Oncology, Bonn Comprehensive Cancer Network, Germany. · Oncology (Williston Park). · Pubmed #15934497 No free full text.

Abstract: Granulocyte-macrophage colony-stimulating factor (GM-CSF, sargramostim [Leukine]) is a powerful cytokine that is able to stimulate the generation of dendritic cells. Adjuvant treatment with continuous low-dose GM-CSF has been shown to prolong survival of stage III/IV melanoma patients. Data on continuous low-dose GM-CSF therapy in tumors other than prostate cancer are still lacking. This pilot trial was initiated in order to evaluate the efficacy and tolerability of continuous low-dose GM-CSF as salvage in various chemotherapy-refractory carcinomas. A total of 19 patients who had failed a median of 4 prior chemotherapies were included. Their malignancies included metastatic breast cancer, recurrent ovarian carcinoma, metastatic endometrial carcinoma, and recurrent squamous cell cancer of the cervix uteri. Continuous low-dose GM-CSF was delivered subcutaneously at a daily starting dose of 125 microg. GM-CSF was increased at 25-microg increments until a maximum of 250 microg was reached or when mild leukocytosis (10-20 g/L) was achieved, providing that the relative eosinophil count did not exceed 15%. Therapy was continued until progression or refusal by the patient. Toxicity was generally mild. Only one patient was withdrawn due to grade 3 fatigue. In three additional patients, temporary dose reduction was necessary because of grade 1 injection site reactions, which recovered spontaneously. Mild to moderate leukocytosis was obvious in 10 patients. Systemic hypersensitivity-like reactions did not occur and no patient required hospitalization for other life-threatening side effects. The objective response rate was 37%: 1 complete and 6 partial responses, 4 disease stabilizations, 8 progression of disease. Median response duration was 6 months. Notably, 6 of 7 responders but only 1 of 8 patients with disease progression developed leukocytosis during therapy. Therefore, we conclude that continuous low-dose GM-CSF has substantial activity in heavily pretreated patients with either metastatic breast cancer or female genital tract cancer. Achievement of mild leukocytosis seems to be a predictor of response.

Pföhler C, Cree IA, Ugurel S, Kuwert C, Haass N, Neuber K, Hengge U, Corrie PG, Zutt M, Tilgen W, Reinhold U. · Saarland University Hospital, Department of Dermatology, Homburg/Saar, Germany. · Anticancer Drugs. · Pubmed #12782938 No free full text.

Abstract: No effective treatment currently exists for metastatic uveal melanoma. However, recent results obtained by an ATP-based tumor chemosensitivity assay have shown consistent activity of treosulfan+gemcitabine in up to 80% of tumor specimens tested. In this study we describe the first clinical results observed with this drug combination at different European centers in patients with metastatic uveal melanoma. Clinical case series of patients with metastatic uveal melanoma were treated with treosulfan+gemcitabine at seven different centers. Fourteen patients, 13 previously untreated and one pretreated with chemoimmunotherapy, were included in the study. Patients received treosulfan+gemcitabine in four different dose regimens. The response rates, progression-free and overall survival, and toxicity were evaluated. The analysis of 14 patients revealed one complete response, three partial responses and a stable disease in eight cases. The objective response rate was 28.6%, the median overall survival was 61 weeks [95% confidence interval (CI) 54-133 weeks], the progression-free survival was 28.5 weeks (95% CI 13-62 weeks) and the 1-year survival rate was 80%. The drugs were well tolerated. The most common side-effects were leuko- and thrombocytopenia. These preliminary results suggest potential therapeutic benefit of treosulfan+gemcitabine treatment in metastatic uveal melanoma and warrant further controlled studies.

Neuber K, Reinhold U, Deutschmann A, Pföhler C, Mohr P, Pichlmeier U, Baumgart J, Hauschild A. · Department of Dermatology, University Hospital Hamburg, Martinistr. 52, D-20246 Hamburg, Germany. · Melanoma Res. · Pubmed #12569289 No free full text.

Abstract: The purpose of this multicentre phase II trial was to evaluate time to progression, survival time, rate of objective tumour response and toxicity of second-line intravenous treosulfan chemotherapy in stage IV melanoma patients. Thirty-one patients with measurable stage IV malignant melanoma and prior chemotherapy with a dacarbazine-containing regimen were included. Of this group, 26 patients were evaluable. All patients received treosulfan (8 g/m intravenously on day 1; cycle repeated every 28 days up to six courses). Patients were evaluated for tumour response, survival time and toxicity. No objective responses (complete or partial) were observed. Five patients (19%) showed no change and 21 had progressive disease after treosulfan treatment. Four patients experienced a minor or mixed response. The median time to progression was 1.8 months (95% confidence interval [CI] 1.6-2.1 months) and the median overall survival was 6.5 months (95% CI 3.1-10 months). The 1 year survival rate was 33.9% (95% CI 15.4-52.3%). Leukocytopenia and thrombocytopenia (Common Toxicity Criteria grades 3 and 4) occurred in 15% and 18% of cases, respectively. The non-haematological toxicity of this outpatient regimen was mild. In conclusion, intravenous treosulfan treatment does not induce objective response rates when used as a second-line treatment of metastatic malignant melanoma.

Reinhold U, Schmitz B, Kurbacher C, Nagel W, Schmidt M, Malaisse WJ. · Medical Center Bonn Friedensplatz, D-53111 Bonn, Germany. · Oncol Rep. · Pubmed #19020739 No free full text.

Abstract: The serum 25-hydroxyvitamin D [25(OH)D] concentration was measured in 20 patients with prostatic carcinoma, compared to 75 subjects with prostatic hyperplasia, in 24 male and 17 female patients with melanoma, in 26 female patients with breast cancer, 7 patients with ovarian carcinoma and 3 patients with cervix carcinoma among subjects followed in a German polyclinical centre. In >50% of these 174 subjects, 25(OH)D concentration was < 20 microg/l. In most subject groups, a seasonal decrease of 25(OH)D concentration was observed during the winter period. An age-related decrease in such a concentration was also observed in subjects with prostatic hyperplasia examined in the late summer/early autumn period and in female cancer subjects, at the exclusion of patients with breast cancer. In the latter patients, however, a positive correlation prevailed between age and 25(OH)D concentration. Hence, it is proposed that an abnormally low serum 25(OH)D concentration represents a preferential risk factor, in middle-aged women, for breast cancer, as compared to other neoplasic manifestations in female subjects.

Garbe C, Schadendorf D, Stolz W, Volkenandt M, Reinhold U, Kortmann RD, Kettelhack C, Frerich B, Keilholz U, Dummer R, Sebastian G, Tilgen W, Schuler G, Mackensen A, Kaufmann R, Hauschild A. · Sektion Dermatologische Onkologie, Universitäts-Hautklinik, Liebermeister Strasse 25, D-72076 Tübingen, Germany. · J Dtsch Dermatol Ges. · Pubmed #18801142 No free full text.

This publication has no abstract.

Garbe C, Hauschild A, Volkenandt M, Schadendorf D, Stolz W, Reinhold U, Kortmann RD, Kettelhack C, Frerich B, Keilholz U, Dummer R, Sebastian G, Tilgen W, Schuler G, Mackensen A, Kaufmann R. · University Department of Dermatology, Tübingen, Germany. · Melanoma Res. · Pubmed #17992123 No free full text.

Abstract: Melanoma is a malignant tumor that arises from melanocytic cells and primarily involves the skin. The most important exogenous etiological factor is exposure to ultraviolet irradiation. Diagnosis of melanoma is based primarily on its clinical features, and the A-B-C-D rule is useful in identifying pigmented lesions, which are suspicious for melanoma (Asymmetry, Border irregular, Color inhomogeneous and Diameter more than 5 mm). Dermoscopy is very helpful in clarifying the differential diagnosis of pigmented lesions. About 90% of melanomas are diagnosed as primary tumors without any evidence for metastasis. The tumor-specific 10-year survival for all such tumors is about 75-85%. The most important prognostic factors for primary melanoma without metastases are vertical tumor thickness (Breslow depth) as measured on the histological specimen, presence of histopathologically recognized ulceration, invasion level (Clark level) and identification of micrometastases in the regional lymph nodes via sentinel lymph node biopsy. The current tumor node metastasis classification for the staging of primary melanoma is based on these factors. Melanomas can metastasize either by the lymphatic or by the hematogenous route. About two-thirds of metastases are originally confined to the drainage area of regional lymph nodes. A regional metastasis can appear as satellite metastases up to 2 cm from the primary tumor, as intransit metastases in the skin between the site of the primary tumor and the first lymph node and as regional lymph node metastases. In the stage of regional metastasis, the differentiation between micrometastasis and macrometastasis and the number of lymph nodes involved are crucial. As soon as distant metastasis develops, prognosis depends on the site of the metastasis and on the lactate dehydrogenase levels in the blood. The frequency and extent of follow-up examinations is based on the initial tumor parameters. In thin primary melanomas up to 1-mm tumor thickness, clinical examinations at 6-month intervals are sufficient and in thicker primary melanomas, at 3-month intervals. Lymph node sonography as well as determination of the tumor marker protein S100beta are recommended. Additionally, in the stage of regional metastasis, whole body imaging should be performed every 6 months; in the stage of distant metastasis, surveillance has to be scheduled individually.

Buzug TM, Schumann S, Pfaffmann L, Reinhold U, Ruhlmann J. · Dept. of Math. & Technol., RheinAhrCampus Remagen, Germany. · Conf Proc IEEE Eng Med Biol Soc. · Pubmed #17945738 No free full text.

Abstract: Annually 133.000 people world-wide get sick on malign melanoma, tendency increasing. The purpose of this study is the early diagnosis of malignant skin cancer. At the moment the dermatologists are screening for anomalies at the relevant lesion by examining the skin area with a microscope. To determine changes, another scan has to be taken in a follow-up session after a time period of about 15-20 weeks. Today's visual diagnostic decision is based on the pragmatic ABCD approach (Asymmetry, Border, Colour, and Diameter). However, there is no adequate and sound non-invasive way to find out, if a skin spot is either malign or benign. If the visual approach corroborates a suspicion of skin cancer, histology is needed to make explicit diagnosis. To avoid unnecessary surgeries (on false positive alarm) and to initiate necessary surgeries in early stages a new diagnostic screening approach is presented here. Based on the fact that malign melanoma have higher metabolism as well as increased blood flow, it has been conjectured that malign melanoma have slightly higher temperature compared to the healthy skin that can be measured by high resolution functional infrared imaging.

Ladewig G, Reinhold U, Thirkill CE, Kerber A, Tilgen W, Pföhler C. · Department of Dermatology, The Saarland University Hospital , 66421 Homburg/Saar, Germany. · Br J Dermatol. · Pubmed #15888149 No free full text.

Abstract: BACKGROUND: Patients with melanoma-associated retinopathy (MAR) experience different visual symptoms caused by the production of antitumoral antibodies that cross-react with retinal epitopes. Immunofluorescence assays of serum from patients with MAR on sectioned monkey or human retina characteristically reveal antibody activity located within the inner nuclear layer, with a focus of activity upon the membranes of bipolar cells. OBJECTIVES: We inquired into the association with disease of this serological abnormality by evaluating sera from patients with melanoma with no MAR-like signs or symptoms. METHODS: Groups of patients were selected with different stages of melanoma (American Joint Committee on Cancer stages I-IV). Seventy-seven serum samples from 51 patients with melanoma were examined by indirect immunohistochemical assay on sections of human retina. RESULTS: Of the 77 serum samples, 53 were found to contain antibodies reactive with various components of retina. Eight were from 17 sera from patients in stage I or II, 14 were from 23 sera from patients in stage III, and 31 were from 37 sera from patients in stage IV. Statistical analysis revealed a correlation between antibody activity and the stage of disease, with a higher percentage of antibody activity in advanced stages (P = 0.002). CONCLUSIONS: The presence of antiretinal antibodies in patients with melanoma without ocular symptoms appears to be more common than previously suspected. Antibody activity similar to that ascribed to the MAR syndrome appears in some patients with melanoma who have no MAR-like retinopathy. Follow-up studies will determine if patients with antiretinal antibodies go on to develop MAR and if staining intensity and staining patterns change over the course of the disease.

Ugurel S, Uhlig D, Pföhler C, Tilgen W, Schadendorf D, Reinhold U. · Skin Cancer Unit, German Cancer Research Center, Heidelberg, Germany. · Cancer Immunol Immunother. · Pubmed #14727087 No free full text.

Abstract: Antigen-presenting cells are crucial for the induction of an antigen-specific antitumoral immune response. Deteriorations in the expression pattern of cell surface molecules important for the presentation of antigens might therefore be indicative of an impaired immune response status in cancer patients. In the present study we investigated the expression of MHC class I and class II molecules, of the costimulatory molecules CD80/B7-1 and CD86/B7-2, of the adhesion molecule CD11c, and of the marker of activation CD71 on CD14+ peripheral blood monocytes (PBMs) from 144 melanoma patients in different stages of disease and 43 healthy controls, by flow cytometric analysis. We found a decreased expression of HLA-DR (p<0.0005), HLA-DQ (p=0.006), HLA-DP (p<0.0005), and CD86/B7-2 (p=0.001) on PBMs from melanoma patients compared with healthy controls, whereas no significant difference could be detected in the expression of HLA class I antigens and CD80/B7-1. This down-regulated expression was associated with disease progression. In contrast, CD71 expression was stage-dependently increased on PBMs from melanoma patients compared with healthy controls (p=0.024). No correlation was found between the PBM surface expression pattern and age, gender, tumor load, and current mode of therapy of the patients. The observed down-regulation of HLA class II and CD86/B7-2 on melanoma patients' PBMs might reflect an ineffective antigen-presenting function contributing to an impaired antigen-specific immune response in these patients.

Pföhler C, Haus A, Palmowski A, Ugurel S, Ruprecht KW, Thirkill CE, Tilgen W, Reinhold U. · Department of Dermatology, The Saarland University Hospital, 66421 Homburg/Saar, Germany. · Br J Dermatol. · Pubmed #12890197 No free full text.

Abstract: BACKGROUND: Melanoma-associated retinopathy (MAR) is a paraneoplastic syndrome with symptoms of night blindness, light sensations, visual loss, defect in visual fields, and reduced b-waves in the electroretinogram. Patients with MAR often suffer from a sudden onset of ocular symptoms that are believed to result from antibody production against melanoma-associated antigens that cross-react with corresponding epitopes on retinal depolarizing bipolar cells. OBJECTIVES: To correlate the frequency of subclinical symptoms suggestive of MAR in melanoma patients to different stages of disease, patient age, type and thickness of the primary tumour, form of therapy, S-100 level and tumour burden. METHODS: We analysed 28 patients with melanoma in stages I-IV (according to the American Joint Committee on Cancer tumour classification) for the presence of subclinical MAR symptoms using scotopic electroretinography, static and kinetic perimetry and nyctometry. RESULTS: Seven patients had clinical signs and symptoms consistent with MAR, 18 had some indications, while the remaining three had none. We found no correlation between clinical symptoms and stage of disease, tumour burden or S-100 level, but findings suggestive of MAR were observed more frequently in advanced stages of disease. CONCLUSIONS: Subclinical retinal involvement characteristic of MAR appears to be more common than previously suspected in patients with cutaneous malignant melanoma. Our findings in this small cohort seem to indicate that the percentage of patients with symptoms suggestive of MAR is higher in advanced stages of disease. Further clinical studies are required to evaluate if the presence of subclinical symptoms suggestive of MAR is correlated with a worse prognosis and a shortened progression-free and overall survival.

Ugurel S, Tilgen W, Reinhold U. · Department of Dermatology, The Saarland University Hospital, 66421 Homburg/Saar, Germany. · Recent Results Cancer Res. · Pubmed #12528801 No free full text.

Abstract: The prognosis of patients with metastatic melanoma remains poor. In patients with distant metastases only low response rates between 10% and 15% have been achieved by the most effective cytostatics in single-agent therapy leading to a mean 5-year survival rate of less than 5%. More aggressive treatment regimens using multidrug chemotherapy yielded response rates of up to 40% but failed to show a significant benefit in overall survival compared to single-agent therapy. However, complete remissions of metastatic lesions after multidrug cytostatic regimens have been reported in some cases of melanoma patients. To evaluate an in vitro test system providing information on the drug sensitivity profile of melanoma cells, we examined tumor tissue specimens from 31 metastatic melanoma patients with an ATP-based chemosensitivity assay (ATP-TCA) testing eight anticancer drugs alone or in different combinations. Chemosensitivity was assessed using a luciferin-luciferase- based luminescence assay providing individual chemosensitivity indices for each test drug. We found a heterogeneous chemosensitivity in the melanoma tissue samples tested. The highest sensitivity was detected for the combination of treosulfan and gemcitabine, with 76% of the tissue samples revealing high sensitivity and 10% resistance, followed by the combination of paclitaxel and doxorubicine (66%/0%), gemcitabine and cisplatin (55%/21%),and paclitaxel and cisplatin (46%/8%). Our data indicate that the ATP-TCA can be used to select patients who might benefit from an individually adapted cytostatic therapy. On the basis of these results a multicenter trial has recently been initiated to evaluate the feasibility and predictive value of an ATP-TCA directed chemotherapy in metastatic melanoma patients.

Palmowski AM, Haus AH, Pföhler C, Reinhold U, Allgayer R, Tilgen W, Ruprecht KW, Thirkill CE. · Department of Ophthalmology, Saarland University Hospital, D-66421 Homburg/Saar, Germany. · Arch Ophthalmol. · Pubmed #12470158 No free full text.

This publication has no abstract.

Rebmann V, Ugurel S, Tilgen W, Reinhold U, Grosse-Wilde H. · Institute of Immunology, University Hospital of Essen, Essen, Germany. · Int J Cancer. · Pubmed #12124808 No free full text.

Abstract: Despite numerous therapeutic options, the prognosis of malignant melanoma, once metastasized, is still poor. Thus, the search for reliable methods to identify patients with high risk of disease progression as early as possible is of major importance. In our study, we analyzed the predictive value of soluble HLA-DR (sHLA-DR) in comparison to S100-beta in serum from 183 melanoma patients of different stages of disease and with or without current therapy using immunosorbent assays. sHLA-DR serum levels of 121 healthy individuals served as controls. We found significantly (p < 0.0005) reduced sHLA-DR serum levels in melanoma patients (0.70 +/- 0.08 SEM microg/ml) compared to controls (1.49 +/- 0.10 SEM microg/ml). Reduced sHLA-DR and increased S100-beta levels were associated with advanced disease stages and tumor load. S100-beta was increased under cytostatic therapy (p < 0.0005), whereas sHLA-DR was not influenced by therapy modalities. Univariate analysis showed an association of sHLA-DR < 0.3 microg/ml and S100-beta > 0.12 microg/l with poor overall (p = 0.021 and p = 0.0009) and progression-free survival (p < 0.0005 and p = 0.0025). Multivariate analysis revealed disease stage (p = 0.0093) and tumor burden (p < 0.0005) as independent predictive factors for overall survival, and sHLA-DR (p = 0.0007) and tumor burden (p = 0.0015) for progression-free survival. In contrast to S100-beta, sHLA-DR serum concentrations < 0.3 microg/ml were strongly associated (p = 0.0001) with poor progression-free survival in a subgroup of 60 nonmetastasized patients. In conclusion, our results suggest sHLA-DR as a potent prognostic serum marker in melanoma patients superior to S100-beta in helping to identify early-stage patients at high risk of disease progression.

Reinhardt MJ, Kensy J, Frohmann JP, Willkomm P, Reinhold U, Grünwald F, Biersack HJ, Bender H. · Department of Nuclear Medicine, University Hospital Bonn. · Nuklearmedizin. · Pubmed #12109034 No free full text.

Abstract: AIM: The S-100B protein is commonly used in the immunohistochemical diagnosis of malignant melanoma and its metastases and has recently been introduced as a tumor marker in peripheral blood, whereas 18F-FDG PET is currently the most sensitive in-vivo imaging method for melanoma staging. Thus, the efficiency of serum S-100B and 18F-FDG PET in the detection of metastatic disease in melanoma patients are compared. METHODS: Serum S-100B was measured with a commercially available immunoradiometric assay. As part of primary tumour staging whole-body positron emission tomography (PET) with 18F labeled fluorodeoxy-D-glucose (18F-FDG) was performed in 67 patients suffering from cutaneous melanoma with a tumour thickness > 0.75 mm and a Clark-level III-V. Final diagnosis based on histology, morphologic imaging results and/or clinical follow-up after at least six months. RESULTS: No evidence of disease was seen in 43 of 67 patients (64.2%), 11 patients (16.4%) presented with lymph node metastases, 13 patients (19.4%) had one or more distant metastases. Alltogether, 18 of 67 patients showed S-100B values > 0.2 microgram/l, including two patients without metastatic disease, 3 of 11 patients with lymph node metastases, and the 13 patients with distant metastases. One patient showed false-positive FDG-uptake in the mediastinum, but presented with S-100B values off curve. CONCLUSION: Our data indicate that serum S-100B determination might be helpful in identifying melanoma patients with distant metastases. In comparison to 18F-FDG PET, the value of serum S-100B for lymph-node staging is limited.

Rappl G, Hasselmann DO, Rössler M, Ugurel S, Tilgen W, Reinhold U. · Department of Dermatology, The Saarland University Hospital, Homburg/Saar, Germany. · Ann N Y Acad Sci. · Pubmed #11708477 No free full text.

Abstract: It has been suggested that extracellular mRNA might be released from tumor cells and might be protected from serum RNase by proteins or proteolipid complexes. Our group has recently assessed the potential value of extracellular RNA detection by RT-PCR in the peripheral blood of patients with metastatic melanoma.

Max N, Willhauck M, Wolf K, Thilo F, Reinhold U, Pawlita M, Thiel E, Keilholz U. · Department of Medicine III, University Hospital Benjamin Franklin, Free University Berlin, Hindenburgdamm 30, 12200 Berlin, Germany. · Melanoma Res. · Pubmed #11479425 No free full text.

Abstract: For the molecular detection of rare tumour cells in clinical samples, real-time reverse transcription-polymerase chain reaction (RT-PCR) offers two important advantages over conventional RT-PCR assays: the results are quantitative and, perhaps more importantly, it facilitates exact sensitivity controls on a per sample basis as well as exact comparison of different assay protocols. We report here on quantitative results obtained with different protocols for RNA isolation and cDNA synthesis for amplification of beta2-microglobulin transcripts using the light cycler system. Furthermore, housekeeping gene-specific PCRs were compared with PCRs specific for an artificial transcript (internal standard) detected simultaneously at a level comparable to the wild-type sequence. Artificial tyrosinase transcripts derived from a vector construct stably transfected into a human lymphoma cell line were used as a model to test the usefulness of artificial internal standards as an alternative to housekeeping genes. The highest RNA yields were obtained using a combination of phenol-chloroform extraction and the High Pure RNA Isolation Kit. Analysing beta2-microglobulin transcript-specific RT-PCRs, the highest sensitivity was obtained for cDNAs generated with Omniscript reverse transcriptase and oligo-p(dT)15 primer. Regarding patient blood samples, RT-PCRs specific for beta2-microglobulin, porphobilinogen deaminase and artificial tyrosinase transcripts provided quantitative data for all, for 18 out of 21, and for 10 out of 21 samples, respectively. Quantification of beta2-microglobulin transcripts by the light cycler system defined the protocol revealing the highest cDNA quality. Comparisons of quantitative data from RT-PCRs specific for beta2-microglobulin, porphobilinogen deaminase and artificial tyrosinase transcripts enabled us to determine a close range for crossing points within which sufficient cDNA quality can be guaranteed, even for the detection of rare transcripts. PCRs specific for the artificial internal standard are ideally suited for cDNA quality assessment on a per sample basis.

Hasselmann DO, Rappl G, Tilgen W, Reinhold U. · Department of Dermatology, The Saarland University Hospital, 66421 Homburg/Saar, Germany. · Clin Chem. · Pubmed #11468248 links to  free full text

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Ugurel S, Rebmann V, Ferrone S, Tilgen W, Grosse-Wilde H, Reinhold U. · Department of Dermatology, The Saarland University Hospital, Homburg/Saar, Germany. · Cancer. · Pubmed #11466692 links to  free full text

Abstract: BACKGROUND: The nonclassic human major histocompatibility complex class I antigens human leukocyte antigen (HLA)--G are proposed to protect tumor cells from natural killer cell lysis. In the current study, the authors measured soluble HLA-G molecules (sHLA-G) in serum from patients with malignant melanoma. METHODS: Soluble HLA-G was determined in serum samples of 190 melanoma patients with various stages of disease, with or without current therapy including interferon (IFN)-alpha and different cytostatics in comparison to 126 healthy controls by using a two-step enzyme-linked immunoadsorbent assay. RESULTS: Serum sHLA-G was significantly (P < 0.0005) elevated in melanoma patients (mean +/- standard error of the mean [SEM] = 41.95 +/- 2.15 ng/mL) compared with healthy controls (mean +/- SEM = 22.92 +/- 1.51 ng/mL). Univariate analysis revealed a correlation of sHLA-G serum level with advanced stages of disease (P < 0.001) and tumor load (P < 0.05). Patients undergoing immunotherapy with IFN-alpha (n = 31) showed an increased serum sHLA-G (mean +/- SEM = 62.05 +/- 7.58 ng/mL; P < 0.0005), whereas other treatment regimens (n = 24) did not influence sHLA-G serum concentrations. Multivariate analysis revealed treatment with IFN-alpha as the only impact factor for elevated serum sHLA-G, lacking any correlation with stage of disease or tumor burden. Furthermore, IFN-alpha was found to upregulate HLA-G cell surface expression on circulating monocytes. sHLA-G serum level was not associated with recurrence free or overall survival. CONCLUSIONS: This study shows increased sHLA-G serum concentrations in melanoma patients and additional enhancement upon treatment with IFN-alpha. The level of serum sHLA-G, however, had no negative impact on patients' prognosis.

Ugurel S, Rappl G, Tilgen W, Reinhold U. · Department of Dermatology, The Saarland University Hospital, 66421 Homburg/Saar, Germany. · Clin Cancer Res. · Pubmed #11350895 links to  free full text

Abstract: Functional impairment of the Fas/CD95 receptor-ligand system is associated with the development and progression of malignancies. One possible cause might be the inhibition of the formation of a functional Fas/CD95-FasL complex by soluble Fas/CD95 molecules (sFas/CD95). In the present study we determined sFas/CD95 serum concentration in 125 melanoma patients of different clinical stages of disease compared with 30 healthy controls using an ELISA. sFas/CD95 serum level was significantly elevated (P < 0.0005) in melanoma patients (mean +/- SE = 8.60 +/- 0.26 ng/ml) compared with healthy controls (mean +/- SE = 6.27 +/- 0.25 ng/ml). Univariate analysis revealed a correlation of sFas/CD95 serum concentration with advanced stages of disease (P = 0.009). Only a slight increase in sFas/CD95 serum level (P = 0.057) could be observed in regard to the tumor burden. Patients undergoing current treatment with cytostatics (n = 18) revealed a strong increase in sFas/CD95 serum level (P < 0.0005), whereas treatment with IFN-alpha alone or combined with cytostatics (n = 19) showed no change in serum sFas/CD95 concentration. According to univariate analysis, elevated sFas/CD95 serum levels were associated with a poor overall (P < 0.005) and a progression-free (P < 0.0005) survival. Multivariate analysis revealed sFas/CD95 serum concentration as an independent predictive factor for progression-free (P = 0.011), but not overall (P = 0.078), survival. Our results show a prognostic relevance of serum sFas/CD95 in melanoma patients, indicating that the evaluation of sFas/CD95 serum level may be important for the selection of therapeutic strategies.

Reinhold U, Berkin C, Bosserhoff AK, Deutschmann A, Garbe C, Gläser R, Hein R, Krähn G, Peter RU, Rappl G, Schittek B, Seiter S, Ugurel S, Volkenandt M, Tilgen W. · Department of Dermatology, The Saarland University Hospital, Homburg/Saar, Germany. · J Clin Oncol. · Pubmed #11251002 No free full text.

Abstract: PURPOSE: Reverse transcription-polymerase chain reaction (RT-PCR)-based detection of tyrosinase mRNA is the most frequently used laboratory method for the detection of circulating tumor cells in melanoma patients. However, previously published results showed considerable variability in the PCR positivity rates. MATERIALS AND METHODS: We designed a collaborative study to assess the sensitivity, specificity, and clinical relevance of a new standardized RT-PCR-based enzyme-linked immunosorbent assay (ELISA) for the detection of circulating melanoma cells. Blood samples of healthy donors mixed with cells of a melanoma cell line were prepared in a blinded fashion, and aliquots were sent to seven participating laboratories experienced in RT-PCR. RESULTS: The results demonstrate a high sensitivity (1 melanoma cell/mL blood) and specificity (no false-negatives and 7.4% [2 of 28] false-positives) of the assay and a satisfactory rate of interlaboratory reproducibility. The analysis of aliquots of blinded samples derived from 60 melanoma patients identified tyrosinase mRNA in 17 of 60 (28.3%): three (20%) of 15 stage I patients, two (13.3%) of 15 stage II patients, five (35.7%) of 14 stage III patients, and seven (43.8%) of 16 stage IV patients. The interlaboratory reproducibility of positive samples, however, was extremely low and indicates the presence of low amounts of target mRNA. CONCLUSION: Reverse transcriptase-PCR ELISA has a high sensitivity and specificity for the detection of tyrosinase mRNA in peripheral blood cells. The low interlaboratory reproducibility for the detection of tumor cells in blood samples of melanoma patients, however, raises the question of relevance of this assay for clinical use.