Constraints on the evolution of phenotypic plasticity: limits and costs of phenotype and plasticity.

Phenotypic plasticity is ubiquitous and generally regarded as a key mechanism for enabling organisms to survive in the face of environmental change. Because no organism is infinitely or ideally plastic, theory suggests that there must be limits (for example, the lack of ability to produce an optimal trait) to the evolution of phenotypic plasticity, or that plasticity may have inherent significant costs. Yet numerous experimental studies have not detected widespread costs. Explicitly differentiating plasticity costs from phenotype costs, we re-evaluate fundamental questions of the limits to the evolution of plasticity and of generalists vs specialists. We advocate for the view that relaxed selection and variable selection intensities are likely more important constraints to the evolution of plasticity than the costs of plasticity. Some forms of plasticity, such as learning, may be inherently costly. In addition, we examine opportunities to offset costs of phenotypes through ontogeny, amelioration of phenotypic costs across environments, and the condition-dependent hypothesis. We propose avenues of further inquiry in the limits of plasticity using new and classic methods of ecological parameterization, phylogenetics and omics in the context of answering questions on the constraints of plasticity. Given plasticity's key role in coping with environmental change, approaches spanning the spectrum from applied to basic will greatly enrich our understanding of the evolution of plasticity and resolve our understanding of limits.

Parallel invasions produce heterogenous patterns of life history adaptation: rapid divergence in an invasive insect.

Biotic invasions provide a natural experiment in evolution: when invasive species colonize new ranges, they may evolve new clines in traits in response to environmental gradients. Yet it is not clear how rapidly such patterns can evolve and whether they are consistent between regions. We compare four populations of the invasive cabbage white butterfly (Pieris rapae) from North America and Japan, independently colonized by P. rapae 150 years ago and 300 years ago, respectively. On each continent, we employed a northern and southern population to compare the effects of latitude on body mass, development rate and immune function. For each population, we used a split-sibling family design in which siblings were reared at either warm (26.7 °C) or cool (20 °C) temperatures to determine reaction norms for each trait. Latitudinal patterns in development time were similar between the two continents. In contrast, there were strong geographical differences in reaction norms for body size, but no consistent effects of latitude; there were no detectable effects of latitude or continent on immune function. These results imply that some life history traits respond consistently to selection along climatic gradients, whereas other traits may respond to local environmental factors, or not at all.

Environmental determinants of population divergence in life-history traits for an invasive species: climate, seasonality and natural enemies.

Invasive species cope with novel environments through both phenotypic plasticity and evolutionary change. However, the environmental factors that cause evolutionary divergence in invasive species are poorly understood. We developed predictions for how different life-history traits, and plasticity in those traits, may respond to environmental gradients in seasonal temperatures, season length and natural enemies. We then tested these predictions in four geographic populations of the invasive cabbage white butterfly (Pieris rapae) from North America. We examined the influence of two rearing temperatures (20 and 26.7 °C) on pupal mass, pupal development time, immune function and fecundity. As predicted, development time was shorter and immune function was greater in populations adapted to longer season length. Also, phenotypic plasticity in development time was greater in regions with shorter growing seasons. Populations differed significantly in mean and plasticity of body mass and fecundity, but these differences were not associated with seasonal temperatures or season length. Our study shows that some life-history traits, such as development time and immune function, can evolve rapidly in response to latitudinal variation in season length and natural enemies, whereas others traits did not. Our results also indicate that phenotypic plasticity in development time can also diverge rapidly in response to environmental conditions for some traits.

High-dose pulse corticosteroid therapy in the treatment of severe alopecia areata.

OBJECTIVE: The present monocenter prospective study was designed to evaluate the efficacy of intravenous high-dose methylprednisolone pulse therapy in patients with severe alopecia areata (AA). METHODS: 30 patients (aged 14-56 years) were treated with methylprednisolone (8 mg/kg body weight) intravenously on 3 consecutive days at 4-week intervals for at least 3 courses. RESULTS: 67% of patients with AA plurifocalis showed >50% regrowth of hair. None of the patients with AA totalis or universalis and only 1 patient with ophiasic AA responded to therapy. In patients with AA plurifocalis, higher response rates could be observed in those suffering from long-term disease compared to patients treated during their first episodes of AA (73 vs. 57%). CONCLUSION: High-dose methylprednisolone pulse therapy is an effective and well-tolerated treatment for patients with severe AA plurifocalis but might be less beneficial for patients with ophasic AA, AA totalis or universalis.

Dacarbazine and interferon alpha with or without interleukin 2 in metastatic melanoma: a randomized phase III multicentre trial of the Dermatologic Cooperative Oncology Group (DeCOG).

In several phase II-trials encouraging tumour responses rates in advanced metastatic melanoma (stage IV; AJCC-classification) have been reported for the application of biochemotherapy containing interleukin 2. This study was designed to compare the efficacy of therapy with dacarbazine (DTIC) and interferon alpha (IFN-alpha) only to that of therapy with DTIC and IFN-alpha with the addition of interleukin 2 (IL-2) in terms of the overall survival time and rate of objective remissions and to provide an elaborated toxicity profile for both types of therapy. 290 patients were randomized to receive either DTIC (850 mg/m(2)every 28 days) plus IFN-alpha2a/b (3 MIU/m(2), twice on day 1, once daily from days 2 to 5; 5 MIU/m(2)3 times a week from week 2 to 4) with or without IL-2 (4.5 MIU/m(2)for 3 hours i.v. on day 3; 9.0 MIU/m(2) i.v. day 3/4; 4.5 MIU/m(2) s.c. days 4 to 7). The treatment plan required at least 2 treatment cycles (8 weeks of therapy) for every patient. Of 290 randomized patients 281 were eligible for an intention-to-treat analysis. There was no difference in terms of survival time from treatment onset between the two arms (median 11.0 months each). In 273 patients treated according to protocol tumour response was assessable. The response rates did not differ between both arms (P = 0.87) with 18.0% objective responses (9.7% PR; 8.3% CR) for DTIC plus IFN-alpha as compared to 16.1% (8.8% PR; 7.3% CR) for DTIC, IFN-alpha and IL-2. Treatment cessation due to adverse reactions was significantly more common in patients receiving IL-2 (13.9%) than in patients receiving DTIC/IFN-alpha only (5.6%). In conclusion, there was neither a difference in survival time nor in tumour response rates when IL-2, applied according to the combined intravenous and subcutaneous schedule used for this study, was added to DTIC and IFN-alpha. However, toxicity was increased in melanoma patients treated with IL-2. Further phase III trials with continuous infusion and higher dosages must be performed before any final conclusions can be drawn on the potential usefulness of IL-2 in biochemotherapy of advanced melanoma.

Efficacy of local versus systemic application of antibody-cytokine fusion proteins in tumor therapy.

Application of immunocytokines [fusion proteins (FuPs)] where the cytokine has been coupled to an antibody may not produce the severe side effects frequently observed during systemic application of cytokines in cancer therapy. However, it has not been explored whether FuPs are sufficient for intratumoral activation of leukocytes or whether intratumoral versus systemic application may be of greater efficacy. Interleukin 2 (IL2) or tumor necrosis factor (TNF) coupled to an anti-epidermal growth factor receptor monoclonal antibody (IL2-FuP or TNF-FuP) were tested in SCID mice bearing a human epidermal growth factor receptor-positive melanoma transplant and being reconstituted with human HLA-matched peripheral blood leukocytes. Whole-body autoradiography revealed larger accumulation and prolonged retention of i.v. or intratumorally applied IL2-FuP or TNF-FuP compared with the antibody. Even with low doses of FuP, tumor growth was significantly retarded, with the survival time being further prolonged by the intratumoral application. Furthermore, outgrowth of the tumor was prevented in approximately 50% of mice as long as they received weekly injections of peripheral blood leukocytes concomitantly with the FuPs, which confirmed that it was the donor leukocytes activated in vivo that retarded tumor growth. An in vitro analysis revealed that the IL2-FuP supported mainly proliferation and lymphokine-activated killer cell activity, whereas TNF-FuP stimulated cytokine production and cytotoxic activity of monocytes and, to a low degree, of T cells. Both TNF-FuP and IL2-FuP significantly accumulated in the tumor, which led to retardation of tumor growth. The therapeutic effect was improved by intratumoral application. Importantly, the efficacy of both IL2-FuP and TNF-FuP depended on the induction of an immune response in vivo.

Interlaboratory evaluation of a new reverse transcriptase polymerase chain reaction-based enzyme-linked immunosorbent assay for the detection of circulating melanoma cells: a multicenter study of the Dermatologic Cooperative Oncology Group.

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.

[Ocular manifestation of progressive essential telangiectases]. / Manifestation oculaire de télangiectasies essentielles progressives.

Generalized essential telangiectasia is a rare skin disorder characterized by the development of dilated venules beginning at the lower extremities and progressively spreading out to the rest of the body. It is not related to any known affection and is therefore considered to be essential. The condition tends to affect mostly women in their midthirties. We report the case of a 49-year-old healthy man worried by the progressive extension of dilated vessels on the conjunctivae of both eyes and the progressive worsening at the condition over the last five years. On clinical examination he presented also telangiectasies on both arms and legs which had also grown worse over the last five years. All further investigations and complementary examinations showed no disease. A cutaneous biopsy showed dilated venules in the superior dermis supporting the diagnosis of telangiectases. Several differential diagnoses are discussed. With confirmation of the clinical diagnosis we started treatment with oral tetracycline. Because of the poor response we stopped treatment after a period of three months.

Cloning of the human homologue of the metastasis-associated rat C4.4A.

We have previously described a rat metastasis-associated molecule, C4.4A, which has some common features with the uPAR. Because of its restricted expression in non-transformed tissues a search for the human homologue became of interest. Human C4.4A was cloned from a placental cDNA library. As in the rat, the human uPAR and the human C4.4A genes appear to belong to the same family. Both genes are located on chromosome 19q13.1-q13.2 and both molecules have a glycolipid anchor site and are composed of three extracellular domains. Only domains one and two of the human C4.4A and the uPAR protein show a significant degree of identity. Expression of the human C4.4A was observed by RT-PCR and Northern blotting in placental tissue, skin, esophagus and peripheral blood leukocytes, but not in brain, lung, liver, kidney, stomach, colon and lymphoid organs. Yet, tumors derived from the latter tissues frequently contained C4.4A mRNA. As demonstrated for malignant melanoma, C4.4A mRNA expression correlated with tumor progression. While nevi were negative and only a minority of primary malignant melanoma expressed C4.4A, all metastases were C4.4A-positive. Taking into account the high degree of homology between rat and human C4.4A, the conformity of the expression profiles and the association of rat C4.4A with tumor progression, human C4.4A might well become a prognostic marker and possibly a target of therapy.

Upregulation of C4.4A expression during progression of melanoma.

We have previously described the human homolog of a rat metastasis-associated molecule, hC4.4 A, with a weak homology to the urokinase-type plasminogen activator receptor. By the restricted expression in nontransformed tissues as opposed to expression in roughly 50% of a variety of carcinoma lines of different origins, a possible correlation between hC4.4 A and tumor progression emerged. This was explored in more detail in melanoma by quantitative polymerase chain reaction and in situ hybridization. As shown before, normal human skin weakly expresses hC4.4 A. Melanocytes and nevi are negative, but up to 60% of primary malignant melanoma and 100% of lymph node and skin metastases of melanoma are hC4.4 A positive. Signal intensity in both polymerase chain reaction and in situ hybridization varied considerably between individual samples, which is indicative for regulated expression of hC4.4 A. To test the hypothesis, melanoma lines were incubated with human serum. Whereas expression of hC4.4 was not influenced by heat-inactivated human serum, all melanoma lines responded to noninactivated human serum with upregulation of hC4.4 A expression. Regulated expression with highest level expression on metastases is a feature that hC4.4 A shares with the urokinase-type plasminogen activator receptor. This feature points towards functional activity of hC4.4 A in tumor progression.

Facts and pitfalls in the detection of tyrosinase mRNA in the blood of melanoma patients by RT-PCR.

Reverse transcription (RT) of tyrosinase mRNA and specific cDNA amplification to facilitate the early detection of circulating tumor cells in melanoma patients have been reported. The significance and practical value of these procedures for the diagnosis of tumor dissemination in melanoma patients is, however, still unclear. We analyzed peripheral blood samples of melanoma patients of different clinical stages for the presence of tyrosinase mRNA by reverse transcriptase polymerase chain reaction (RT-PCR). In addition to a nested RT-PCR-based system, we evaluated the new PCR enzyme-linked immunosorbent assay tyrosinase system for sensitivity and specificity in detecting circulating melanoma cells. Our results showed a high sensitivity and specificity for this system in detecting one melanoma cell in 1 ml of whole blood. Using different methods of detection, no tyrosinase mRNA was detectable in blood samples of patients with primary melanoma and regional lymph node metastases. In a small number of patients with visceral metastases (10-30%), we found tyrosinase mRNA-positive results. Analyses of different blood samples taken at 2-h intervals indicate that tumor cells persist only transiently in the peripheral blood. Successful establishment of melanoma cell growth from tyrosinase mRNA-positive samples indicates that viable tumor cells exist in melanoma patients' peripheral blood. Our results indicate a low amount of tyrosinase-specific transcripts in a small subset of stage IV patients and suggest that the analysis of tyrosinase mRNA in peripheral blood samples is not helpful as a prognostic marker or monitoring tool in melanoma patients.

Treatment with an anti-CD44v10-specific antibody inhibits the onset of alopecia areata in C3H/HeJ mice.

A murine CD44v10-neutralizing antibody has been reported to impair delayed-type hypersensitivity reactions. Because alopecia areata is characterized by a delayed-type hypersensitivity-like T cell mediated immune response, we addressed the question whether an anti-CD44v10-antibody influences the onset of alopecia areata. Therefore, we used the C3H/HeJ mouse model with the induction of alopecia areata in unaffected mice by the grafting of lesional alopecia areata mouse skin. Six grafted mice were injected (intraperitoneally) with anti-CD44v10, six grafted mice with anti-CD44standard, and six with phosphate-buffered saline only. After 11 wk phosphate-buffered saline injected animals on average had developed alopecia areata on 36.8% of their body. The onset of hair loss was slightly delayed and its extent reduced to 17.2% of their body in anti-CD44standard-treated mice. By contrast, five of six anti-CD44v10-treated mice did not show any hair loss and one mouse developed alopecia areata on only 1% of the body. Immunohistochemical examination revealed a marked reduction of perifollicular CD8+ lymphocytes and, to a lesser degree, CD4+ cells as well as a decreased expression of major histocompatibility complex class I on hair follicle epithelium in anti-CD44v10-treated mice as compared with phosphate-buffered saline or anti-CD44 standard-treated mice. Our data show that anti-CD44v10 is able to inhibit the onset of alopecia areata in C3H/HeJ mice. This might be accomplished by an anti-CD44v10-triggered impairment of immune cell homing (e.g., CD8+ T cells), resulting in a decrease of their number in target tissues.

Use of high-dose methylprednisolone pulse therapy in patients with progressive and stable vitiligo.

BACKGROUND: Although there are several treatment options available for patients with generalized vitiligo, their efficacy is still a matter of debate. Although shown to be effective, corticosteroids applied either systemically or topically carry the risk of significant side-effects in long-term therapy. We evaluated the effectiveness of intravenous methylprednisolone (8 mg/kg body weight) administered on three consecutive days in patients with generalized vitiligo. MATERIALS AND METHODS: A total of 14 patients with progressive or static vitiligo were included in a prospective, open, clinical study. RESULTS: Eighty-five per cent of the patients presenting with progressive disease showed cessation of disease progression after the infusion therapy. Repigmentation was observed in 71% of patients with progressive vitiligo. None of the six patients presenting with static disease showed any repigmentation in response to this form of treatment. The therapy was well tolerated in all but one patient who developed intermittent arterial hypertension during therapy. CONCLUSIONS: High-dose glucocorticoid pulse therapy may represent a therapeutic option in patients with generalized progressive vitiligo, and should be further evaluated in a prospective, randomized, clinical trial.

Interferon-alpha-associated development of bullous lesions in mycosis fungoides.

We report a 67-year-old woman with mycosis fungoides (MF) who was receiving subcutaneous injections of recombinant interferon alpha-2b (IFN-alpha). After 2 months of IFN-alpha treatment, bullous lesions appeared on her trunk and extremities. A skin biopsy specimen from the trunk revealed histopathologic features of bullous MF. Bullous lesions with specific infiltrates of MF are very rare; to the best of our knowledge, this is the first case showing specific bullous lesions of MF developed under IFN-alpha treatment.

Randomized phase III study of temozolomide versus dacarbazine in the treatment of patients with advanced metastatic malignant melanoma.

PURPOSE: To compare, in 305 patients with advanced metastatic melanoma, temozolomide and dacarbazine (DTIC) in terms of overall survival, progression-free survival (PFS), objective response, and safety, and to assess health-related quality of life (QOL) and pharmacokinetics of both drugs and their metabolite, 5-(3-methyltriazen-1-yl)imidazole-4-carboximide (MTIC). PATIENTS AND METHODS: Patients were randomized to receive either oral temozolomide at a starting dosage of 200 mg/m(2)/d for 5 days every 28 days or intravenous (IV) DTIC at a starting dosage of 250 mg/m(2)/d for 5 days every 21 days. RESULTS: In the intent-to-treat population, median survival time was 7.7 months for patients treated with temozolomide and 6.4 months for those treated with DTIC (hazards ratio, 1.18; 95% confidence interval [CI], 0.92 to 1.52). Median PFS time was significantly longer in the temozolomide-treated group (1.9 months) than in the DTIC-treated group (1.5 months) (P =.012; hazards ratio, 1.37; 95% CI, 1.07 to 1.75). No major difference in drug safety was observed. Temozolomide was well tolerated and produced a noncumulative, transient myelosuppression late in the 28-day cycle. The most common nonhematologic toxicities were mild to moderate nausea and vomiting, which were easily managed. Temozolomide therapy improved health-related QOL; more patients showed improvement or maintenance of physical functioning at week 12. Systemic exposure (area under the curve) to the parent drug and the active metabolite, MTIC, was higher after treatment with oral temozolomide than after IV administration of DTIC. CONCLUSION: Temozolomide demonstrates efficacy equal to that of DTIC and is an oral alternative for patients with advanced metastatic melanoma.

The CD44 variant isoforms CD44v6 and CD44v7 are expressed by distinct leukocyte subpopulations and exert non-overlapping functional activities.

We have described recently that anti-CD44s, anti-CD44v6 and anti-CD44v7 interfere with delayed-type hypersensitivity (DTH) reactions. Yet, TNBS-induced colitis can be cured only by anti-CD44v7. To clarify the mechanisms underlying the divergent functional activities of CD44v6 and CD44v7 we explored their contribution to lymphocyte activation in vivo and in vitro. CD44v6 and CD44v7 are distinctly expressed on subpopulations of activated lymphocytes. Expression of CD44v6 is mainly restricted to T cell blasts. CD44v7 has been detected on CD4(+) cells, B cells and monocytes. Mitogenic and antigenic stimulation of lymphocytes in vitro was impaired in the presence of anti-CD44v6 and anti-CD44v7. Accordingly, anti-CD44v6 and anti-CD44v7 mitigated the DTH reaction in 2,4-dinitro-1-fluorobenzene-sensitized and challenged mice. However, the seemingly similar effects of CD44v6- and CD44v7-specific antibodies resulted from different activities. Anti-CD44v6 treatment led to a down-regulation of IL-2 and IFN-gamma production predominantly by CD8(+) cells. In anti-CD44v7-treated mice expression of IL-12 was decreased. Elevated levels of IL-10 accompanied this reduction. The latter resulted from an anti-CD44v7-mediated blockade of interactions between CD4(+) cells and monocytes as well as an active triggering of B cells. Thus, anti-CD44v6 and anti-CD44v7 interfere with lymphocyte activation at very specific points. CD44v6 functions predominantly at the T cell level. CD44v7 influences production of proinflammatory cytokines by B cells as well as an interaction between CD4(+) cells and antigen-presenting cells. As CD44 isoforms do not differ in their intracytoplasmatic tail, the distinct activities must result from expression on different leukocyte subsets and interactions with distinct ligands.

Successful treatment of progressive vitiligo with high-dose intravenous methylprednisolone 'pulse' therapy.

We report on the successful therapy of rapid progressive generalized vitiligo in a 24-year-old woman. After intravenous injection of 500 mg of methylprednisolone on 3 consecutive days, the disease progression was stopped and repigmentation was induced. The treatment was repeated monthly and after 6 cycles, repigmentation was achieved on vast parts of most vitiligo lesions. The treatment was well tolerated by the patient, and no side effects were noted. We conclude that high-dose glucocorticoid pulse therapy may represent a therapeutic alternative in selected patients with generalized rapid progressive vitiligo.

Multiple granular cell tumors and growth hormone deficiency in a child.

Granular cell tumor is an uncommon benign tumor occurring on the skin as a single nodule. Multiple tumors are very rare, particularly in children. We describe a child with multiple granular cell tumors on the skin in association with growth hormone deficiency. The occurrence of multiple granular cell tumors in association with other clinical manifestations in childhood is discussed.

Mitigation of delayed-type hypersensitivity reactions by a CD44 variant isoform v3-specific antibody: blockade of leukocyte egress.

In allergic alterations of human skin the majority of infiltrated leukocytes express CD44v3, but no other CD44 variant isoform. Vessel endothelium, too, is brightly stained with a CD44v3-specific antibody. Being concerned about therapeutic intervention, it became of importance to define whether expression of CD44v3 on the endothelial cells or on the leukocytes or on both is of functional importance. As expression of CD44v3 in the mouse on activated endothelium and on subpopulations of activated CD4+ cells, B cells and monocytes was similar to the expression in the human, we answered the question in a mouse delayed-type hypersensitivity model. The effect of anti-CD44v3 was compared with the effect of anti-CD44s and anti-CD44v10, both known to suppress delayed-type hypersensitivity reactions. Anti-CD44v3 mitigated the delayed-type hypersensitivity reaction in dinitrofluorobenzene sensitized and challenged mice comparable with anti-CD44s and anti-CD44v10. The seemingly similar effects of CD44 isoform-specific antibodies, however, resulted from a distinct modulation of response. Anti-CD44s mainly suppressed T cell activation and interleukin-2 as well as interferon-gamma expression. Anti-CD44v10 inhibited the activation of monocytes in the draining lymph nodes and in the infiltrate, which led to a strong reduction in the proinflammatory cytokines tumor necrosis factor-alpha and interleukin-12 and in edema formation. Anti-CD44v3 had only a weak effect on cytokine expression by isolated subpopulations of leukocytes, but suppressed cytokine production by helper T cells when cocultured with antigen-presenting cells, i.e., blocked an interaction between antigen-presenting cells and helper T cells. The dominating effect of anti-CD44v3, however, relied on a blockade of leukocyte extravasation. As leukocytes transferred into dinitrofluorobenzene sensitized, anti-CD44v3-treated and lethally irradiated mice did not infiltrate the sensitized skin, anti-CD44v3 most likely prevented leukocyte extravasation by blocking CD44v3 on endothelial cells.

Heterogenous susceptibility to CD95-induced apoptosis in melanoma cells correlates with bcl-2 and bcl-x expression and is sensitive to modulation by interferon-gamma.

The expression and functionality of the Fas receptor (CD95/APO-1) play an important role for the maintenance of tissue homeostasis. Various types of tumor cells have been shown to escape immune recognition by constitutive resistance to CD95-mediated apoptosis. Furthermore, several apoptosis-related proteins have been reported to influence CD95 sensitivity. We tested an unselected panel of 11 melanoma cell lines for sensitivity to CD95 and the corresponding expression of CD95, CD95L, bcl-2, bcl-x, bcl-xS, bax and FLIP proteins. Despite detection of CD95 cell-surface expression in 9 out of the 11 cell lines tested, only 3 melanoma cell lines were sensitive to anti-CD95-MAb-induced cell death. Apoptosis-related proteins CD95L, bcl-2, bcl-x, bcl-xS and bax were found to be heterogenously expressed in different melanoma cell lines tested. The susceptibility of melanoma cells to anti-CD95-MAb-mediated apoptosis was associated with low protein expression of both bcl-2 and bcl-x. The level of CD95 cell-surface expression in melanoma cells was no indicator for CD95 sensitivity. Furthermore, FLIP protein was detectable in 7 out of the 11 cell lines, but showed no correlation to CD95 sensitivity. Certain cytokines have been described as modulating the susceptibility of tumor cells to CD95-induced cell death. Since IFN-alpha was proved to be clinically efficient in melanoma therapy, we tested whether interferons have the ability to induce sensitivity to CD95 in primarily resistant melanoma cell lines. Here we show that IFN-gamma, but not IFN-alpha, is able to increase the susceptibility of sensitive cell lines and to induce CD95 sensitivity in resistant melanoma cell lines, accompanied by up-regulation of the protein expression level of CD95 and/or bcl-xS.