Trends in hepatocyte growth factor, insulin-like growth factor 1, thyroid-stimulating hormone, and leptin expression levels in uveal melanoma patient serum and tumor tissues: correlation to disease progression.

This exploratory study was carried out to determine the expression levels of hepatocyte growth factor (HGF), insulin-like growth factor 1, thyroid-stimulating hormone (TSH), and leptin in serum and tumor samples from patients with uveal melanoma and to investigate the potential association of these expression levels with disease progression and patient survival. Seventeen patients, including nine nonmetastatic and eight metastatic, were included in the study. Eighteen healthy individuals served as controls. The levels of these four proteins in serum and tissue samples were determined by enzyme-linked immunosorbent assays and immunohistochemical staining, respectively. Associations between protein levels and survival, disease progression, and other clinicopathological factors were analyzed statistically. Serum levels of HGF were significantly higher and TSH levels were lower in uveal melanoma patients than in healthy individuals, but the level of neither protein differed significantly between metastatic and nonmetastatic groups. Of the four proteins tested, only serum TSH was significantly associated with patient survival. No correlation was observed between the tissue and serum levels of each protein. The levels of HGF in serum may be markers of uveal melanoma development. The prognostic and predictive values of these potential markers need to be determined in a larger cohort.

Elevated Serum Leptin Levels are Associated With an Increased Risk of Sentinel Lymph Node Metastasis in Cutaneous Melanoma.

The metabolic hormone leptin has been implicated in the pathogenesis of various malignancies and may contribute to the high rate of cancer in obese individuals. We reported that leptin and its receptor are expressed by melanoma tumors and cell lines, and that leptin stimulates proliferation of cultured melanoma cells. Here, we tested the hypothesis that leptin contributes to early melanoma progression by assessing its association with sentinel node positivity in cutaneous melanoma patients. The study enrolled 72 patients who were scheduled to undergo lymphatic mapping and sentinel node biopsy. Fasting blood was obtained before surgery, and serum leptin levels were measured by enzyme-linked immunosorbent assay (ELISA) with a "raw" (assay value) and an "adjusted" value (raw value divided by body mass index). Leptin levels and other clinicopathologic parameters were compared between sentinel node positive and negative groups. Logistic regression models were used to predict sentinel node status using leptin and other relevant clinical parameters. The raw and adjusted leptin levels were significantly higher in the 15 patients with positive sentinel nodes. These findings could not be attributed to differences in body mass indices. Univariate models revealed raw leptin, adjusted leptin, Breslow thickness, and mitotic rate as significant predictors of sentinel node status. Leptin levels and Breslow thickness remained significant in multivariate models. Survival and follow-up analysis revealed more aggressive disease in diabetic patients. Elevated serum leptin levels predict sentinel node metastasis in melanoma. Validation of this finding in larger cohorts should enable better stratification of early stage melanoma patients.

Association of activated c-Met with NRAS-mutated human melanomas.

Cutaneous melanomas can be divided into three mutually exclusive genetic subsets: tumors with mutated BRAF, tumors with mutated NRAS and tumors wild type at both loci (wt/wt). Targeted therapy for melanoma has been advancing with agents directed to mutated BRAF, accounting for 50% of melanoma patients. The c-Met pathway is known to play a role in melanoma tumorigenesis and preliminary data from our laboratory suggested that this pathway is preferentially activated in NRAS-mutated tumors. The objective of this study was to test the hypothesis that melanomas carrying the mutated NRAS genotype are uniquely sensitively to c-Met inhibition, thus providing rationale for therapeutic targeting of c-Met in this patient cohort. Using primary human melanomas with known BRAF/NRAS genotypes, we observed greater immunostaining for phosphorylated (activated) c-Met in NRAS-mutated and wt/wt tumors, compared to BRAF-mutated tumors. NRAS-mutated and wt/wt cell lines also demonstrated more robust c-Met activation in response to hepatocyte growth factor (HGF). Knock-down of mutated N-Ras, but not wild type N-Ras, by RNA interference resulted in decreased c-Met phosphorylation. Compared to BRAF mutants, NRAS-mutated melanoma cells were more sensitive to pharmacologic c-Met inhibition in terms of c-Met activation, Akt phosphorylation, tumor cell proliferation, migration and apoptosis. This enhanced sensitivity was observed in wt/wt cells as well, but was a less consistent finding. On the basis of these experimental results, we propose that c-Met inhibition may be a useful therapeutic strategy for melanomas with NRAS mutations, as well as some tumors with a wt/wt genotype.

Clinical correlates of NRAS and BRAF mutations in primary human melanoma.

PURPOSE: NRAS and BRAF mutations are common in cutaneous melanomas, although rarely detected mutually in the same tumor. Distinct clinical correlates of these mutations have not been described, despite in vitro data suggesting enhanced oncogenic effects. This study was designed to test the hypothesis that primary human cutaneous melanomas harboring mutations in NRAS or BRAF display a more aggressive clinical phenotype than tumors wild type at both loci. EXPERIMENTAL DESIGN: Microdissection of 223 primary melanomas was carried out, followed by determination of the NRAS and BRAF mutational status. Genotypic findings were correlated with features known to influence tumor behavior including age, gender, Breslow depth, Clark level, mitotic rate, the presence of ulceration, and American Joint Committee on Cancer (AJCC) staging. RESULTS: Breslow depth and Clark level varied significantly among the genotypes, with NRAS mutants showing the deepest levels and wild-type tumors the least depth. Ulceration also differed significantly among the genotypes, with BRAF mutants demonstrating the highest rate. In addition, tumors with mutated NRAS were more likely to be located on the extremities. Patients whose tumors carried either mutation presented with more advanced AJCC stages compared with patients with wild-type tumors, and specifically, were more likely to have stage III disease at diagnosis. Overall survival did not differ among the 3 groups. CONCLUSIONS: Distinct clinical phenotypes exist for melanomas bearing NRAS and BRAF mutations, whether considered together or separately, and are associated with features known to predict aggressive tumor behavior. The impact of these mutations is most evident at earlier stages of disease progression.

Promotion of melanoma growth by the metabolic hormone leptin.

We have previously shown that melanoma cells proliferate in response to the metabolic hormones TRH and TSH. The objective of the present study was to test the hypothesis that a third metabolic hormone, leptin, serves as a growth factor for melanoma. Using western blotting, indirect immunofluorescence, and RT-PCR, leptin receptors were found to be expressed by human melanoma cells. In contrast, cultured melanocytes expressed message for the receptor without detectable protein. Melanoma cells responded to treatment with leptin by activating the MAPK pathway and proliferating. Melanoma cells but not melanocytes, also expressed leptin protein, creating a potential autocrine loop. Examination of human melanoma tumors by immunohistochemistry revealed that melanomas and nevi expressed leptin at a high frequency. Melanomas also strongly expressed the leptin receptor, whereas nevi expressed this receptor to a much lesser degree. We conclude that leptin is a melanoma growth factor and that a leptin autocrine-loop may contribute to the uncontrolled proliferation of these cells.

Autologous tumor-derived heat-shock protein peptide complex-96 (HSPPC-96) in patients with metastatic melanoma.

BACKGROUND: Glycoprotein-96, a non-polymorphic heat-shock protein, associates with intracellular peptides. Autologous tumor-derived heat shock protein-peptide complex 96 (HSPPC-96) can elicit potent tumor-specific T cell responses and protective immunity in animal models. We sought to investigate the feasibility, safety, and antitumor activity of HSPPC-96 vaccines prepared from tumor specimens of patients with metastatic melanoma. METHODS: Patients with a Karnofsky Performance Status >70% and stage III or stage IV melanoma had to have a metastasis >3 cm in diameter resectable as part of routine clinical management. HSPPC-96 tumor-derived vaccines were prepared in one of three dose levels (2.5, 25, or 100 microg/dose) and administered as an intradermal injection weekly for 4 consecutive weeks. In vivo induction of immunity was evaluated using delayed-type hypersensitivity (DTH) to HSPPC-96, irradiated tumor, and dinitrochlorobenzene (DNCB). The gamma-interferon (IFNgamma) ELISPOT assay was used to measure induction of a peripheral blood mononuclear cell response against autologous tumor cells at baseline and at the beginning of weeks 3, 4, and 8. RESULTS: Among 36 patients enrolled, 72% had stage IV melanoma and 83% had received prior systemic therapy. The smallest tumor specimen from which HSPPC-96 was prepared weighed 2 g. Twelve patients (including 9 with stage IV and indicator lesions) had a negative DNCB skin test result at baseline. All 36 patients were treated and evaluable for toxicity and response. There were no serious toxicities. There were no observed DTH responses to HSPPC-96 or to autologous tumor cells before or during treatment. The IFNgamma-producing cell count rose modestly in 5 of 26 patients and returned to baseline by week 8, with no discernible association with HSPPC-96 dosing or clinical parameters. There were no objective responses among 16 patients with stage IV disease and indicator lesions. Among 20 patients treated in the adjuvant setting, 11 with stage IV melanoma at baseline had a progression-free and overall survival of 45% and 82%, respectively, with a median follow-up of 10 years. CONCLUSION: Treatment with autologous tumor-derived HSPPC-96 was feasible and safe at all doses tested. Observed immunological effects and antitumor activity were modest, precluding selection of a biologically active dose. Nevertheless, the 25-microg dose level was shown to be practical for further study.

Genetic deficiency of complement isoforms C4A or C4B predicts improved survival of metastatic renal cell carcinoma.

PURPOSE: Autoimmune phenomena during immunotherapy are associated with favorable outcomes in patients with metastatic renal cell carcinoma. We have reported improved survival in patients with stage IV renal cell carcinoma who carry autoimmunity associated HLA class II haplotypes. We propose that the clinical benefit is mediated by products of other autoimmunity associated genes linked to these haplotypes. A candidate gene is complement C4, which replicates as part of the RCCX module, can be present in multiple copies and exists as C4A and C4B isoforms. Deficiencies of either isoform are associated with autoimmunity. In the current study we tested the hypothesis that C4A or C4B deficiency predicts improved survival of patients with RCC. MATERIALS AND METHODS: The total C4 copy number was determined by simultaneous amplification of RP1 and TNXA/RP2 to quantitate RCCX modules. C4A and C4B alleles were distinguished by PshAI restriction fragment length polymorphism. RESULTS: Genetic complotypes were determined in 61 patients. Individuals with a solitary copy of either C4 isoform experienced longer survival. Median survival from the diagnosis of metastatic disease in patients with a solitary copy of C4A or C4B was 7.75 years vs 1.25 in the comparison group (p = 0.001). This was independent of the benefit derived from autoimmune class II genotypes. CONCLUSIONS: Improved survival is seen in patients with C4A or C4B deficiency and renal cell carcinoma treated with cytokine therapy with or without surgery. These data support our hypothesis that patients with renal cell carcinoma who have autoimmune genotypes have favorable outcomes resulting from autoimmune mechanisms directed to the tumor.

Lack of maturation with anti-leptin receptor antibody in melanoma but not in nevi.

We have previously shown thryotropin-releasing hormone expression in nevi and melanoma. Thryotropin-releasing hormone regulation by leptin has been shown in the hypothalamus. The present study was therefore undertaken to evaluate leptin and leptin receptor in nevi and melanoma. Leptin receptor expression as assessed using an anti-leptin receptor antibody showed uniform expression throughout the lesion in 14 of 17 melanomas; 3 melanomas lacked leptin receptor immunoreactivity. In contrast, out of 15 nevi, 10 showed weak to moderate leptin receptor immunoreactivity, with positivity present only in the superficial dermal component. Thus, maturation was present in nevi but not in melanoma with the anti-leptin receptor antibody (P<0.0001). Anti-leptin antibody, in contrast, did not show a significant difference in maturation between nevi and melanoma. We also compared leptin receptor in Spitz nevi and melanoma, as the two can sometimes be difficult to distinguish. Spitz nevi showed moderate to strong immunopositivity. Of 19 Spitz nevi, 7 showed lack of maturation, a finding statistically significant from both melanoma and nevi. Our results suggest a role for leptin receptor in melanoma, and we show for the first time that melanomas show more intense immunoreactivity as compared to nevi (but not Spitz nevi) and that maturation with anti-leptin receptor antibody may be a diagnostically useful tool in distinguishing melanomas, especially nevoid ones, from nevi in difficult cases.

Frequencies of NRAS and BRAF mutations increase from the radial to the vertical growth phase in cutaneous melanoma.

A lack of consensus exists with regards to the relative rates of NRAS and BRAF mutations in the radial (RGP) and vertical (VGP) growth phases of individual melanoma tumors. This study was conducted to test the hypothesis that mutations are acquired with progression from RGP to VGP. Using laser capture microdissection, pure tumor DNA was obtained from 15 in situ melanomas, and from the RGP and VGP of 29 invasive tumors. NRAS exon 2 and BRAF exon 15 DNA were amplified by PCR and sequenced. Mutations were present in 6 of 15 in situ melanomas (40%). Of 29 invasive tumors, 16 exhibited RGP mutations (55.2%); 22 showed VGP mutations (75.9%). Paired RGP/VGP mutation analysis revealed a trend toward discordance in the distribution of mutations, favoring VGP localization (P=0.07). Of 15 samples, 12 with mutations in both phases had an increased proportion of mutated DNA in the VGP, measured on DNA chromatograms (P=0.08). Limitations of this study include a relatively small sample cohort selected for technical reasons from a larger population, presenting the risk of selection bias. These concerns notwithstanding our findings support the hypothesis that NRAS and BRAF mutations increase with tumor progression from superficial to invasive disease. JID JOURNAL CLUB ARTICLE: For questions, answers, and open discussion about this article, please go to http://network.nature.com/group/jidclub.

NF-kappaB mediates mitogen-activated protein kinase pathway-dependent iNOS expression in human melanoma.

Tumor expression of inducible nitric oxide synthase (iNOS) predicts poor outcomes for melanoma patients. We have reported the regulation of melanoma iNOS by the mitogen-activated protein kinase (MAPK) pathway. In this study, we test the hypothesis that NF-kappaB mediates this regulation. Western blotting of melanoma cell lysates confirmed the constitutive expression of iNOS. Western blot detected baseline levels of activated nuclear extracellular signal-regulated kinase and NF-kappaB. Indirect immunofluorescence confirmed the presence of NF-kappaB p50 and p65 in melanoma cell nuclei, with p50 being more prevalent. Electrophoretic mobility shift assay demonstrated baseline NF-kappaB activity, the findings confirmed by supershift analysis. Treatment of melanoma cells with the MEK inhibitor U0126 decreased NF-kappaB binding to its DNA recognition sequence, implicating the MAPK pathway in NF-kappaB activation. Two specific NF-kappaB inhibitors suppressed iNOS expression, demonstrating regulation of iNOS by NF-kappaB. Several experiments indicated the presence of p50 homodimers, which lack a transactivation domain and rely on the transcriptional coactivator Bcl-3 to carry out this function. Bcl-3 was detected in melanoma cells and co-immunoprecipitated with p50. These data suggest that the constitutively activated melanoma MAPK pathway stimulates activation of NF-kappaB hetero- and homodimers, which, in turn, drive iNOS expression and support melanoma tumorigenesis.

Changes in pERK1/2 and pAKT expression in melanoma lesions after imatinib treatment.

Response to treatment with imatinib mesylate has been associated in preclinical models with the inhibition of two signaling pathways that promote cellular survival - the phosphatidylinositol 3-kinase/AKT pathway and the mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) pathway. We sought to evaluate the extent of inhibition of these two pathways in metastatic melanoma specimens from patients treated with imatinib. Metastatic melanoma tumor samples were obtained before and during the second week of imatinib treatment from patients enrolled in a phase II study. A tissue microarray was constructed using formalin-fixed, paraffin-embedded tissues, and immunohistochemical analysis was performed using standard techniques to detect phosphorylated (p) ERK1/2 and pAKT expression. Of 21 patients who were treated with imatinib, tumor samples adequate for analysis were available both at baseline and during the second week of treatment from 10 patients for pERK1/2 expression and from nine patients for pAKT expression. No consistent pattern of change in pAKT or pERK expression after treatment with imatinib was observed. No apparent correlation between the clinical benefit of imatinib treatment and changes in pAKT and pERK1/2 expression was observed. A better understanding of the AKT and mitogen-activated protein kinase pathways is needed to optimize the clinical benefit of targeted therapy, such as imatinib.

Human melanoma cells express functional receptors for thyroid-stimulating hormone.

We have reported a high prevalence of hypothyroidism in the cutaneous melanoma population, suggesting that the pathologic hormonal environment of hypothyroidism promotes melanoma growth. The objective of this study was to test the hypothesis that TSH, which circulates at elevated levels in hypothyroid individuals, stimulates the growth of melanoma cells. Our results show that TSH receptors (TSHR) are expressed by virtually all cutaneous melanocytic lesions, including benign nevi, dysplastic nevi, and melanomas, with higher expression found in malignant and pre-malignant lesions. The finding of TSHR expression by human tumors is confirmed in cultured melanoma cells and melanocytes, in which TSHR expression is demonstrated by immunofluorescent staining, western blotting, and reverse transcriptase-PCR. Melanoma TSHR are functional, as evidenced by the ability of TSH to induce the formation of cAMP and to activate the mitogen-activated protein kinase (MAPK) pathway. Cultured melanoma cells, but not melanocytes, are induced to proliferate at a physiologically relevant concentration of TSH. Taken together, these data support the hypothesis that TSH is a growth factor for human melanoma. Our findings have broad clinical implications for the prevention of melanoma and the management of established disease.

Tumor iNOS predicts poor survival for stage III melanoma patients.

Inducible nitric oxide synthase (iNOS) produces nitric oxide, which has growth promoting activity in melanoma. A preliminary study of tumors from patients with Stage III melanoma who had received neo-adjuvant therapy revealed an association of tumor iNOS expression with shortened survival. The objective of the present study was to determine whether iNOS expression in tumors of newly diagnosed, untreated Stage III patients is predictive of survival. iNOS expression was examined by immunohistochemistry in tumors from 132 patients. The staining was evaluated for percentage of positive cells (Number score) and the intensity of staining (Intensity score). The association of iNOS expression with overall and disease-specific survival was tested in univariate and multivariate Cox proportional hazards regression models that included other known prognostic factors. Results of the univariate analysis demonstrated that the presence of iNOS in a patient's tumor, whether graded on the basis of Number or Intensity score, was associated with a significant increase in the hazard ratio of death from melanoma. These findings were corroborated by median survival data estimated from Kaplan Meier analysis. In the multivariate model including iNOS number or intensity, gender, age, number of lymph nodes, macroscopic disease and in-transit disease, only iNOS expression predicted survival. We conclude that a significant association exists between tumor iNOS expression and shortened survival in untreated Stage III melanoma patients. The ability of iNOS to predict outcomes for these patients may be independent of other known prognostic factors, providing a new molecular marker with significant potential for clinical utility.

Expression of thyrotropin-releasing hormone by human melanoma and nevi.

PURPOSE: Thyrotropin-releasing hormone (TRH) is a tripeptide hormone produced by the hypothalamus in response to hypothyroidism. RNA transcripts for the TRH prohormone have recently been described in melanoma cell lines. To expand these findings, we have examined cultured melanoma cells and melanocytes, human melanoma tumors, and nevi for the expression of TRH. EXPERIMENTAL DESIGN: Five melanoma cell lines were analyzed by reverse transcription-PCR/Southern blotting for preproTRH message. The same melanoma lines and two melanocyte lines were examined by immunocytochemistry for TRH protein expression and for growth response to exogenous TRH. Immunohistochemistry was used to test for TRH protein in sections of 19 melanomas, 33 dysplastic nevi, and 27 benign nevi. RESULTS: TRH message and protein were detected in all melanoma cell lines examined. Melanocytes were also found to express TRH protein. Four of the five melanoma cell lines but neither melanocyte line responded with a increase in proliferation to low concentrations of exogenous TRH. TRH immunoreactivity was observed in 12 of 19 melanomas (63%), 23 of 33 (69.7%) dysplastic nevi, and 14 of 27 (51.9%) benign nevi. Expression in dysplastic nevi was significantly greater than in benign nevi. Upon separate analysis of nevi from melanoma patients, the difference between dysplastic and benign nevi was even more significant. However, in healthy individuals, no difference between dysplastic and benign nevi was observed. Furthermore, dysplastic nevi from melanoma patients had a significantly higher percentage of TRH-positive cells when compared with healthy individuals. CONCLUSIONS: TRH is commonly expressed by melanomas and dysplastic nevi and may function as a melanoma autocrine growth factor. The presence of TRH in dysplastic nevi may be predictive for the development of melanoma. Our findings have significant clinical and biological implications for future research into the early stages of melanoma initiation and progression.

High prevalence of hypothyroidism among patients with cutaneous melanoma.

We have previously reported a high prevalence of hypothyroidism among patients with uveal melanoma. The objectives of the present study were to determine if a similar pattern of thyroid pathology exists among patients with cutaneous melanoma as well. To address this question, the medical records of all patients registered at the University of Texas M.D. Anderson Cancer Center with a diagnosis of cutaneous melanoma during the years 1997 and 1998 were examined for a history of overt hypothyroidism, defined as a requirement for thyroid hormone replacement. Data regarding stage and site of the primary tumor were obtained for these patients and for age/gender matched euthyroid controls from the same melanoma study population. Among 1,580 cutaneous melanoma patients (948 M/632 F), 111 (7.0%) gave a history of hypothyroidism [23/948 M (2.4%) and 88/632 F (13.9%)]. The prevalences of hypothyroidism for both males and females were significantly higher than those reported for the general population. Characteristics of the primary tumor did not differ between cases and controls, although there was a trend for a lower rate of primary tumor ulceration among the hypothyroid case subjects. We conclude that hypothyroidism of varied etiologies is common among patients with cutaneous melanoma. These data suggest that melanoma may be responsive to hormones of the thyroid hormone control loop, raising many questions of clinical and biologic importance.

Heterozygosity or homozygosity for 2 HLA class II haplotypes predict favorable outcomes for renal cell carcinoma treated with cytokine therapy.

PURPOSE: Durable responses to cytokine therapy occur in a small subset of patients with renal cell carcinoma. We determined if a common HLA genotype existed among these patients which might be associated with response and survival. MATERIALS AND METHODS: The study population consisted of 80 patients with metastatic renal cell carcinoma who had received cytokine therapy. DNA obtained from these patients was used for high resolution typing of HLA A, B, C, DRB1, DQA1 and DQB1 alleles. RESULTS: The class II alleles from patients with prolonged disease-free survival were predominantly composed of haplotype DRB1*0301/DQA1*0501/DQB1*0201 and DRB1*1501/DQA1*0102/DQB1*0602. The frequency of heterozygosity or homozygosity for these alleles was significantly greater in the good outcome group of patients than in those whose disease progressed during therapy. Heterozygosity or homozygosity at these loci was also associated with significant prolongation of survival. CONCLUSIONS: We conclude that heterozygosity or homozygosity for the class II haplotypes DRB1*0301/DQA1*0501/DQB1*0201 and DRB1*1501/DQA1*0102/DQB1*0602 is associated with durable response and survival in patients with metastatic renal cell carcinoma treated with cytokine therapy.

Negative association of melanoma differentiation-associated gene (mda-7) and inducible nitric oxide synthase (iNOS) in human melanoma: MDA-7 regulates iNOS expression in melanoma cells.

The melanoma differentiation association gene-7 (mda-7) is a novel tumor suppressor gene of which the protein expression decreases to nearly undetectable levels in metastatic melanoma. In contrast, expression of inducible nitric oxide synthase (iNOS) is increased in advanced stages of melanoma, and iNOS expression has been proposed as a potential prognostic marker in this disease. Thus, expression of these molecules in the same tumor appears to exhibit reciprocal characteristics. We hypothesize that the relative ratios of these melanoma progression molecules may define either tumor progression or tumor suppression in human melanoma. The first goal of this study was to determine whether MDA-7 expression in melanoma negatively correlates with iNOS expression. The second goal was to determine whether iNOS expression could be regulated by MDA-7 expression in melanoma cells. Expression of MDA-7 and iNOS proteins were evaluated by immunohistochemistry in a total of 81 tumor samples: 38 primary melanomas and 43 metastatic melanomas. Evaluation of these melanoma patient samples showed that expression of MDA-7 and iNOS exhibits a significant negative correlation (P = 0.03). In vitro studies revealed that iNOS expression in melanoma cell lines is lost in a dose-dependent fashion after treatment with an adenoviral vector encoding the mda-7 gene (Ad-mda7) or with rhMDA-7 protein, demonstrating that MDA-7 down-regulates iNOS expression. Furthermore, we demonstrate that the STAT-3-modulated expression of IFN regulatory factors 1 and 2 is regulated by MDA-7, which may alter iNOS gene expression. These studies demonstrate that expression of the MDA-7 tumor suppressor can negatively regulate iNOS expression in malignant melanoma cell lines.

Phase I and pharmacokinetic study of ABI-007, a Cremophor-free, protein-stabilized, nanoparticle formulation of paclitaxel.

PURPOSE: ABI-007 is a novel Cremophor-free, protein-stabilized, nanoparticle formulation of paclitaxel. The absence of Cremophor EL may permit ABI-007 to be administered without the premedications used routinely for the prevention of hypersensitivity reactions. Furthermore, this novel formulation permits a higher paclitaxel concentration in solution and, thus, a decreased infusion volume and time. This Phase I study examines the toxicity profile, maximum tolerated dose (MTD), and pharmacokinetics of ABI-007. EXPERIMENTAL DESIGN: ABI-007 was administered in the outpatient setting, as a 30-min infusion without premedications. Doses of ABI-007 ranged from 135 (level 0) to 375 mg/m2 (level 3). Sixteen patients participated in pharmacokinetic studies. RESULTS: Nineteen patients were treated. No acute hypersensitivity reactions were observed during the infusion period. Hematological toxicity was mild and not cumulative. Dose-limiting toxicity, which occurred in 3 of 6 patients treated at level 3 (375 mg/m2), consisted of sensory neuropathy (3 patients), stomatitis (2 patients), and superficial keratopathy (2 patients). The MTD was thus determined to be 300 mg/m2 (level 2). Pharmacokinetic analyses revealed paclitaxel C(max) and area under the curve(inf) values to increase linearly over the ABI-007 dose range of 135-300 mg/m2. C(max) and area under the curve(inf) values for individual patients correlated well with toxicity. CONCLUSIONS: ABI-007 offers several features of clinical interest, including rapid infusion rate, absence of requirement for premedication, and a high paclitaxel MTD. Our results provide support for Phase II trials to determine the antitumor activity of this drug.

Phase II trial of 9-nitrocamptothecin (RFS 2000) for patients with metastatic cutaneous or uveal melanoma.

The camptothecin derivative 9-nitrocamptothecin (9-NC) has demonstrated clinical activity in patients with ovarian and pancreatic carcinomas. Preclinical studies have shown promising activity of 9-NC for melanoma. We have thus conducted a phase II clinical trial of 9-NC for patients with metastatic cutaneous and uveal melanoma. Twenty-eight patients were enrolled in the trial, with diagnoses evenly divided between the two types of melanoma. 9-NC was administered orally at a starting dose of 1.5 mg/m(2)/day for 5 consecutive days of each week. No complete or partial responses were observed. Stabilization of disease was achieved in four individuals (15%) for durations of 3, 4, 6 and 8 months. Hematologic toxicity was moderate. Gastrointestinal side effects were common with 43% of the patients experiencing grade 3 or 4 diarrhea and 18% reporting grade 3 or 4 vomiting. In contrast to other 9-NC clinical trials, no patients developed chemical cystitis with gross hematuria. We conclude that, in keeping with the general chemoresistance of melanoma, 9-NC at the dose and schedule studied in this trial is significantly toxic and is not active for metastatic melanoma of cutaneous or uveal origin.

Loss of MDA-7 expression with progression of melanoma.

PURPOSE: Ectopic transfer of the melanoma differentiation-associated gene-7 (mda-7) has been shown in vitro to suppress growth and induce apoptosis in a variety of human tumor cell lines; similar effects are not elicited in normal cells. Thus, the mda-7 gene seems to function as a novel tumor suppressor, and there is interest in the potential of mda-7 gene transfer as cancer therapy. The objective of this study was to determine if MDA-7 protein is lost during primary melanoma progression from superficial to invasive stages and from localized to metastatic tumor. As a secondary objective, we analyzed MDA-7 protein expression in primary melanomas for correlation with predictors of outcome and with survival. MATERIALS AND METHODS: MDA-7 protein expression was evaluated by immunohistochemistry in 41 primary melanomas and 41 metastases, including 24 paired samples. Each sample was scored for the percentage of positive cells and the overall intensity of immunolabeling. RESULTS: Significant decreases in MDA-7 immunostaining, reflected in both number and intensity scores, were observed when comparing the intraepidermal and superficially invasive portions with the deeply invasive portions of primary tumors. Significant differences were also observed when comparing primary tumors to paired metastases. CONCLUSION: Downregulation of MDA-7 expression in primary melanomas facilitates progression to invasive and metastatic stages. These data support the development of Ad-mda7 as gene therapy for advanced melanoma.