A phase 2 trial of complete resection for stage IV melanoma: results of Southwest Oncology Group Clinical Trial S9430.

BACKGROUND: On the basis of retrospective experience at individual centers, it appears that patients with stage IV melanoma who undergo complete resection have a favorable outcome compared with patients with disseminated stage IV disease. The Southwest Oncology Group (SWOG) performed a prospective trial in patients with metastatic melanoma who were enrolled before complete resection of their metastatic disease and provided prospective outcomes in the cooperative group setting. METHODS: Based on their physical examination and radiologic imaging studies, patients with a stage IV melanoma judged amenable to complete resection underwent surgery within 28 days of enrollment. All eligible patients were followed with scans (computed tomography or positron emission tomography) every 6 months until relapse and death. RESULTS: Seventy-seven patients were enrolled from 18 different centers. Of those, 5 patients were ineligible; 2 had stage III disease alone; and 3 had no melanoma in their surgical specimen. In addition, 8 eligible patients had incompletely resected tumor. Therefore, the primary analysis included 64 completely resected patients. Twenty patients (31%) had visceral disease. With a median follow-up of 5 years, the median relapse-free survival was 5 months (95% CI, 3-7 months) whereas median overall survival was 21 months (95% CI, 16-34 months). Overall survivals at 3 and 4 years were 36% and 31%, respectively. CONCLUSIONS: In a prospective multicenter setting, appropriately selected patients with stage IV melanoma achieved prolonged overall survival after complete surgical resection. Although median relapse-free survival was only 5 months, patients could still frequently undergo subsequent surgery for isolated recurrences. This patient population is appropriate for aggressive surgical therapy and for trials evaluating adjuvant therapy.

A case of recurrent pseudolymphomatous folliculitis: A mimic of cutaneous lymphoma.

Pseudolymphomatous folliculitis is a rare entity. We present a 62-year-old man with a recurrent solitary nodule on his nose requiring multiple excisions. Microscopic examination of the excisions showed a dense lymphocytic infiltrate containing numerous histiocytes and S100+, CD1a+ dendritic cells that surrounded and infiltrated hypertrophic hair follicles. Diffuse sheets of CD3+ T cells and nodular clusters of CD20+ B cells were also seen. There was normal reactive pattern of follicular centers. Light chain restriction was not detected. T-cell receptor and immunoglobulin heavy chain gene rearrangements by polymerase chain reaction revealed negative findings. A diagnosis of pseudolymphomatous folliculitis was made based on the hypertrophic hair follicles, periadnexal S100+ and CD1a+ dendritic cells, and negative clonal gene rearrangement study findings. This case of recurrent pseudolymphomatous folliculitis is instructive because of the resemblance to cutaneous lymphomas and cutaneous lymphoid hyperplasias, and the need for correct diagnosis to avoid overtreatment of this indolent condition.

Detection of copy number alterations in metastatic melanoma by a DNA fluorescence in situ hybridization probe panel and array comparative genomic hybridization: a southwest oncology group study (S9431).

PURPOSE: Gene copy number alteration (CNA) is common in malignant melanoma and is associated with tumor development and progression. The concordance between molecular cytogenetic techniques used to determine CNA has not been evaluated on a large set of loci in malignant melanoma. EXPERIMENTAL DESIGN: A panel of 16 locus-specific fluorescence in situ hybridization (FISH) probes located on eight chromosomes was used to identify CNA in touch preparations of frozen tissue samples from 19 patients with metastatic melanoma (SWOG-9431). A subset (n = 11) was analyzed using bacterial artificial chromosome (BAC) array comparative genomic hybridization (aCGH) of DNA isolated directly from touch-preparation slides. RESULTS: By FISH, most samples showed loss near or at WISP3/6p21, CCND3/6q22, and CDKN2A/9p21 (>75% of samples tested). More than one third of CDKN2A/9p21 losses were biallelic. Gains of NEDD9/6p24, MET/7q31, and MYC/8q24 were common (57%, 47%, and 41%, respectively) and CNA events involving 9p21/7p12.3 and MET were frequently coincident, suggesting gain of the whole chromosome 7. Changes were confirmed by aCGH, which also uncovered many discreet regions of change, larger than a single BAC. Overlapping segments observed in >45% of samples included many of the loci analyzed in the FISH study, in addition to other WNT pathway members, and genes associated with TP53 pathways and DNA damage response, repair, and stability. CONCLUSIONS: This study outlines a set of CNAs at the gene and regional level, using FISH and aCGH, which may provide a benchmark for future studies and may be important in selection of individual therapy for patients with metastatic malignant melanoma.

Failure of senescence in the dysplasia-melanoma sequence: demonstration using a tissue microarray and a revised paradigm for melanoma.

In order to correlate changes in morphology to changes in molecular attributes, we constructed a tissue microarray of thin and thick melanomas selected to represent progression from dysplasia to early and advanced melanoma. Hematoxylin and eosin (H&E) staining and immunohistology with antibodies to cyclin D1, p16, Ki67, and Bcl-2 were performed. Observations were interpreted using a revised paradigm for the dysplasia-melanoma sequence in which the early steps of melanomatous growth develop in an accretive fashion similar to the growth of the common acquired nevus. The co-expression of cyclin D1 and p16 persisted from dysplasia to early melanomatous vertical growth. Malignant transformation characterized by absence of p16 and presence of increased cyclin D1 and increased Ki67 and confirmed by clinically documented metastasis occurred during the process of evolving melanomatous vertical growth. The interplay of mutated BRAF, cyclin D1, and p16 with anti-apoptosis and failure of senescence may account for the existence of nevi and dysplastic nevi and for their relationship to melanoma, and may indirectly account for the infrequency of nevi in the lentiginous melanomas that lack mutated BRAF. These observations suggest a need for more detailed study of transformation to malignancy in the various subsets of melanoma.

Adjuvant immunotherapy of resected, intermediate-thickness, node-negative melanoma with an allogeneic tumor vaccine: overall results of a randomized trial of the Southwest Oncology Group.

PURPOSE: Patients with clinically negative nodes constitute over 85% of new melanoma cases. There is no adjuvant therapy for intermediate-thickness, node-negative melanoma patients. PATIENTS AND METHODS: The Southwest Oncology Group conducted a randomized phase III trial of an allogeneic melanoma vaccine for 2 years versus observation in patients with intermediate-thickness (1.5 to 4.0 mm or Clark's level IV if thickness unknown), clinically or pathologically node-negative melanoma (T3N0M0). RESULTS: Six hundred eighty-nine patients were accrued over 4.5 years; 89 patients (13%) were ineligible. Surgical node staging was performed in 24%, the remainder were clinical N0. Thirteen eligible patients refused assigned treatment: seven on the observation arm and six on the vaccine arm. Most vaccine patients experienced mild to moderate local toxicity, but 26 (9%) experienced grade 3 toxicity. After a median follow-up of 5.6 years, there were 107 events (tumor recurrences or deaths) among the 300 eligible patients randomized to vaccine compared with 114 among the 300 eligible patients randomized to observation (hazard ratio, 0.92; Cox-adjusted P(2) = 0.51). There was no difference in vaccine efficacy among patients with tumors < or = 3 mm or > 3 mm. CONCLUSION: This represents one of the largest randomized, controlled trials of adjuvant vaccine therapy in human cancer reported to date. Compliance with randomization was excellent, with only 2% refusing assigned therapy. There is no evidence of improved disease-free survival among patients randomized to receive vaccine, although the power to detect a small but clinically significant difference was low. Future investigations of adjuvant vaccine approaches for patients with intermediate-thickness melanoma should involve larger numbers of patients and ideally should include sentinel node biopsy staging.

Resistance to interferons in melanoma cells does not correlate with the expression or activation of signal transducer and activator of transcription 1 (Stat1).

Defects in expression or activation signal transducer and activator of transcription-1 (Stat1) in response to interferon-alpha2 (IFN-alpha2) have been implicated as a mechanism for IFN resistance in melanoma cells. To further determine the significance of this observation, 17 melanoma cell lines sensitive or resistant to the antiproliferative effects of IFN-alpha2 and IFN-beta, as well as 30 melanoma patient samples, were analyzed for Stat1 levels by either Western blot analysis or immunohistochemistry. Although the expression level varied between samples, all the cell lines except one and all melanoma biopsy specimens expressed Stat1. IFN-stimulated levels of Stat1 and Stat2, which constitute the transcriptional activation complexes, such as, gamma activated factor (GAF) and IFN-stimulated gene factor 3 (ISGF3), for IFN-stimulated gene (ISG) induction were assessed in melanoma cell lines. Both IFN-alpha2 and INF-beta induced equivalent amounts of Stat1 and Stat2 proteins in cell lines, although compared with IFN-alpha2, IFN-beta had greater antiproliferative effects. No significant differences were observed in tyrosine or serine phosphorylation of Stat1 or the formation of GAF or ISGF3 complexes following IFN-alpha2 or IFN-beta treatment of IFN-resistant or IFN-sensitive cell lines. Comparable induction of two ISGs, ISG54 and IFN regulatory factor-1 (IRF-1), was observed in both sensitive WM9 and resistant A375 cells. Therefore, we report that defects in expression or activation of Stat1 or Stat2 were infrequent in melanoma cell lines and tumor samples and did not correlate with IFN resistance. Cellular resistance to IFNs likely results from defective quantitative or qualitative expression of specific ISGs.

Risk assessment in localized primary cutaneous melanoma: a Southwest Oncology Group study evaluating nine factors and a test of the Clark logistic regression prediction model.

We studied 9 clinical and pathologic factors in 259 patients using Cox model regression analysis to determine which factors have independent predictive value. Median follow-up time in all patients still alive was 12.3 years (range, 1.7 to 16.7 years). Tumor-infiltrating lymphocytes (P = .005), primary site (P = .006), and thickness (P = .02) had independent predictive value. Ulceration (P = .06) and age (P = .07) had marginal value. We used 6 of those factors to test the Clark logistic regression prediction model, which accurately predicted 8-year survival in 121 (72.9%) of 166 patients and accurately predicted melanoma-specific mortality in 32 (43%) of 74 patients. The combined or overall accuracy of the Clark model was only 64%.

Adjuvant vaccine immunotherapy of resected, clinically node-negative melanoma: long-term outcome and impact of HLA class I antigen expression on overall survival.

Associations between HLA class I antigen expression and the efficacy of a melanoma vaccine (Melacine; Corixa Corp.) were initially described in stage IV melanoma. Similar associations were observed in S9035, a phase III adjuvant trial evaluating Melacine for 2 years compared with observation in patients with stage II melanoma. This report provides long-term results. The effects of treatment on relapse-free survival (RFS) and overall survival (OS) were evaluated, and prespecified analyses investigated associations between treatment and HLA expression. Multivariable analyses were adjusted for tumor thickness, ulceration and site, method of nodal staging, and sex. P = 0.01 was considered statistically significant in subset analyses to account for multiple comparisons. For the entire study population of 689 patients, there were no significant differences in RFS or OS by treatment arm. HLA serotyping was performed on 553 (80%) patients (vaccine, 294; observation, 259). Among the subpopulation with HLA-A2 and/or HLA-Cw3 serotype, vaccine arm patients (n = 178) had marginally improved RFS (adjusted P = 0.02) and significantly improved OS compared with observation arm patients (n = 145), with 10-year OS of 75% and 63%, respectively [hazard ratio (HR), 0.62; 99% confidence interval (CI), 0.37-1.02; P = 0.01]. There was no impact of HLA-A2 and/or HLA-Cw3 expression on observation arm patients. An analysis of mature data from S9035 indicates a significant OS benefit from adjuvant vaccine therapy for patients with HLA-A2- and/or HLA-Cw3-expressing melanoma. The possibility of interactions between HLA type and outcome should be considered in future immunotherapy trials. Further investigations of melanoma-associated antigens present in Melacine and presented by HLA-A2 and HLA-Cw3 may be warranted.

Southwest Oncology Group S0008: a phase III trial of high-dose interferon Alfa-2b versus cisplatin, vinblastine, and dacarbazine, plus interleukin-2 and interferon in patients with high-risk melanoma--an intergroup study of cancer and leukemia Group B, Children's Oncology Group, Eastern Cooperative Oncology Group, and Southwest Oncology Group.

PURPOSE: High-dose interferon (IFN) for 1 year (HDI) is the US Food and Drug Administration-approved adjuvant therapy for patients with high-risk melanoma. Efforts to modify IFN dose and schedule have not improved efficacy. We sought to determine whether a shorter course of biochemotherapy would be more effective. PATIENTS AND METHODS: S0008 (S0008: Chemotherapy Plus Biological Therapy in Treating Patients With Melanoma) was an Intergroup phase III trial that enrolled high-risk patients (stage IIIA-N2a through IIIC-N3), randomly assigning them to receive either HDI or biochemotherapy consisting of dacarbazine, cisplatin, vinblastine, interleukin-2, IFN alfa-2b (IFN-α-2b) and granulocyte colony-stimulating factor given every 21 days for three cycles. Coprimary end points were relapse-free survival (RFS) and overall survival (OS). RESULTS: In all, 432 patients were enrolled. Grade 3 and 4 adverse events occurred in 57% and 7% of HDI patients and 36% and 40% of biochemotherapy patients, respectively. At a median follow-up of 7.2 years, biochemotherapy improved RFS (hazard ratio [HR], 0.75; 95% CI, 0.58 to 0.97; P = .015), with a median RFS of 4.0 years (95% CI, 1.9 years to not reached [NR]) versus 1.9 years for HDI (95% CI, 1.2 to 2.8 years) and a 5-year RFS of 48% versus 39%. Median OS was not different (HR, 0.98; 95% CI, 0.74 to 1.31; P = .55), with a median OS of 9.9 years (95% CI, 4.62 years to NR) for biochemotherapy versus 6.7 years (95% CI, 4.5 years to NR) for HDI and a 5-year OS of 56% for both arms. CONCLUSION: Biochemotherapy is a shorter, alternative adjuvant treatment for patients with high-risk melanoma that provides statistically significant improvement in RFS but no difference in OS and more toxicity compared with HDI.

Acantholytic dyskeratosis occurring within an epidermal nevus.

A 25-year-old woman presented with a large area of flesh-colored verrucous plaques following the lines of Blaschko on the left side of the body that had been present since 6 months of age. The plaques had been stable and grew proportionately with the patient's body until she reached 20 years of age when they began to thicken and enlarge. Her medical and family history was unremarkable. A shave biopsy revealed a papillomatous epidermis with 3 discrete foci of acantholytic dyskeratosis, with corps ronds and grains that were similar to the histologic findings of Darier disease (DD). Epidermolytic hyperkeratosis was not identified. Our patient's lack of a family history of DD, early-onset disease, and linear presentation along the lines of Blaschko all favored a diagnosis of acantholytic dyskeratotic epidermal nevi (ADEN) versus localized DD.

A unique cause of intestinal and splenic infarction in a sickle cell trait patient.

Sickle-cell trait is a common genetic abnormality in the African American population. A sickle-cell crisis in a patient with sickle-cell trait is uncommon at best. Abdominal painful crises are typical of patients with sickle cell anemia. The treatment for an abdominal painful crisis is usually medical and rarely surgical. We present the case of a cocaine-induced sickle-cell crisis in a sickle-cell trait patient that resulted in splenic, intestinal, and cerebral infarctions and multisystem organ failure necessitating a splenectomy, subtotal colectomy, and small bowel resection. This case highlights the diagnostic dilemma that abdominal pain can present in the sickle-cell population and illustrates the importance of recognizing the potential for traditionally medically managed illnesses to become surgical emergencies.

Re-inventing intratumoral immunotherapy for melanoma.

Immunotherapeutics have been applied intratumorally to manage accessible lesions and to induce systemic immunity in malignant melanoma. Intratumoral bacillus Calmette-Guérin (BCG) has been used for 40 years, and intratumoral BCG, IL-2, IFN-α and imiquimod are recommended as treatment options for patients with in-transit melanoma metastases. Regression of cutaneous metastases can be achieved. Subcutaneous metastases are more refractory, and regression of uninjected, visceral metastases is infrequent. Other microbial products, cytokines, chemicals, immune cells, antibody and viral and plasmid vectors expressing immunologically active molecules have been tested. Antitumor activity has not been demonstrated to be superior to that of intratumoral BCG. There are few controlled trials, and whether survival is impacted with any approach has not yet been established. The immunotherapeutics applied and the intratumoral administration procedure itself can activate responses that are immune inhibitory. More rigorous clinical testing and improved understanding and modulation of regulatory immune responses are necessary.

Adenoid cystic carcinoma of the base of the tongue: Late metastasis to the pancreas.

Adenoid cystic carcinoma (ACC) is a relatively rare epithelial tumor of the salivary glands. We present a 64-year-old gentleman with ACC of the tongue who following resection and radiotherapy, presented 10 years later with a lung metastasis and underwent operative intervention and further radiotherapy. Five years later he presented with obstructive jaundice found to be metastatic ACC. We believe this to be the first report of an ACC metastasizing to the pancreas.

A phase II trial of imatinib mesylate in merkel cell carcinoma (neuroendocrine carcinoma of the skin): A Southwest Oncology Group study (S0331).

BACKGROUND: Imatinib mesylate (Gleevec) was evaluated as a treatment for Merkel cell carcinoma (MCC, neuroendocrine carcinoma of the skin) based on the identification of strong c-KIT staining of these neoplasms. METHODS: Eligibility included patients with measurable metastatic or unresectable MCC, c-KIT (CD117) expression and a Zubrod performance status of 0 to 2. Imatinib 400 mg daily was administered orally in 28-day cycles to 23 patients. RESULTS: Overall, imatinib was well tolerated with grade 1 or 2 nausea, diarrhea, and hematologic toxicity as the most frequent side effects. A partial response was seen in 1 patient (4%; 95% CI: 0%-22%). Median progression-free survival was 1 month (95% CI: 1-2 months). Median overall survival was 5 months (95% CI: 2-8 months). One patient achieved a partial response and another had prolonged disease stabilization while receiving treatment. CONCLUSIONS: The majority of patients progressed rapidly within 1 to 2 cycles of treatment. The observed progression-free survival and overall survival were not adequate to conclude that this agent was active in advanced MCC, and thus the planned second stage of patient accrual was not opened.

Dual-color, break-apart fluorescence in situ hybridization for EWS gene rearrangement distinguishes clear cell sarcoma of soft tissue from malignant melanoma.

Clear cell sarcoma of soft tissue (malignant melanoma of soft parts) is a soft tissue sarcoma with melanocytic differentiation that typically occurs in the tendons and aponeuroses of young adults. As demonstrated by cytogenetics and reverse-transcriptase polymerase chain reaction, between 70% and over 90% of clear cell sarcomas have a t(12;22) translocation, fusing the EWS and ATF1 genes on chromosomes 22q12 and 12q13, respectively. Identification of this translocation distinguishes clear cell sarcoma from histologic mimics, most importantly conventional malignant melanoma. We report our experience with a commercially available, dual-color, break-apart fluorescence in situ hybridization (FISH) probe, which allows detection of EWS (22q12) gene rearrangement in formalin-fixed, paraffin-embedded tissues. Histologically and immunophenotypically well-characterized cases of clear cell sarcoma (n = 10) and malignant melanoma (n = 32) were evaluated with a 22q12 dual-color, break-apart probe (Vysis, Downer's Grove, IL, USA), which spans the known common breakpoints in the EWS gene on chromosome 22 (introns 7-10). Signals from tumor cell nuclei were counted under a fluorescence microscope and the presence of red-green break-apart signals was recorded. Of the clear cell sarcoma cases, seven of 10 showed evidence of an EWS gene rearrangement with a mean of 81.6% positive cells per sample (range: 60-95%). All cases of malignant melanoma (n = 32) showed virtually absent break-apart signals in the EWS gene (less than 4% cells per case). FISH detects EWS gene rearrangement in a substantial proportion of clear cell sarcomas, with excellent specificity. Importantly, EWS FISH is negative in malignant melanoma, a clinically dissimilar tumor, which may closely mimic clear cell sarcoma histologically and immunohistochemically. As the studied probe can be utilized in routinely processed tissue, FISH provides an excellent alternative to reverse-transcriptase polymerase chain reaction in cases where fresh tissue is unavailable.

Efficient identification of T-cell clones associated with graft-versus-host disease in target tissue allows for subsequent detection in peripheral blood.

Graft-versus-host disease (GVHD) causes severe morbidity and mortality in allogeneic haematopoietic stem cell transplantation (HSCT) because of destruction of recipient tissues by donor alloreactive T cells. We hypothesized that GVHD-specific T-cell clones are expanded within affected tissue of HSCT patients and can also be detected in blood at the time of active disease. A multiplex polymerase chain reaction (PCR) was used to amplify T-cell receptor (TCR) variable beta (VB) chain rearrangements in skin biopsies from eight allogeneic HSCT patients. Molecular analysis of the complementarity-determining region 3 (CDR3) of amplified products defined expanded, potentially disease-associated 'clonotypes' and enabled the design of clonotype-specific PCR assays. We detected immunodominant clones in seven of eight GVHD-positive skin biopsies. In serial skin biopsies from the same patient, the identical clone was found in each biopsy. In a patient who underwent two successive HSCTs from different donors, distinct clones were identified for each engraftment. Using clonotypic PCR assays, individual tissue-derived clones could be identified in peripheral blood samples obtained during active GVHD. We hypothesize that clonotypic sequences derived from target tissue can serve as markers for GVHD and may have utility in diagnosis and monitoring response to therapy, as well as enable future therapies targeted against pathogenic clones.

Loss of heterozygosity, microsatellite instability, and mismatch repair protein alterations in the radial growth phase of cutaneous malignant melanomas.

Little is known about genomic alterations during development of the radial growth phase (RGP) of cutaneous malignant melanomas (CMMs). In this investigation polymerase chain reaction-based microsatellite assays were applied to analyze 13 RGP-CMMs with 18 microsatellite markers at six chromosomal regions: 1p, 3p, 4q, 6q, 9p, and 10q. Loss of heterozygosity (LOH) was found in eight cases (62%), at 9p22, 1p36, and 10q11, suggesting the presence of tumor-suppressor genes at these regions. LOH was encountered frequently at the interferon-alpha (31%) and D10S249 loci (15%). Low-level microsatellite instability (MSI) (11-16% of investigated loci unstable) was noted in three cases (23%). Two MSI banding patterns were seen: band shift and the presence of additional bands. To investigate the underlying mechanisms of the low-level MSI pattern, we analyzed the lesions for expression of mismatch repair (MMR) proteins with immunoperoxidase methods and mouse monoclonal antibodies. The average percentages of positively stained cells for human MutL homolog 1 (hMLH1), human MutS homolog 2 (hMSH2), and human MutS homolog 6 (hMSH6) in RGP-CMM (75.6 +/- 3.4%, 67.20 +/- 7.71%, and 76.6 +/- 2.1%, respectively) were reduced compared with benign nevi. No statistically significant differences in MMR protein expression were found between microsatellite-stable and low-level MSI lesions (P = 0.173, P = 0.458, and P = 0.385 for hMLH1, hMSH2, and hMSH6, respectively). There was a direct correlation between values for percentages of positively stained cells for hMSH2 and hMSH6 (r = +0.9, P = 0.03), suggesting that common mechanisms regulate their expression. In conclusion, LOH, MSI, and reduced MMR protein expression appear to be present in at least some RGP-CMMs and may play a role in their pathogenesis. Further studies are necessary to support these finding and to determine their diagnostic and prognostic significance.