Robert E. Allen

The gene expression signatures of melanoma progression

Because of the paucity of available tissue, little information has previously been available regarding the gene expression profiles of primary melanomas. To understand the molecular basis of melanoma progression, we compared the gene expression profiles of a series of nevi, primary melanomas, and melanoma metastases. We found that metastatic melanomas exhibit two dichotomous patterns of gene expression, which unexpectedly reflect gene expression differences already apparent in comparing laser-capture microdissected radial and vertical phases of a large primary melanoma. Unsupervised hierarchical clustering accurately separated nevi and primary melanomas. Multiclass significance analysis of microarrays comparing normal skin, nevi, primary melanomas, and the two types of metastatic melanoma identified 2,602 transcripts that significantly correlated with sample class. These results suggest that melanoma pathogenesis can be understood as a series of distinct molecular events. The gene expression signatures identified here provide the basis for developing new diagnostics and targeting therapies for patients with malignant melanoma.

Recombinant human granulocyte macrophage-colony stimulating factor (rhGM-CSF) and autologous melanoma vaccine mediate tumor regression in patients with metastatic melanoma.

In mice, significant immunoprotection was achieved using B16 melanoma cells transfected with granulocyte-macrophage colony-stimulating factor (GM-CSF) as vaccines (Dranoff G, Jaffee E, Lazenby A, et al. Vaccination with irradiated tumor cells engineered to secrete murine granulocyte-macrophage colony-stimulating factor stimulates potent, specific, and long-lasting anti-tumor immunity. Proc Natl Acad Sci USA 1993;90:3539-43). The aim of this study is to test the hypothesis that recombinant human GM-CSF (rhGM-CSF) injected with autologous melanoma vaccine may result in tumor rejection in melanoma patients. Twenty stage IV melanoma patients were treated as outpatients with multiple cycles of autologous melanoma vaccine and bacillus Calmette-Guérin (BCG) plus rhGM-CSF injection in the vaccine sites. Two patients (10%) showed a complete response, with one patient showing resolution of subcutaneous, hepatic, and splenic metastases. In the second patient, buccal, subcutaneous, pulmonary, paraaortic, hepatic, splenic, and retroperitoneal metastases regressed completely. Two patients (10%) showed partial response, with regression of a paraaortic metastasis in one patient. In the second patient, there was shrinkage (> 75%) of a large hepatic lesion. One patient has been rendered free of disease after resection of a single pulmonary metastatic nodule. Three patients (15%) had stable disease during treatment but subsequently developed progression of disease. In 12 patients (60%), the disease progressed. Side effects were minimal. In a separate pilot study, 15 stage IV melanoma patients were also treated with autologous melanoma vaccine with BCG but not with rhGM-CSF; none responded. The fact that four patients showed objective responses to active specific immunotherapy with rhGM-CSF demonstrates that melanoma patients bearing a significant tumor burden may respond specifically to their autologous melanoma.

Clinical Significance of Occult Metastatic Melanoma in Sentinel Lymph Nodes and Other High-risk Factors Based on Long-term Follow-up

Selective sentinel lymphadenectomy (SSL) following preoperative lymphoscintigraphy is the most significant recent advance in the management of patients with primary melanoma. This study evaluates the prognostic value of sentinel lymph node (SLN) status and other risk factors in predicting survival and recurrence in patients with primary cutaneous melanoma. From October 1993 to July 1998 a series of 412 patients with primary invasive melanoma underwent SSL at the UCSF/ Mt. Zion Melanoma Center. The outcome of 363 evaluable patients is summarized in this study. The factors related to survival and disease recurrence were analyzed by Cox proportional hazard regression models. The overall incidence of patients with positive SLNs was 18%. Over a median follow-up of 4.8 years, the overall mortality rate in patients with primary cutaneous melanoma was 18.7%, and 74 recurrences occurred (20.4%). Mortality was significantly related to SLN status [HR = 2.06; 95% Confidence interval (CI) 1.18, 3.58], angiolymphatic invasion (HR = 2.21; 95% CI 1.08, 4.55), ulceration (HR = 1.79; 95% CI 1.02, 3.15), mitotic index (HR =1.38; 95% CI 1.01, 1.90), and tumor thickness (HR = 2.20, 95% CI 1.21, 3.99). Factors significantly related to disease-free survival included SLN status (HR = 2.09; 95% CI 1.31, 3.34), tumor thickness (HR = 1.89; 95%. CI 1.20,2.98), and age (HR= 1.26 95% CI 1.08, 1.47). SLN status was the most significant factor for melanoma recurrence and death. Other important predictors include tumor thickness, ulceration, lymphatic invasion, and mitotic index.

Recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF, sargramostim) administered for 3 years as adjuvant therapy of stages II(T4), III, and IV melanoma.

A hypothesis generating study was conducted to evaluate the safety and efficacy of prolonged (3 y) administration of granulocyte-macrophage colony-stimulating factor (GM-CSF, sargramostim) as surgical adjuvant therapy in patients with melanoma at high risk of recurrence. Ninety-eight evaluable patients with stages II(T4), III, or IV melanoma were given prolonged treatment with GM-CSF after surgical resection of disease. The GM-CSF was administered subcutaneously in 28-day cycles, such that a dose of 125 μg/m2 was delivered daily for 14 days followed by 14 days rest. Treatment cycles continued for 3 years or until disease recurrence, which could not be surgically excised. Patients were evaluated for toxicity, disease-free survival, and melanoma-specific survival. Prolonged administration of GM-CSF was well tolerated; grade 1 or 2 side effects occurred in 82% of the patients. There were no grade 3 or 4 treatment-related side effects. Two patients developed acute myelogenous leukemia after completion of 3 years of GM-CSF administration. With a median follow-up of 5.3 years, the median melanoma-specific survival has not yet been reached. The 5-year melanoma-specific survival rate was 60%. The current study has expanded the preliminary evidence on GM-CSF as adjuvant therapy of patients with melanoma who are at high risk for recurrence.

Heterogeneous patterns of lymphatic drainage to sentinel lymph nodes by primary melanoma from different anatomic sites

We want to define the patterns of lymphatic drainage for primary melanoma to sentinel lymph nodes (SLNs) based on a large lymphoscintigraphic database. Preoperative lymphoscintigraphy was used to identify and classify SLN drainage basins and patterns of drainage. Methods: Lymphoscintigraphy using intradermally administered technetium-99m labeled sulfur colloid was performed on 400 consecutive patients with malignant melanoma to define lymphatic drainage channels and draining SLN basins before surgery. Primary tumor sites consisted of head and neck, upper extremity, trunk, and lower extremity. Different types of drainage patterns were classified and correlated with different anatomic sites. Results: SLN(s) were identified in over 98% of the patients, whereas lymphatic drainage channels were successfully identified in 90% of the patients. Drainage from the primary site to a single SLN through a single lymphatic channel (type IA) was seen in 186 of 400 patients (47%) as the most common type. In patients with a single SLN within a single basin (type I–V), the percentage of patients with primary lesions in the head and neck, upper extremity, trunk, and lower extremity regions were 61%, 79%, 55%, and 78%, respectively. In cases of multiple lymphatic channels (type VI–VII), the percentages of patients with primary lesions in the head and neck, upper extremity, trunk, and lower extremity regions were 24%, 8%, 36%, and 19%, respectively. Conclusion: Various drainage patterns were noted from primary melanomas in different anatomic sites. Preoperative lymphoscintigraphy is important in establishing the SLN basins for harvesting the SLN(s).

Low-Dose Outpatient Chemobiotherapy With Temozolomide, Granulocyte-Macrophage Colony Stimulating Factor, Interferon-α2b, and Recombinant Interleukin-2 for the Treatment of Metastatic Melanoma

PURPOSE The objective of this study was to further investigate the efficacy and safety of low-dose outpatient chemobiotherapy in patients with unresectable metastatic melanoma. PATIENTS AND METHODS Thirty-one patients with histologically confirmed unresectable measurable metastatic melanoma were enrolled onto an open-label, multicenter phase II study. The treatment regimen consisted of oral temozolomide followed by subcutaneous biotherapy with granulocyte macrophage colony-stimulating factor, interferon-alfa, and recombinant interleukin-2 (rIL-2). RESULTS Twenty-eight patients (90%) had M1c disease, and 58% had three or more sites of metastasis. Four patients (13%), all with M1c disease, had a complete response, and four patients had a partial response. The median progression-free survival was 4.9 months and the median overall survival was 13.1 months. Two patients (6%) developed CNS metastasis as the first site of disease progression, and 7 (23%) of 30 experienced CNS progression after receiving chemobiotherapy. A total of 112 cycles of therapy were administered. Toxicity occurred in 78% of the cycles and was grade 1 or 2 in the majority of cases and easily managed. Grade 4 toxicity occurred in 3% of the cycles. CONCLUSION This low-dose chemobiotherapy combination produces clinical responses in patients with metastatic melanoma, even in those with M1c disease, is well tolerated, and allows home dosing. It offers a reasonable alternative to high-dose regimens, such as high-dose biochemotherapy or rIL-2 requiring prolonged periods of hospitalization, or single agent outpatient regimens, such as dacarbazine, which is usually not effective in patients with M1c disease. Furthermore, it may protect against the development of brain metastases.

Delayed harvesting of sentinel lymph nodes after previous wide local excision of extremity melanoma.

Background:Harvesting the sentinel lymph node (SLN) is important in the management of patients with primary cutaneous melanoma. Selective sentinel lymphadenectomy (SSL) is generally performed at the time of wide local excision (WLE). The aim of our study was to determine whether delayed SSL is useful in detecting micrometastasis to the regional basin in patients with previous WLE of an extremity melanoma.Methods:Of 203 patients with a primary melanoma site located on the upper or lower extremity seen at the University of California, San Francisco/Mount Zion Melanoma Center from May 17, 1994, to March 23, 1999, 24 patients had a WLE of their extremity melanoma with adequate margins before referral. SSL was performed to assess micrometastasis in the regional lymph node basin after preoperative lymphoscintigraphy.Results:At least 1 SLN was identified in all 24 patients. At a median follow-up of 3 years, two patients showed micrometastasis in the SLNs. One of these two patients developed recurrence, and all remaining patients showed no evidence of disease.Conclusions:Although it is generally advised that WLE should be performed simultaneously with SSL, delayed SSL after WLE of an extremity melanoma can still provide valuable staging information, which is critical for management of the patient.

A randomized, double-blind, placebo-controlled trial of transfer factor as adjuvant therapy for malignant melanoma

One hundred and sixty‐eight evaluable patients participated in a randomized, double‐blind study of transfer factor (TF) versus placebo as surgical adjuvant therapy of Stage I and Stage II malignant melanoma. Eighty‐five patients received TF prepared from the leukocytes of healthy volunteer donors; eighty‐three participants received placebo. Therapy was initiated within 90 days of resection of all evident tumor and continued until 2 years of disease‐free survival or the occurrence of unresectable dissemination of melanoma. Known prognostic variables were similarly distributed in the treatment and control groups, documenting the randomization efficacy. Three endpoints were analyzed: disease‐free interval, time to Stage III metastasis, and survival. After a median follow‐up period of 24.75 months, there was a trend in favor of the placebo group with regard to all three endpoints and this was significant (P ≦ 0.05) for time to Stage III metastasis. These findings indicate that TF is not effective as surgical adjuvant therapy of malignant melanoma.

Phase II study of low-dose outpatient chemobiotherapy with temozolomide (TMZ), granulocyte macrophage colony stimulating factor (GM-CSF), Interferon-alpha-2b (IFN), and Interleukin-2 (IL-2) for the treatment of metastatic melanoma

7546 Background: Metastatic melanoma responds poorly to currently available therapies, many of which require hospitalization. de Gast reported that chemobiotherapy with TMZ, GM-CSF, IL-2, and IFN i...

INV 23 Granulocyte-macrophage colony-stimulating factor (GM-CSF) as adjuvant therapy of melanoma

remains unconfirmed. Cases of vulval melanoma were associated with HPV-16 DNA. In addition, VZV exhibits tropism for the melanocyte (R. Harson and C. Grose, 1995). Although none of these clinical or in-vitro observations establish a viral cause for cutaneous melanoma, they nevertheless support the hypothesis of viral involvement. On this basis, sera of patients with melanoma and additional unrelated cancers have been stored for viral studies. This hypothesis was conceived when a patient with melanoma of the nasal mucosa in the author’s series, reported the contemporaneous development of nasal melanoma on his dog. The search for a viral cause in the pathogenesis of melanoma, a hitherto neglected approach in favour of the solar hypothesis, will facilitate the rational development of effective vaccines against the disease.