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BackgroundSeveral protein markers, including vimentin, have been used to diagnose human melanoma. Because melanoma often has metastasized by the time of diagnosis, early markers prognostic for metastatic potential need to be identified. Commonly, vimentin is found in mesenchymal cells, and keratins are present in epithelial cells, but recent studies report coexpression of vimentin and keratin(s) in epithelial and nonepithelial neoplasms, including some melanomas.PurposeOur purpose was to determine whether coexpression of vimentin and keratin(s) is correlated with tumor cell invasion and metastatic behavior.MethodsWe evaluated nine human melanoma cell lines expressing vimentin and other markers of aggressive tumor behavior (HMB-45, S-100, HLA-ABC class I and HLA-DR class II histocompatibility antigens, and K8 and K18 keratins). Levels of K8 and K18 keratins were determined in the highly metastatic C8161 cell line, the poorly metastatic A375P line, and the moderately metastatic A375M line. To determine whether the presence of keratin affects migratory ability, we altered the conformational structure of keratin filaments in C8161 cells by transfection with a mutant K18 complementary DNA. We also determined messenger RNA levels of human type IV collagenase, an enzyme marker for invasion and metastasis.ResultsIn A375P cells, two-dimensional electrophoresis with Coomassie-stained gels, immunoblotting, and immunofluorescence staining showed no detectable levels of K8 or K18. A375M cells showed low levels of K8 and K18 by Western and Northern blotting, with a distinctive fluorescent subpopulation of cells. In comparison, K8 and K18 levels in C8161 cells were high in all cells. Type IV collagenase messenger RNA levels were lowest in A375P cells and highest in C8161 cells, correlating with invasive ability in vitro and metastatic potential in athymic nude mice. The transfectant clones C1070-10 and C1070-14 derived from the C8161 parent line showed dramatic morphological changes, disrupted keratin filaments, and decreased invasive and metastatic potential directly correlated with a reduction in migratory activity.ConclusionThese findings show a correlation between the coexpression of vimentin with K8 and K18 keratins and the invasive and metastatic behavior of three representative human melanoma cell lines.

The optimal staging and treatment of the internal mammary nodes (IMNs) among patients with invasive breast cancer (IBC) is controversial. Although medial tumors have been reported to more commonly drain to IMNs, other variables predictive for IMN drainage may help identify those patients who may benefit from further IMN assessment. Factors associated with IMN drainage were analyzed among 141 patients who underwent lymphatic mapping and selective sentinel lymphadenectomy using intradermal injection (ID) or peritumoral (PT) injection. Fourteen of 83 patients (17%) receiving PT injections had IMN drainage, compared to none among the 58 patients who underwent ID injection alone (p = 0.0004). There were no differences in patient or tumor variables detected between the two groups. Among patients receiving PT injections, no factors examined were significantly associated with IMN drainage on univariate analysis. Using the multivariate logistic regression model, palpable disease was the most important factor associated with IMN drainage (risk ratio [RR] = 6.02; 95% confidence interval [CI] 0.64-56.34; p = 0.05). In addition, lymphatic/vascular invasion (LVI) and age less than 50 years were associated with IMN drainage (RR = 6.17; 95% CI 1.02-37.50; p = 0.09 and RR = 2.94; 95% CI 0.82-10.49; p = 0.09, respectively). IMN drainage occurred in a significant proportion of patients after PT injection, but not ID injection. In the final model, palpable disease was the most important factor associated with IMN drainage; LVI and age less than 50 years were of borderline significance. These factors may aid in the selection of patients who might benefit from further staging or treatment of the IMNs. [ABSTRACT FROM AUTHOR]

Background: Guidelines for melanoma recommend sentinel lymph node biopsy (SLNB) in patients with melanomas ≥1 mm thickness. Recent single institution studies have found tumors <1.5 mm a low-risk group for positive SLNB., Methods: A retrospective review of the Sentinel Lymph Node Working Group multicenter database identified patients with intermediate thickness melanoma (1.01-4.00 mm) who had SLNB, and assessed predictors for positive SLNB., Results: 3460 patients were analyzed, 584 (17%) had a positive SLNB. Univariate factors associated with a positive SLNB included age <60 (p < .001), tumor on the trunk/lower extremity (p < .001), Breslow depth ≥2 mm (p < .001), ulceration (p < .001), mitotic rate ≥1/mm 2 (p = .01), and microsatellitosis (p < .001). Multivariate analysis revealed age, location, and Breslow depth as significant predictors. Patients ≥75 with lesions 1.01-1.49 mm on the head/neck/upper extremity and 1.5-1.99 mm without high-risk features had <5% risk of SLN positivity., Conclusions: Intermediate thickness melanoma has significant heterogeneity of SLNB positivity. Low-risk subgroups can be found among older patients in the absence of high-risk features., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Purpose: To validate the prognostic impact of combined expression levels of three markers (SPP1, RGS1, and NCOA3) in melanoma specimens from patients enrolled in the E1690 clinical trial of high-dose or low-dose IFNα-2b versus observation. Experimental Design: Tissue was available from 248 patients. Marker expression was determined by digital imaging of immunohistochemically stained slides. The prognostic impact of each marker was first assessed by recording its expression value relative to the median. A multimarker index was then developed to combine marker expression levels by counting for each patient the number of markers with high expression. The impact of the multimarker index on relapse-free survival (RFS) and overall survival (OS) was assessed using Kaplan-Meier analysis, and both univariate and multivariate Cox regression analyses. Results: By Kaplan-Meier analysis, high multimarker expression scores were significantly predictive of RFS ( P < 0.001) and OS ( P < 0.001). Stepwise multivariate Cox regression analysis with backward elimination that included routine clinical and histologic prognostic factors revealed high multimarker expression scores and tumor thickness as the only factors significantly and independently predicting RFS and OS. Stepwise multivariate Cox regression analyses that also included treatment type and number of positive lymph nodes generated identical results for both RFS and OS. In the molecularly defined low-risk subgroup, patients treated with high-dose IFN had a significantly improved RFS compared with patients in the other two subgroups ( P < 0.05). Conclusions: These results validate the independent impact of combined expression levels of SPP1, RGS1, and NCOA3 on survival of melanoma in a prospectively collected cohort. Clin Cancer Res; 23(22); 6888-92. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Melanoma patients with additional positive lymph nodes in the completion lymph node dissection (CLND) following a positive sentinel lymph node (SLN) biopsy would have a poorer prognosis than patients with no additional positive lymph nodes. We hypothesize that the progression of disease from the SLN to the non-SLN compartment is orderly and is associated with the worsening of the disease status. Thus, the SLN and non-SLN compartments are biologically different in that cancer cells, in general, arrive in the SLN compartment before spreading to the non-SLN compartment. To validate this concept, we used a large cohort of melanoma patients from our prospective SLN database in an academic tertiary medical center. Adult cutaneous melanoma patients (n = 291) undergoing CLND after a positive SLN biopsy from 1994 to 2009 were analyzed. Comparison of 5-year disease-free survival and 5-year overall survival between positive (n = 66) and negative (n = 225) CLND groups was made. The 5-year disease-free survival rates were 55% (95% CI 49-62%) for patients with no additional LN on CLND versus 14% (95% CI 8-26%) in patients with positive LN on CLND (p < 0.0001, log-rank test). The median disease-free survival time was 7.4 years with negative CLND (95% CI 4.4-15+ years) and 1.2 years with positive CLND (95% CI 1.0-1.8 years). The 5-year overall survival rates were 67% (95% CI 61-74%) for negative CLND versus 38% (95% CI 28-52%) for positive CLND (p < 0.0001, log-rank test). The median overall survival time was 12.1 years for negative CLND (95% CI 9.3-15+ years) and 2.5 years for positive CLND (95% CI 2.2-5.7 years). This study shows that CLND status is a significant prognostic factor for patients with positive SLNs undergoing CLND. Also, it suggests an orderly progression of metastasis from the SLN to the non-SLN compartment. Thus, the SLN in the regional nodal basin draining the primary melanoma may serve as an important gateway for metastasis to the non-SLN compartment and beyond to the systemic sites.

Objective: There is currently no consensus regarding how to address pelvic sentinel lymph nodes (PSLNs) in melanoma. Thus, our objectives were to identify the incidence and clinical impact of PSLNs., Methods: Retrospective review of a prospectively collected multi-institutional melanoma database., Results: Of 2476 cases of lower extremity and trunk melanomas, 227 (9%) drained to PSLNs (181 to both PSLNs and superficial (inguinal or femoral) sentinel lymph nodes (SSLN) and 46 to PSLNs alone). Seventeen (7.5%) of 227 PSLN cases were positive for nodal metastasis, 8 of which drained to PSLNs only while 9 drained to both PSLNs and SSLNs. Complication rates between PSLN and SSLN biopsy were similar (15% vs. 14% respectively). In 181 cases with drainage to both SSLNs and PSLNs, PSLN biopsy upstaged one patient (0.6%), and completion dissection based on a positive PSLN did not upstage any., Conclusions: PSLN biopsy is safe, however in the setting of negative SSLNs there is minimal clinical impact. We therefore recommend PSLN biopsy when the SSLNs are positive or when the tumor drains to PSLNs alone., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Ulceration is an important prognostic factor in melanoma whose biologic basis is poorly understood. Here we assessed the prognostic impact of pleckstrin homology domain-interacting protein (PHIP) copy number and its relationship to ulceration. PHIP copy number was determined using fluorescence in situ hybridization (FISH) in a tissue microarray cohort of 238 melanomas. Elevated PHIP copy number was associated with significantly reduced distant metastasis-free survival (DMFS; P=0.01) and disease-specific survival (DSS; P=0.009) by Kaplan-Meier analyses. PHIP FISH scores were independently predictive of DMFS (P=0.03) and DSS (P=0.03). Increased PHIP copy number was an independent predictor of ulceration status (P=0.04). The combined impact of increased PHIP copy number and tumor vascularity on ulceration status was highly significant (P<0.0001). Stable suppression of PHIP in human melanoma cells resulted in significantly reduced glycolytic activity in vitro, with lower expression of lactate dehydrogenase 5, hypoxia-inducible factor 1 alpha subunit, and vascular endothelial growth factor, and was accompanied by reduced microvessel density in vivo. These results provide further support for PHIP as a molecular prognostic marker of melanoma, and reveal a significant linkage between PHIP levels and ulceration. Moreover, they suggest that ulceration may be driven by increased glycolysis and angiogenesis.