Progression in cutaneous malignant melanoma is associated with distinct expression profiles: a tissue microarray-based study

Abstract

Cutaneous malignant melanoma remains the leading cause of skin cancer death in industrialized countries. Clinical and histological variables that predict survival, such as Breslow's index, tumor size, ulceration, or vascular invasion have been identified in malignant melanoma. Nevertheless, the potential relevance of biological variables still awaits an in-depth exploration. Using tissue microarrays (TMAs), we retrospectively analyzed 165 malignant melanoma samples from 88 patients corresponding to distinct histological progression phases, radial, vertical, and metastases. A panel of 39 different antibodies for cell cycle, apoptosis, melanoma antigens, transcription factors, DNA mismatch repair, and other proteins was used. Integrating the information, the study has identified expression profiles distinguishing specific melanoma progression stages. Most of the detected alterations were linked to the control of cell cycle G1/S transition; cyclin D1 was expressed in radial cases 48% (12 of 25) with significant lost of expression in vertical cases 14% (9 of 65), P = 0.002; whereas p16(INK4a) (89% in vertical versus 71% in metastatic cases, P = 0.009) and p27(KIP1) (76% in radial versus 45% in vertical cases, P = 0.010) were diminished in advanced stages. The study also defines a combination of biological markers associated with shorter overall survival in patients with vertical growth phase melanoma, that provided a predictor model with four antibodies (Ki67, p16(INK4a), p21(CIP1), and Bcl-6). This predictor model was validated using an independent series of 72 vertical growth phase melanoma patients.

Figure 1

TMA design for melanoma progression. Cylinders corresponding to nevus (A), radial growth phase (B), vertical growth phase (C), and metastases (D). Immunohistochemical patterns. Core samples with positive and negative (or high and low) expression for selected antibodies are shown. All of the cases are representative of nevus-column A, and of each malignant melanoma progression step-columns B, C, and D. Note the high expression of cyclin D1 in radial (B) and metastases (D) when compared with vertical cases (C) and the loss of p16^INK4a expression in metastases. Note also the loss of p27^KIP1 and caveolin with tumor progression and the nuclear survivin expression in melanomas when compared with nevi. Original magnification for A–D, ×40; insets, ×600 and ×800, respectively. HE, hematoxylin and eosin.

Figure 2

Signature in melanoma progression. Representative proteins differentially expressed in cutaneous malignant melanoma identified by tissue array analysis. Subheadings indicate protein function and arrows represent up- or down-regulation in radial growth phase melanoma relative to nevus; vertical growth phase relative to radial growth phase and metastasis versus vertical growth phase. If there was no difference in protein expression between one phase and the comparative phase, the name of the marker is not included. See also Table 2 for differences in protein expression. A total of 175 specimens were evaluated, including 10 nevus, 28 radial malignant melanoma, 66 vertical malignant melanoma, and 71 metastases (34 skin metastases and 37 nodal metastases).

Figure 3

Kaplan-Meier estimation of overall survival according to the assigned probability in the multivariate analysis. A score was built-up according with the multivariate model. The survival curves for the original and the validation groups are presented. Patients are grouped collapsing the score values into three categories (low, intermediate, and high risk patients) obtained by dividing the whole hazard ratio range into three equally spaced categories (groups).