• ezh2
• histopathological
• immunotherapy
• melanoma
• nf1
• prognosis
• target

In patients affected by melanoma prognosis and treatment decisions rely on a combination of clinical, laboratory, and histopathological factors. Despite several investigations, no single predictive factor can reliably determine a patient’s response to oncological treatments, except for the BRAF mutation for target therapies.
Key prognostic factors include disease burden, metastatic site, and LDH levels. Specific clinical factors, such as a patient’s performance status (PS), BMI, concomitant medications, and the location of metastases, also affect survival. Molecular markers like PD-L1 expression, tumor mutational burden (TMB), and circulating tumor DNA (ctDNA) have emerged as predictive biomarkers of response to treatment, especially for immunotherapy.
The treatment of melanoma has been revolutionized by biological therapies, including immune checkpoint inhibitors (such as anti-PD-1 and anti-CTLA-4 antibodies) and target therapies (such as BRAF and MEK inhibitors). Although sequence studies have been conducted to determine the right order in which to administer the available treatments, the correct choice between immunotherapy and targeted therapies in first-line treatment remains a subject of investigation, in the absence of predictive biomarkers that alone influence the choice.
This retrospective study aims to evaluate the clinical outcomes of metastatic melanoma patients treated at the Oncology Department of the University Hospital of Pisa. The study focuses on identifying clinical, biohumoral, and histopathological factors that may influence response to treatment, disease progression and survival.
The population included patients with metastatic melanoma who received first-line immunotherapy or targeted therapy at the University Hospital of Pisa. To be included, patients needed to have available pre-treatment histological samples stored at the hospital's Pathological Anatomy departments. The study included both patients with metastatic melanoma at diagnosis and those who experienced relapse during follow-up or after adjuvant therapy.
We collected clinical and anamnestic data that characterized individuals and their diseases. Immunohistochemical analysis was conducted on the available histological samples at the Pathological Anatomy departments of the University Hospital of Pisa. The study specifically focused on: cytoplasmatic NF1 expression and nuclear EZH2 Expression. Nuclear expression of the histone methyltransferase EZH2 was analyzed using an anti-KMT6/EZH2 antibody. Double immunostaining was performed using an automated immunostainer (Ventana Discovery Ultra System) to detect these proteins in the melanoma tissues.
Categorical data were described with absolute frequency and percentage, continuous data with mean and standard deviation. Two survival endpoints were analyzed: OS and EFS. Analysis of explanatory factors for OS and EFS was performed using univariate Cox regression, followed by multivariable analysis. Significance was set at 0.05 and all analyzes were performed with SPSS version 29 technology.
The results obtained highlighted that the analysis of NF1 and EZH2 identified populations with different survival outcomes. In particular, low expression of NF1 seemed to correlate with a better outcome in patients undergoing immunotherapy, on the contrary, high expression of EZH2 seemed to identify patients with a better outcome among those who received target therapy. Although the statistical correlation was significant only in the univariate analysis for NF1 (for both Overall Survival and Event Free Survival) in patients treated with immunotherapy and was close to significance for EZH2 (only for Overall Survival) in patients undergoing at target therapy, the significance of these correlations were not confirmed in the multivariate analysis. Nonetheless, we believe that this may be due to the small sample size and that the differences in outcomes can still be considered clinically relevant. We believe it may be useful to investigate the expression of NF1 and EZH2 in larger population samples to establish the true extent of the correlation of NF1 and EZH2 expression with survival outcomes in patients with metastatic melanoma.