A multi-factorial genetic model for prognostic assessment of high risk melanoma patients receiving adjuvant interferon

Purpose: IFNa was the first cytokine to demonstrate anti-tumor activity in advanced melanoma. Despite the ability of high-dose IFNa reducing relapse and mortality by up to 33%, large majority of patients experience side effects and toxicity which outweigh the benefits. The current study attempts to identify genetic markers likely to be associated with benefit from IFN-a2b treatment and predictive for survival. Experimental design: We tested the association of variants in FOXP3 microsatellites, CTLA4 SNPs and HLA genotype in 284 melanoma patients and their association with prognosis and survival of melanoma patients who received IFNa adjuvant therapy. Results: Univariate survival analysis suggested that patients bearing either the DRB1*15 or HLA-Cw7 allele suffered worse OS while patients bearing either HLA-Cw6 or HLA-B44 enjoyed better OS. DRB1*15 positive patients suffered also worse RFS and conversely HLA-Cw6 positive patients had better RFS. Multivariate analysis revealed that a five-marker genotyping signature was prognostic of OS independent of disease stage. In the multivariate Cox regression model, HLA-B38 (p = 0.021), HLA-C15 (p = 0.025), HLA-C3 (p = 0.014), DRB1*15 (p = 0.005) and CT60*G/G (0.081) were significantly associated with OS with risk ratio of 0.097 (95% CI, 0.013-0.709), 0.387 (95% CI, 0.169-0.889), 0.449 (95% CI, 0.237-0.851), 1.948 (95% CI, 1.221-3.109) and 1.484 (95% IC, 0.953-2.312) respectively. Conclusion: These results suggest that gene polymorphisms relevant to a biological occurrence are more likely to be informative when studied in concert to address potential redundant or conflicting functions that may limit each gene individual contribution. The five markers identified here exemplify this concept though prospective validation in independent cohorts is needed

Evaluation of six CTLA-4 polymorphisms in high-risk melanoma patients receiving adjuvant interferon therapy in the He13A/98 multicenter trial

<p>ABSTRACT</p> <p>Purpose</p> <p>Interferon is approved for adjuvant treatment of patients with stage IIb/III melanoma. The toxicity and uncertainty regarding survival benefits of interferon have qualified its acceptance, despite significant durable relapse prevention in a fraction of patients. Predictive biomarkers that would enable selection of patients for therapy would have a large impact upon clinical practice. Specific CTLA-4 polymorphisms have previously shown an association with response to CTLA-4 blockade in patients with metastatic melanoma and the development of autoimmunity.</p> <p>Experimental design</p> <p>286 melanoma patients and 288 healthy controls were genotyped for six CTLA-4 polymorphisms previously suggested to be important (AG 49, CT 318, CT 60, JO 27, JO30 and JO 31). Specific allele frequencies were compared between the healthy and patient populations, as well as presence or absence of these in relation to recurrence. Alleles related to autoimmune disease were also investigated.</p> <p>Results</p> <p>No significant differences were found between the distributions of CTLA-4 polymorphisms in the melanoma population compared with healthy controls. Relapse free survival (RFS) and overall survival (OS) did not differ significantly between patients with the alleles represented by these polymorphisms. No correlation between autoimmunity and specific alleles was shown. The six polymorphisms evaluated where strongly associated (Fisher's exact p-values < 0.001 for all associations) and significant linkage disequilibrium among these was indicated.</p> <p>Conclusion</p> <p>No polymorphisms of CTLA-4 defined by the SNPs studied were correlated with improved RFS, OS, or autoimmunity in this high-risk group of melanoma patients.</p

FUTURE PERSPECTIVES IN MELANOMA RESEARCH. Meeting report from the “Melanoma Research: a bridge from Naples to the World. Napoli, December 5th–6 th2011”

After more than 30 years, landmark progress has been made in the treatment of cancer, and melanoma in particular, with the success of new molecules such as ipilimumab, vemurafenib and active specific immunization

Regulatory T cell frequency in patients with melanoma with different disease stage and course, and modulating effects of high-dose interferon-α 2b treatment

<p>Abstract</p> <p>Background</p> <p>High-dose interferon-alpha 2b (IFN-α 2b) is the only approved systemic therapy in the United States for the adjuvant treatment of melanoma. The study objective was to explore the immunomodulatory mechanism of action for IFN-α 2b by measuring serum regulatory T cell (Treg), serum transforming growth factor-β (TGF-β), interleukin (IL)-10, and autoantibody levels in patients with melanoma treated with the induction phase of the high-dose IFN-α 2b regimen.</p> <p>Methods</p> <p>Patients with melanoma received IFN-α 2b administered intravenously (20 MU/m²each day from day 1 to day 5 for 4 consecutive weeks). Serum Treg levels were measured as whole lymphocytes in CD4⁺cells using flow cytometry while TGF-β, IL-10, and autoantibody levels were measured using enzyme-linked immunosorbent assays.</p> <p>Results</p> <p>Twenty-two patients with melanoma received IFN-α 2b treatment and were evaluated for Treg levels. Before treatment, Treg levels were significantly higher in patients with melanoma when compared with data from 20 healthy subjects (<it>P </it>= 0.001; Mann-Whitney test). Although a trend for reduction of Treg levels following IFN-α 2b treatment was observed (average decrease 0.29% per week), statistical significance was not achieved. Subgroup analyses indicated higher baseline Treg levels for stage III versus IV disease (<it>P </it>= 0.082), early recurrence versus no recurrence (<it>P </it>= 0.017), deceased versus surviving patients (<it>P = </it>0.021), and preoperative neoadjuvant versus postoperative adjuvant treatment groups (not significant). No significant effects were observed on the levels of TGF-β, IL-10, and autoantibodies in patients with melanoma treated with IFN-α 2b.</p> <p>Conclusions</p> <p>Patients with melanoma in this study showed increased basal levels of Treg that may be relevant to their disease and its progression. Treg levels shifted in patients with melanoma treated with IFN-α 2b, although no firm conclusions regarding the role of Tregs as a marker of treatment response or outcome can be made at present.</p

Modulation of SOCS protein expression influences the interferon responsiveness of human melanoma cells

<p>Abstract</p> <p>Background</p> <p>Endogenously produced interferons can regulate the growth of melanoma cells and are administered exogenously as therapeutic agents to patients with advanced cancer. We investigated the role of negative regulators of interferon signaling known as suppressors of cytokine signaling (SOCS) in mediating interferon-resistance in human melanoma cells.</p> <p>Methods</p> <p>Basal and interferon-alpha (IFN-α) or interferon-gamma (IFN-γ)-induced expression of SOCS1 and SOCS3 proteins was evaluated by immunoblot analysis in a panel of n = 10 metastatic human melanoma cell lines, in human embryonic melanocytes (HEM), and radial or vertical growth phase melanoma cells. Over-expression of SOCS1 and SOCS3 proteins in melanoma cells was achieved using the PINCO retroviral vector, while siRNA were used to inhibit SOCS1 and SOCS3 expression. Tyr⁷⁰¹-phosphorylated STAT1 (P-STAT1) was measured by intracellular flow cytometry and IFN-stimulated gene expression was measured by Real Time PCR.</p> <p>Results</p> <p>SOCS1 and SOCS3 proteins were expressed at basal levels in melanocytes and in all melanoma cell lines examined. Expression of the SOCS1 and SOCS3 proteins was also enhanced following stimulation of a subset of cell lines with IFN-α or IFN-γ. Over-expression of SOCS proteins in melanoma cell lines led to significant inhibition of Tyr⁷⁰¹-phosphorylated STAT1 (P-STAT1) and gene expression following stimulation with IFN-α (IFIT2, OAS-1, ISG-15) or IFN-γ (IRF1). Conversely, siRNA inhibition of SOCS1 and SOCS3 expression in melanoma cells enhanced their responsiveness to interferon stimulation.</p> <p>Conclusions</p> <p>These data demonstrate that SOCS proteins are expressed in human melanoma cell lines and their modulation can influence the responsiveness of melanoma cells to IFN-α and IFN-γ.</p