PD-L1 expression by tumor cell lines: A predictive marker in melanoma.

Prognostic biomarkers for patients with melanoma after lymph node resection are of clinical relevance and could thus enable the identification of patients who therefore would most benefit from adjuvant treatment. The aim of this work was to determine, using an in vitro model, whether immune-related biomarkers, such as MHC-class I and II, melanoma-associated antigens, IDO1 and PD-L1, could also be relevant to predict the risk of relapse of patients with stage III melanoma after lymph node resection. We established tumor cell lines from metastatic lymph nodes of 50 patients with melanoma. The expression of investigated biomarkers was determined on untreated and IFN-γ treated melanoma cell lines using flow cytometry. Among the selected biomarkers, the IFN-γ-induced expression of PD-L1 and IDO1 was associated with an increased risk of relapse (P = .0001 and P = .013, respectively) and was also associated with death for IDO1 (P = .0005). In the future, this immunologic signature could permit the identification of patients at higher risk of relapse and justifying an adjuvant treatment using immunotherapy.

Clinical significance of BRAF mutation status in circulating tumor DNA of metastatic melanoma patients at baseline.

Circulating tumor DNA is a promising non-invasive tool for cancer monitoring. The main objective of our work was to investigate the relationship between mutant BRAF DNA in plasma and clinical response. Thirty-eight stage IV patients with a V600 mutated BRAF melanoma were included prior to any treatment. DNA was extracted from plasma and mutant DNA was detected using the amplification-refractory mutation system method. Before the beginning of any treatment, the corresponding BRAF mutation was detected in 29 of the 38 tested plasma samples (76.3% positive per cent agreement). We observed a strong correlation between the presence of circulating mutated DNA and overall survival (OS; P=.02), and with the number of metastatic sites (P=.01). The presence of circulating mutated DNA was also strongly correlated with serum LDH activity (P<.01) and S100 protein concentration (P<.01). Finally, seven patients presented discordant BRAF status in different tumor sites. In all these patients, the test performed on ctDNA was positive, suggesting that ctDNA analysis might be less sensitive to tumor heterogeneity. Altogether, these results suggest that plasmatic mutant BRAF DNA is a prognostic factor of OS, correlated with tumor burden. In addition, it represents an interesting alternative source of DNA to detect BRAF mutations before treatment.

Prognostic value of tumor-infiltrating Foxp3+ T-cell subpopulations in metastatic melanoma.

Regulatory T cells have already been associated with poor prognosis in various types of cancer. It was previously reported, in ovarian carcinoma, that quantification of Foxp3 identified a subgroup of patients characterized by a significantly worse prognosis in terms of overall survival (OS) and progression-free survival (PFS), suggesting that high expression levels of Foxp3 might represent a surrogate marker for an immunosuppressive microenvironment contributing to tumor immune escape. The main objective of the present study was to precise the prognostic value of Foxp3 regarding PFS and OS in stage III (AJCC) melanoma patients. Total RNA was isolated from 102 metastatic melanoma lymph nodes and from eight tumor-free lymph nodes. Real-time PCR for Foxp3 was performed and correlated with patients' outcome. Quantification of Foxp3 identified a patient subgroup (>90th percentile), which is characterized by a significantly worse prognosis in terms of PFS (P = 0.000271) but not in terms of OS (P = 0.11). In conclusion, quantification of Foxp3 expression using qPCR appears as an independent prognostic factor for PFS in stage III melanoma patients (AJCC). High Foxp3 expression might thus enable the identification of patients most at risk of relapse.

Propionibacterium acnes activates the IGF-1/IGF-1R system in the epidermis and induces keratinocyte proliferation.

Propionibacterium acnes has a major role in the development of acne lesions. IGF-1 stimulates the proliferation of keratinocytes via an activation of the IGF-1 receptor (IGF-1R). Zinc has been proven to work efficiently against inflammatory acne and to modulate the IGF-1 system. Our objectives were to study the modulation of IGF-1 and IGF-1R expression by P. acnes extracts and to determine their modulation by zinc gluconate. In vivo, we analyzed biopsies of acne lesions and healthy skin, and in vitro we used skin explants incubated with two P. acnes extracts--membrane fraction (MF) and cytosolic proteins--with or without zinc. IGF-1 and IGF-1R expression was evaluated using immunohistochemistry, and the IGF-1 production in supernatants was measured by ELISA. Then, IGF-1 and IGF-1R mRNA levels were analyzed using quantitative PCR on normal human epidermal keratinocytes (NHEKs). IGF-1 and IGF-1R were overexpressed in acne lesions. MF increased IGF-1 and IGF-1R expression in the epidermis of explants and was associated with an overexpression of both Ki-67 and filaggrin. Zinc had the effect of downregulating IGF-1 and IGF-1R levels. These observations were confirmed at the mRNA level for IGF-1R in NHEKs. These results demonstrate that P. acnes can induce the formation of comedones by stimulating the IGF/IGF-1R system. Moreover, zinc downregulates this pathway.

Treatment of metastatic melanoma with autologous Melan-A/MART-1-specific cytotoxic T lymphocyte clones.

Immunotherapy by adoptive T-cell transfer aims at maximizing tumor antigen-specific T-cell responses. We treated 14 patients at the metastatic stage in a phase II study with Melan-A-specific T-cell clones generated from patient blood. During the period required for T-cell clone generation, the patients were treated by dacarbazine. Every patient received a T-cell clone suspension followed by subcutaneous injections of interleukin 2 and interferon alpha. Patients were monitored until disease progression occurred. We succeeded in obtaining autologous Melan-A-specific cytotoxic T lymphocyte clones, which were highly reactive against tumor cells for all the patients. Of the 14 patients treated, six (43%) experienced an objective response (CR + PR) with long-term complete remission for two patients (1 CR for 5 years and 1 CR for 28 months). Furthermore, we showed that all the clinical responses were significantly associated with in vivo expansion of the Melan-A-specific T-cell repertoire. This phenomenon appeared to be significantly associated with clinical responses. Thus, over the course of an adoptive cell transfer, monitoring this melanoma-specific T-cell expansion in patient blood appears crucial for predicting the clinical efficiency of such an immunological approach.