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Despite numerous efforts to define the best therapeutic strategies in advanced melanoma, the response of many patients remains heterogeneous and of short duration. Lenalidomide, an immunomodulating drug, has shown anti-inflammatory, antiangiogenic and anticancer properties in haematological disorders; however, few preclinical data support the rationale for using this drug in melanoma patients. In this study, we investigate lenalidomide's potential role in melanoma by focusing on the in-vitro drug's antiproliferative activity. The antiproliferative action of lenalidomide was evaluated on two melanoma cell lines by MTT assay, cell cycle and apoptosis assay. P21 protein levels were evaluated with droplet digital PCR (ddPCR) and western blot analysis while his interaction with specific cyclin-dependent kinase (CDK) was assessed by immunoprecipitation test. The biological effect and molecular mechanisms of programmed cell death-1 (PD-1) in the regulation of proliferation were evaluated using ddPCR, flow cytometry, western blot and small interfering RNA transfection. We observed that lenalidomide exerts a cytostatic effect in melanoma cell lines by inducing cell cycle arrest in the G0-G1 phase through p21 upregulation and modulation of CDK complexes. Furthermore, we found that lenalidomide has an antiproliferative action through the downregulation of melanoma-PD1 expression and consequently the alteration of intracellular signaling of mammalian target of rapamycin/S6. The present study aims to provide new insights into the role of lenalidomide in melanoma and suggesting to potentially translating these findings into a clinical setting to use immunomodulatory derivatives for blocking the pro-tumorigenic activity of the melanoma through the PD-1/PD-L1 axis., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Bone metastases occur rarely in patients suffering from malignant melanoma, although their onset severely worsens both prognosis and quality of life. Extracellular vesicles (EVs) including exosomes (Exos) are active players in melanoma progression involved in the formation of the pre-metastatic niche., Methods: Trans-well assays explored the basal migratory and invasive potential of four melanoma cell lines and investigated their different propensity to be attracted toward the bone. Exosomes were purified from cell supernatants by ultracentrifugation and explored in their ability to influence the bone tropism of melanoma cells. The molecular machinery activated during this process was investigated by RT-PCR, droplet digital-PCR, flow-cytometry and Western blot, while loss of function studies with dedicated siRNAs defined the single contribute of CXCR4 and CXCR7 molecules., Results: Melanoma cells revealed a variable propensity to be attracted toward bone fragments. Gene profiling of both osteotropic and not-osteotropic cells did not show a different expression of those genes notoriously correlated to chemotaxis and bone metastasis. However, bone conditioned medium significantly increased CXCR4, CXCR7 and PTHrP expression solely to osteotropic cells, while their Exos were able to revert the original poor bone tropism of not-osteotropic cells through CXCR7 up-regulation. Silencing experiments also demonstrated that membrane expression of CXCR7 is required by melanoma cells to promote their chemotaxis toward SDF-1 gradients., Conclusions: Our data correlated the osteotropism of melanoma cells to the activation of the SDF-1/CXCR4/CXCR7 axis following the exposition of tumor cells to bone-derived soluble factors. Also, we demonstrated in vitro that tumor-derived Exos can reprogram the innate osteotropism of melanoma cells by up-regulating membrane CXCR7. These results may have a potential translation to future identification of druggable targets for the treatment of skeletal metastases from malignant melanoma.

The mechanisms leading to immune escape of melanoma have been largely investigated in relation to its tumour immunogenicity and features of inflamed microenvironment that promote the immune suppression during the disease progression. These findings have recently led to advantages in terms of immunotherapy-based approaches as rationale for overcoming the immune escape. However, besides immune checkpoints, other mechanisms including the adenosine produced by ectonucleotidases CD39 and CD73 contribute to the melanoma progression due to the immunosuppression induced by the tumour milieu. On the other hand, CD73 has recently emerged as both promising therapeutic target and unfavourable prognostic biomarker. Here, we review the major mechanisms of immune escape activated by the CD39/CD73/adenosine pathway in melanoma and focus potential therapeutic strategies based on the control of CD39/CD73 downstream adenosine receptor signalling. These evidences provide the basis for translational strategies of immune combination, while CD73 would serve as potential prognostic biomarker in metastatic melanoma.

Melanoma is an immunogenic cancer that overcomes the control of the immune system through the production of tolerogenic cytokines and growth factors in the microenvironment. In melanoma, dendritic cells (DC) show severe alterations in maturation, cross-priming and antigenic presentation, while other accessory cells infiltrating the tumor milieu also suppress DCs through the activation of the STAT pathway by IL-10 and IL-6. Novel immunotherapy strategies blocking cytotoxic T-lymphocyte antigen (CTLA-4) are successful in advanced disease, while melanoma cells carrying the BRAF(V600E) mutation further reinforce the immune suppression by activating MAPKs. Here, we review the major mechanisms involved in the cross-talk between melanoma cells and the immune system as well as the issue of defects in DCs in relation to novel studies aimed at restoring their anti-tumor activity.

Melanoma is an immunogenic tumor whose relationship with immune cells resident in the microenvironment significantly influences cancer cell proliferation, progression, and metastasis. During melanomagenesis, both immune and melanoma cells undergo the immunoediting process that includes interconnected phases as elimination, equilibrium, and escape or immune evasion. In this context, dendritic cells (DCs) are active players that indirectly counteract the proliferation of melanoma cells. Moreover, DC maturation, migration, and cross-priming as well as their functional interplay with cytotoxic T-cells through ligands of immune checkpoint receptors result impaired. A number of signals propagated by highly proliferating melanoma cells and accessory cells as T-cells, natural killer cells (NKs), tumor-associated macrophages (TAMs), T-regulatory cells (T-regs), myeloid-derived suppressor cells (MDSCs), and endothelial cells participate to create an immunosuppressive milieu that results engulfed of tolerogenic factors and interleukins (IL) as IL-6 and IL-10. To underline the role of the immune infiltrate in blocking the melanoma progression, it has been described that the composition, density, and distribution of cytotoxic T-cells in the surrounding stroma is predictive of responsiveness to immunotherapy. Here, we review the major mechanisms implicated in melanoma progression, focusing on the role of DCs. [ABSTRACT FROM AUTHOR]

Treatment with immune checkpoint inhibitors (ICIs) has improved the prognosis of patients with a number of types of cancer, but the frequent development of immune-related adverse effects (irAEs) can worsen the outcome. The most common irAEs involve the gastrointestinal, cutaneous, and endocrine systems, but nephrotoxicity, resulting from damage to the tubule-interstitial compartment, may occur in some patients. The early phases of acute interstitial nephritis (AIN) are characterized by systemic symptoms that indicate a poor clinical state as well as a mild deterioration of renal function. Tubular injury is due to a direct effect mediated by cytotoxic CD8+ T cells, which sustain the local production of pro-inflammatory cytokines that progressively impair renal function. The treatment of AIN is mainly based on high-dose steroids, which in most instances leads to the recovery of renal function. However, the premature discontinuation of ICI therapy may prevent the impact of treatment on the clinical progression of the malignancy. Adequately addressing irAEs requires a standardized therapy that is based on the results of large clinical trials. [ABSTRACT FROM AUTHOR]

The mechanisms leading to immune escape of melanoma have been largely investigated in relation to its tumour immunogenicity and features of inflamed microenvironment that promote the immune suppression during the disease progression. These findings have recently led to advantages in terms of immunotherapy-based approaches as rationale for overcoming the immune escape. However, besides immune checkpoints, other mechanisms including the adenosine produced by ectonucleotidases CD39 and CD73 contribute to the melanoma progression due to the immunosuppression induced by the tumour milieu. On the other hand, CD73 has recently emerged as both promising therapeutic target and unfavourable prognostic biomarker. Here, we review the major mechanisms of immune escape activated by the CD39/CD73/adenosine pathway in melanoma and focus potential therapeutic strategies based on the control of CD39/CD73 downstream adenosine receptor signalling. These evidences provide the basis for translational strategies of immune combination, while CD73 would serve as potential prognostic biomarker in metastatic melanoma. [ABSTRACT FROM AUTHOR]

Immunotherapy is effective in metastatic melanoma (MM) but most studies failed in discovering a biomarker predictive of clinical response. Exosomes (Exo) from melanoma cells are detectable in sera of MM patients similarly to those produced by immune cells that control the tumor progression. Here, we investigated by flow-cytometry the levels of Exo from both T-cells and dendritic cells (DCs) in 59 patients with MM treated with IPI and the relative expression of PD-1, CD28 and ICOS as well as CD80 and CD86. We found a significant increment of PD-1 and CD28 expression in patients achieving a clinical response reflected by improvement of both PFS and OS. Furthermore, MM patients receiving IPI who showed extended PFS underwent increased expression of CD80 and CD86 on DC-derived Exo at the end of treatment. These results suggest a possible association of both PD-1 and CD28 up-regulation on immune cell-derived Exo in patients with better clinical response to IPI. [ABSTRACT FROM AUTHOR]

Background Melanoma is an immunogenic cancer that overcomes the immune surveillance through the production of tolerogenic cytokines and growth factors within microenvironment. Melanoma-derived dendritic cells (DCs) show altered maturation, cross-priming and antigenic presentation and their major defect concerns the activation of the STAT pathway. The prognostic criteria to define melanoma at high-risk of relapse/recurrence include the Breslow depth, the Clark level and number of mitosis. Sentinel lymph node (SLN) characterization is a prognostic factor in melanoma, although false negative occurs in approximately 5% of patients. We investigated the potential prognostic role of DC number variation in relation to clinical stage, and suggest their role as early predictor of high risk melanomas. Methods 84 patients (group A; stage I-IV) with a previously definite diagnosis of melanoma as well as 18 (group B) with highly suspected cutaneous lesion were enrolled into the study. Peripheral blood was collected at study entry in patients of group A, whereas cells from group B were collected before and after the primary tumor exeresis and then before/after any surgical procedure including SLN, lymphadenectomy or metastasis removal. Both myeloid (m) and plasmacytoid (p) DCs were investigated by flow-cytometry using the anti- Lin, -CD11c, -BDCA-1, -CD123, and -BDCA-2 MoAbs. The percentage number of both mDCs and pDCs was correlated to clinical stage as well as to indipendent prognostic factors as histological features including Breslow, Clark level, presence/absence of tumor-infiltrating lymphocytes, and BRAF V600E/K mutations. Results The percentage of both mDCs and pDCs from group A were similar in stage I-III and dramatically reduced in those at stage IV. A. Lack of correlation was also demonstrated with clinical features and prognostic factors. By contrast, data from group B showed: an increase of mDCs and pDCs (p<0.05) after tumor removal as well as in those with negative SLN whereas both subsets resulted unchanged in patients with positive SLN. A weak trend to DC increase occurred in patients undergoing negative lymphoadenectomy. Number of mDCs and pDCs from group B was correlated with histological features and unrelated to mutational status. Conclusions These findings suggest the critical pathogenetic role of DC in melanoma and their measurement may be thus proposed as additional prognostic factor to limit the risk of underscoring the melanoma stage of patients at high risk of relapse/recurrence. [ABSTRACT FROM AUTHOR]