3-nitrotyrosine and Melanoma

Topics: Aldehydes; Animals; Carcinogenesis; DNA Damage; Female; Humans; Melanoma; Metformin; Mice; Oxidative Stress; Radiation Injuries, Experimental; Radiodermatitis; Skin; Skin Neoplasms; Tumor Suppressor Protein p53; Tyrosine; Ultraviolet Rays

Increased expression and prognostic significance of major redox regulator nuclear factor erythroid-2-related factor (Nrf2) is recognized in many cancers. Our aim was to investigate the role of oxidative stress markers in melanoma.. We characterized the immunohistochemical expression of Nrf2, kelch-like ECH-associated protein 1 (Keap1), BRAF(V600E), 8-hydroxy-2'-deoxyguanosine (8-OHdG) and nitrotyrosine in 36 nevi, 14 lentigo maligna and 71 malignant melanomas. We measured Nrf2 expression in melanoma cell lines and conducted cytotoxicity assays combining BRAF/NRAS ablation and H2O2treatment.. Nuclear Nrf2 expression in melanoma correlated with deeper Breslow (p<0.0005), invasive phenotype (Clark III-V) (p=0.011), nodular growth (p=0.001) and worse melanoma-specific survival (p=0.008). Absence of 8-OHdG in the endothelium was a greater significant predictor of poor prognosis (p=0.024) than ulceration (p=0.17) and had a similar impact on prognosis as Breslow (p=0.024). A decrease of Nrf2 followed the BRAF/NRAS inhibition, but combination of inhibitor with H2O2did not increase cytotoxicity.. Nrf2 and 8-OHdG influence prognosis in melanoma.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Aged; Cell Line, Tumor; Deoxyguanosine; Female; Humans; Intracellular Signaling Peptides and Proteins; Kaplan-Meier Estimate; Kelch-Like ECH-Associated Protein 1; Male; Melanoma; NF-E2-Related Factor 2; Oxidative Stress; Proto-Oncogene Proteins B-raf; Signal Transduction; Skin Neoplasms; Tyrosine

This study highlights the systemic oxidative changes in patients submitted to primary cutaneous melanoma removal. Cutaneous melanoma is highly aggressive and its incidence is increasing worldwide. We evaluated systemic oxidative stress (OS) and 3-nitrotyrosine (3-NT) expression in melanoma tissue in relation to the Breslow thickness in patients under surveillance. Forty-three patients with cutaneous melanoma and 50 healthy volunteers were recruited. Patients were divided into two groups according to the tumor's Breslow thickness: T1/T2 (<2 mm) and T3/T4 (≥2 mm). Systemic OS and inflammatory mediators were evaluated in plasma, and the 3-NT expression was analyzed via immunohistochemistry. Compared with the controls, the patients had lower blood levels of reduced glutathione, higher malondialdehyde and thiol levels, and a higher total radical-trapping antioxidant parameter to uric acid ratio. The C-reactive protein and γ-glutamyl transpeptidase were increased only in the T3/T4 group. High levels of 3-NT were present only in T3/T4 patients. Our data suggested that a correlation exists between the Breslow thickness and a systemic pro-oxidant status, and that oxidative changes induced by the melanoma remain in the microenvironment post-surgery, demonstrating a role for oxygen species in melanoma.

Topics: Adult; Aged; Female; Humans; Male; Melanoma; Melanoma, Cutaneous Malignant; Middle Aged; Oxidative Stress; Skin Neoplasms; Tumor Microenvironment; Tyrosine; Young Adult

Human melanoma tumors cells are known to express the enzyme, inducible nitric oxide synthase (iNOS), which is responsible for cytokine induced nitric oxide (NO) production during immune responses. This constitutive expression of iNOS in many patients' tumor cells, as well as its strong association with poor patient survival, have led to the consideration of iNOS as a molecular marker of poor prognosis, as well as a possible target for therapy. The expression of iNOS in patient tumors was found to associate with nitrotyrosine, COX2, pSTAT3, and arginase. Using human melanoma patients' samples as well as cell lines, we have further evidence supporting intracellular NO production by detection of nitrotyrosine and also by use of DAF-2DA staining. Experiments were performed to scavenge the endogenous NO (with c-PTIO) resulting in melanoma cell growth inhibition; this was restored with SIN-1 (NO and O2-donor) providing data to support a functional role of this gas. Our goal is to understand the aberrant biology leading to this curious phenomenon, and to regulate it in favor of patient treatments.

Topics: Apoptosis; Cell Line, Tumor; Cell Proliferation; Cyclooxygenase 2; Humans; Melanoma; Neoplasm Proteins; Nitric Oxide; Nitric Oxide Synthase Type II; STAT3 Transcription Factor; Tumor Cells, Cultured; Tyrosine

Nitric oxide (NO) is a small molecule produced during the conversion of L-arginine to L-citrulline by NO synthase (NOS). Several isoforms of NOS exist, of which the Ca2+-independent, inducible NOS (iNOS or NOS2) is most prominently expressed by macrophages. iNOS activity and increased levels of iNOS have been found in various tumours and tumour cell lines but not in normal tissues; however, the precise role of NO in tumour progression has yet to be elucidated. We studied the expression of iNOS in paraffin sections of 41 benign naevi and 52 primary malignant melanomas (MM) of the skin, as well as in 13 metastatic MM. In addition, nitrotyrosine, indicative of NO production and formation of peroxynitrite, was studied in frozen sections of 13 naevi and 30 MM. Virtually all naevi expressed iNOS, but very few expressed nitrotyrosine, indicating either that iNOS in naevi is functionally inactive, or that naevus cells lack reactive oxygen radicals and thus do not form peroxynitrite. Normal melanocytes in adjacent uninvolved skin were unreactive for both markers. In MM, iNOS was most frequently expressed in the 'pure' and 'invasive' radial growth phase (RGP), whereas expression in the vertical growth phase (VGP) and metastatic phase occurred only in 76% of cases; moreover, in these latest phases of tumour progression, iNOS staining was weak and focal. We conclude that iNOS is expressed de novo in most benign pigment cell lesions. In MM (iNOS-generated) NO appears to play an important part in the early steps of invasion (i.e. the 'invasive' RGP), where it may stimulate neo-angiogenesis and may be a prerequisite for further tumour progression; this view is also supported by the finding of iNOS in the associated blood vessels in the papillary dermis. Finally, our data suggest that (iNOS-generated) NO plays a less significant part in the VGP and in metastatic melanoma.

Topics: Biomarkers, Tumor; Disease Progression; Enzyme Induction; Humans; Isoenzymes; Macrophages; Melanoma; Neoplasm Metastasis; Nevus; Nitric Oxide Synthase; Skin Neoplasms; Tyrosine

Despite recognition of the malignant potential of human melanomas, the mechanisms responsible for the pathobiological characteristics contributing to tumor growth, vascular invasiveness, and distant organ metastasis remain undefined. Recent studies have shown that various human tumors express an inducible form of nitric oxide synthase (iNOS) and nitrotyrosine (NT), which suggests a mechanistic role of tumor-associated nitric oxide (NO) in tumorigenesis. We investigated iNOS and NT expression by immunohistochemistry in 20 human metastatic melanoma tissue specimens specifically with respect to iNOS-expressing cell types in the tumor area, pathological and clinical response to systemic therapy, potential role as a prognostic indicator, and NT formation. Our results showed that melanoma cells from 12 of 20 tumors express iNOS, yet the expression of this molecule in the tumor did not correlate with pathological or clinical response to therapy. More importantly, iNOS and NT expression by the melanoma cells strongly correlated with poor survival in patients with stage 3 disease (P < 0.001 and P = 0.020, respectively), suggesting a pathway whereby iNOS might contribute to enhanced tumor progression. In conclusion, our findings strongly suggest that iNOS expression has potential to be considered as a prognostic factor and NO as a critical mediator of an aggressive tumor phenotype in human metastatic melanomas.

Topics: Adolescent; Adult; Female; Humans; Immunohistochemistry; Male; Melanoma; Middle Aged; Neoplasm Metastasis; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Prognosis; Time Factors; Treatment Outcome; Tyrosine