Gang Li

Summary: The E3 ligase Rad18 is a key regulator for the lesion bypass pathway, which plays an important role in genomic stability. However, the status of Rad18 expression in melanoma is not known. Using melanoma tissue microarray (TMA), we showed that nuclear Rad18 expression was upregulated in primary and metastatic melanoma compared to dysplastic nevi. Rad18 expression was significantly reduced in sun‐exposed sites compared to the sun‐protected sites. Strong Rad18 expression correlated with worse 5‐year patient survival and was an independent prognostic factor for melanoma found in the sun‐protected sites. Furthermore, we showed that melanoma cell proliferation and the expression of pAkt and cyclin D1 were reduced upon Rad18 knockdown. We, for the first time, showed that Rad18 is significantly increased in melanoma and predicts the poor outcome for melanoma in the sun‐protected sites. Rad18 is involved in the regulation of melanoma cell proliferation, which can be exploited in designing new strategy for melanoma treatment.

The INhibitor of Growth (ING) family is an evolutionarily conserved set of proteins, implicated in suppression of initiation and progression of cancers in various tissues. They promote cell cycle arrest, cellular senescence and apoptosis, participate in stress responses, regulate DNA replication and DNA damage responses, and inhibit cancer cell migration, invasion, and angiogenesis of the tumors. At the molecular level, ING proteins are believed to participate in chromatin remodeling and transcriptional regulation of their target genes. However, the best known function of ING proteins is their cooperation with p53 tumor suppressor protein in tumor suppression. All major isoforms of ING family members can promote the transactivition of p53 and the majority of them are shown to directly interact with p53. In addition, ING proteins are thought to interact with and modulate the function of auxiliary members of p53 pathway, such as MDM2, ARF , p300, and p21, indicating their widespread involvement in the regulation and function of this prominent tumor suppressor pathway. It seems that p53 pathway is the main mechanism by which ING proteins exert their functions. Nevertheless, regulation of other pathways which are not relevant to p53, yet important for tumorigenesis such as TGF-β and NF-κB, by ING proteins is also observed. This review summarizes the current understanding of the mutual interactions and cooperation between different members of ING family with p53 pathway and implications of this cooperation in the suppression of cancer initiation and progression.

S-phase kinase protein 2 (Skp2), an F-box protein, targets cell cycle regulators via ubiquitin-mediated degradation. Skp2 is frequently overexpressed in a variety of cancers and associated with patient survival. In melanoma, however, the prognostic significance of subcellular Skp2 expression remains controversial. To investigate the role of Skp2 in melanoma development, we constructed tissue microarrays and examined Skp2 expression in melanocytic lesions at different stages, including 30 normal nevi, 61 dysplastic nevi, 290 primary melanomas and 146 metastatic melanomas. The TMA was assessed for cytoplasmic and nuclear Skp2 expression by immunohistochemistry. The Kaplan-Meier method was used to evaluate the patient survival. The univariate and multivariate Cox regression models were performed to estimate the harzard ratios (HR) at five-year follow-up. Cytoplasmic but not nuclear Skp2 expression was gradually increased from normal nevi, dysplastic nevi, primary melanomas to metastatic melanomas. Cytoplasmic Skp2 expression correlated with AJCC stages (I vs II–IV, P2.00 mm, P0.05). This study demonstrates that cytoplasmic Skp2 plays an important role in melanoma pathogenesis and its expression correlates with patient survival. Our data indicate that cytoplasmic Skp2 may serve as a potential biomarker for melanoma progression and a therapeutic target for this disease.

The lesion bypass pathway, which is regulated by monoubiquitination of proliferating cell nuclear antigen (PCNA), is essential for resolving replication stalling due to DNA lesions. This process is important for preventing genomic instability and cancer development. Previously, it was shown that cells deficient in tumour suppressor p33ING1 (ING1b) are hypersensitive to DNA damaging agents via unknown mechanism. In this study, we demonstrated a novel tumour suppressive function of ING1b in preserving genomic stability upon replication stress through regulating PCNA monoubiquitination. We found that ING1b knockdown cells are more sensitive to UV due to defects in recovering from UV-induced replication blockage, leading to enhanced genomic instability. We revealed that ING1b is required for the E3 ligase Rad18-mediated PCNA monoubiquitination in lesion bypass. Interestingly, ING1b-mediated PCNA monoubiquitination is associated with the regulation of histone H4 acetylation. Results indicate that chromatin remodelling contributes to the stabilization of stalled replication fork and to the regulation of PCNA monoubiquitination during lesion bypass.

BACKGROUND: RUNX3 is a tumor suppressor that plays important roles in cell proliferation, apoptosis, and metastasis. The authors investigated the role of RUNX3 in melanoma pathogenesis and analyzed the prognostic impact of RUNX3 expression in a large series of melanoma patients.METHODS: Two sets of tissue microarrays were constructed, including 440 cases of melanomas (202 for the training set and 238 for the validation set) and 88 cases of nevi (25 normal nevi and 63 dysplastic nevi). RUNX3 expression was evaluated by immunohistochemistry.RESULTS: Positive RUNX3 expression was observed in 56%, 54%, 33%, and 24% of the biopsies in normal nevi, dysplastic nevi, primary melanoma, and melanoma metastases, respectively. Significant differences for positive nuclear RUNX3 staining were observed between dysplastic nevi and primary melanomas (P = .002, chi‐square test), between dysplastic nevi and melanoma metastases (P < .001, chi‐square test), and between primary melanoma and melanoma metastases (P = .045, chi‐square test). Loss of RUNX3 expression was correlated with a worse 5‐year survival of melanoma patients in both training and validation sets. Furthermore, loss of RUNX3 expression was also correlated with a poor 5‐year disease‐specific survival in primary melanoma (P = .001) and metastatic melanoma patients (P = .008). Multivariate Cox regression analysis revealed that positive RUNX3 expression is an independent prognostic factor to predict melanoma patient outcome.CONCLUSIONS: Our findings indicate that RUNX3 plays an important role in melanoma pathogenesis and may serve as a promising prognostic marker for melanoma. Cancer 2011;. © 2010 American Cancer Society.

Cadherin-like protein 22 (CDH22) is a transmembrane glycoprotein implicated in cell-cell adhesion and cancer metastasis. The expression of CDH22 has been shown to be increased in colorectal cancers. However, the role of CDH22 in melanomagenesis is not known. To investigate the role of CDH22 in melanoma progression, we examined the expression of CDH22 in melanocytic lesions at different stages and analysed the correlation between CDH22 expression and clinicopathlogic parameters and patient survival. Using tissue microarray and immunohisto-chemistry, we evaluated CDH22 staining in 76 dysplastic nevi, 247 primary melanomas, and 143 metastatic melanomas. We found that metastatic melanomas had a significantly higher percentage of negative CDH22 staining than dysplastic nevi (P = 0.012) and primary melanomas (P = 0.038). CDH22 expression was also reduced in thick (≥2 mm) and ulcerative melanomas (P = 0.003 and 0.022, respectively). Melanomas of AJCC stage II, III, and IV had a higher percentage of negative CDH22 staining than AJCC stage I melanomas (P = 0.004, P < 0.0001, and P = 0.009, respectively). Melanomas with negative CDH22 expression had significantly poorer disease-specific 5-year survival than those with positive CDH22 staining. Additionally, CDH22 expression depended on AJCC stage to predict patient survival. These data indicate that reduced CDH22 expression is associated with melanoma metastasis and poor patient prognosis.

The application of biomarkers in melanoma prognosis has been well recognized. However the ability of a single biomarker to predict melanoma patient outcome is usually limited. We previously examined the expression of ten biomarkers (Bim, BRG1, BRMS1, CTHRC1, ING4, NQO1, NF-κB-p50, PUMA, SNF5 and SOX4) in melanomas. To assess the value of a combined multiple biomarker system in melanoma prognosis, we compared the expression of each biomarker between various stages of melanoma, and determined the best combination of biomarkers for melanoma prognosis. Although the expression of six biomarkers (Bim, BRMS1, ING4, NQO1, PUMA and SOX4) was significantly decreased in AJCC III–IV stages of melanoma compared to AJCC I–II stages, the combined 6-biomarker index score exhibited higher variations than any individual biomarker in the same comparison. Moreover, the 6-biomarker index score was correlated with melanoma thickness, location and subtype, and predicted the outcome of melanoma patients more accurately than the individual biomarkers. Multivariate Cox regression analysis demonstrated that the 6-biomarker index score is an independent prognostic factor for melanoma. In conclusion, our study suggests that a multi-biomarker system test is valuable for improved outcome prediction in melanoma patients and for the development of novel therapeutic strategies.

The INhibitor of Growth (ING) proteins belong to a well-conserved family which presents in diverse organisms with several structural and functional domains for each protein. The ING family members are found in association with many cellular processes. Thus, the ING family proteins are involved in regulation of gene transcription, DNA repair, tumorigenesis, apoptosis, cellular senescence and cell cycle arrest. The ING proteins have multiple domains that are potentially capable of binding to many partners. It is conceivable, therefore, that such proteins could function similarly within protein complexes. In this case, within this family, each function could be attributed to a specific domain. However, the role of ING domains is not definitively clear. In this review, we summarize recent advances in structure-function relationships in ING proteins. For each domain, we describe the known biological functions and the approaches utilized to identify the functions associated with ING proteins.