Guohong Zhang

Targeting MAPK pathway in melanoma therapy

New drugs targeting the mitogen-activated protein kinase (MAPK) pathway have generated striking clinical response in melanoma therapy. From the discovery of BRAF mutation in melanoma in 2002, to the approval of first BRAF inhibitor vemurafenib for melanoma treatment by the US Food and Drug Administration in 2011, therapies targeting the MAPK pathway have been proven effective in less than a decade. The success of vemurafenib stimulated more intensive investigation of the molecular mechanisms of melanoma pathogenesis and development of new treatment strategies targeting specific molecules in MAPK pathway. Although selective BRAF inhibitors and MEK inhibitors demonstrated improved overall survival of metastatic melanoma patients, limited duration or development of resistance to BRAF inhibitors have been reported. Patients with metastatic melanoma still face very poor prognosis and lack of clarified therapies. Studies and multiple clinical trials on more potent and selective small molecule inhibitory compounds to further improve the clinical effects and overcome drug resistance are underway. In this review, we analyzed the therapeutic potentials of each member of the MAPK signaling pathway, summarized important MAPK-inhibiting drugs, and discussed the promising combination treatment targeting multiple targets in melanoma therapy, which may overcome the drawbacks of current drugs treatment.

Role of EIF5A2, a downstream target of Akt, in promoting melanoma cell invasion

Background:Cutaneous melanoma is a life-threatening skin cancer because of its poorly understood invasive nature and high metastatic potential. This study examines the importance of eukaryotic translation initiation factor 5A2 (EIF5A2) in melanoma pathogenesis.Methods:We examined EIF5A2 expression in 459 melanocytic lesions using tissue microarray. In addition, melanoma cell lines were subjected to invasion and cell proliferation assays, zymography, FACS and real-time PCR to investigate the role of EIF5A2 in cancer progression.Results:Positive EIF5A2 staining increased from dysplastic naevi to primary melanomas (PMs; P=0.001), and further increased in metastatic melanomas (P=0.044). Eukaryotic translation initiation factor 5A2 expression was correlated with melanoma thickness (P<0.001) and was inversely correlated with the 5-year survival of PM patients especially those with tumour⩽2 mm thick. Strikingly, none of the latter died within 5 years in EIF5A2-negative staining group. Cox regression analysis revealed that EIF5A2 is an independent prognostic marker. Further, we found that EIF5A2 is a novel downstream target of phosphorylated Akt. Both melanoma cell invasion and MMP-2 activity increased and decreased with EIF5A2 overexpression and knockdown, respectively.Conclusion:We for the first time showed that EIF5A2, as a target of PI3K/Akt, promotes melanoma cell invasion and may serve as a promising prognostic marker and a potential therapeutic target for melanoma.

A combination of p300 and Braf expression in the diagnosis and prognosis of melanoma

BackgroundTo date only a handful of drugs are available for the treatment of melanoma. Among them vemurafenib, a BrafV600E specific inhibitor, showed promising results in terms of response rate and increase in median survival time. However, its effectiveness is limited by development of resistance and the search for additional drugs for melanoma treatment is ongoing. The present study was performed to analyze the correlation between Braf expression and the expression of p300, a known down stream target of the mitogen activated protein kinase (MAPK) pathway, which was recently shown by us to be a prognostic marker for melanoma progression and patient survival.MethodsThe expression of Braf and p300 expression were correlated and analyzed by Chi-square test. A total of 327 melanoma patient cases (193 primary melanoma and 134 metastatic melanoma) were used for the study. Classification & regression tree (CRT), Kaplan-Meier, and multivariate Cox regression analysis were used to elucidate the significance of the combination of Braf and p300 expression in the diagnosis and prognosis of melanoma.ResultsOur results demonstrate that Braf expression is inversely correlated with nuclear p300 and positively correlated with cytoplasmic p300 expression. Braf and cytoplasmic p300 were found to be associated with melanoma progression, tumor size and ulceration status. CRT analysis revealed that a combination of Braf and p300 expression (nuclear and cytoplasmic), could be used to distinguish between nevi and melanoma, and primary from metastatic melanoma lesions. The combination of Braf and nuclear p300 was significantly associated with patient survival and nuclear p300 was found to be an independent predictor of patient survival.ConclusionOur results indicate a cross-talk between Braf and p300 in melanoma and demonstrate the importance Braf and p300 expression in the diagnosis and prognosis of melanoma.

JWA inhibits melanoma angiogenesis by suppressing ILK signaling and is an independent prognostic biomarker for melanoma

Melanoma is the deadliest cutaneous malignancy because of its high incidence of metastasis. Melanoma growth and metastasis relies on sustained angiogenesis; therefore, inhibiting angiogenesis is a promising approach to treat metastatic melanoma. JWA is a novel microtubule-associated protein and our previous work revealed that JWA inhibited melanoma cell invasion and metastasis. However, the role of JWA in melanoma angiogenesis and the prognostic value are still unknown. Here, we report that JWA in melanoma cells significantly inhibited the tube formation of endothelial cells. In addition, JWA regulated integrin-linked kinase (ILK) through integrin αVβ3 and such regulation was achieved through the transcription factor Sp1. Notably, both in vitro and in vivo angiogenesis assays revealed that JWA dramatically suppressed melanoma angiogenesis by inhibiting ILK signaling. Furthermore, we examined the expression of JWA protein in a large set of melanocytic lesions (n = 505) at different stages by tissue microarray and found an inverse correlation between JWA expression and melanoma progression (P = 5 × 10(-6)). Importantly, reduced JWA expression was correlated with a poorer overall, and disease-specific 5 year survival of patients (P = 0.001 and 0.007, respectively). Multivariate Cox regression analyses indicated that JWA was an independent prognostic marker for melanoma patients. Moreover, we found a significant negative correlation between JWA and ILK in melanoma biopsies, and their concomitant expression was closely correlated with melanoma patient survival (P = 0.004), further indicating the regulation of ILK expression by JWA is critical in melanoma. Taken together, our data highlight the function of JWA in melanoma angiogenesis and reveal the clinical prognostic value of JWA.

Therapeutic efficacy of combined BRAF and MEK inhibition in metastatic melanoma: a comprehensive network meta-analysis of randomized controlled trials.

Background Several recent randomized clinical trials have preliminarily demonstrated that initial targeted therapy with combined BRAF and MEK inhibition is more effective in metastatic melanoma (MM) than single agent. To guide therapeutic decisions, we did a comprehensive network meta-analysis to identify evidence to robustly support whether combined BRAF and MEK inhibition is the best initial targeted therapeutic strategy for patients with MM. Methods The databases of PubMed and trial registries were researched for randomized clinical trials of targeted therapy. Data of outcome were extracted on progression-free survival (PFS), objective response rate (ORR), and overall survival (OS). Network meta-analysis using a Bayesian statistical model was performed to evaluate relative hazard ratio (HR) for PFS and OS, odds ratio (OR) for ORR. Results Finally, 16 eligible trials comprising 5976 participants were included in this meta-analysis. PFS were significantly prolonged in patients who received combined BRAF-MEK inhibition compared with those who received BRAF inhibition (HR: 0.58, 95%CI: 0.51-0.67, P < 0.0001) or MEK inhibition alone (HR: 0.29, 95%CI: 0.22-0.37, P < 0.0001). Combined BRAF-MEK inhibition also improved the OS over BRAF inhibition (HR: 0.67, 95%CI: 0.56-0.81, P < 0.0001) or MEK inhibition alone (HR: 0.48, 95%CI: 0.36-0.65, P < 0.0001). The ORR was superior in combined BRAF and MEK inhibition comparing with BRAF inhibition (OR: 2.00, 95%CI: 1.66-2.44, P < 0.0001) or MEK inhibition alone (OR: 20.66, 95%CI: 12.22-35.47, P < 0.0001). Conclusions This study indicates that concurrent inhibition of BRAF and MEK improved the most effective therapeutic modality as compared as single BRAF or MEK inhibition for patients with MM.

ING4 regulates JWA in angiogenesis and their prognostic value in melanoma patients.

Background:We previously showed that inhibitor of growth family member 4 (ING4) inhibits melanoma angiogenesis, and JWA suppresses the metastasis of melanoma cells. As angiogenesis is essential for tumour metastasis, further investigation of the function of ING4 and JWA in melanoma angiogenesis is needed, and their prognostic value are of great interest.Methods:Western blot, tube-formation assays and luciferase assays were used to investigate the correlation between ING4 and JWA in melanoma angiogenesis. JWA and integrin-linked kinase (ILK) expression was determined on a tissue microarray constructed from 175 biopsies.Results:ING4 promoted JWA expression by activating JWA promoter. Furthermore, the regulation of growth and tube formation of endothelial cells by ING4 was partially JWA dependent. Also, ING4 inhibited the ILK-induced angiogenesis signalling pathway via JWA. Moreover, reduced JWA, or increased ILK, expression was closely associated with 5-year disease-specific survival of melanoma patients (P=0.001 and 0.007, respectively). There was also a positive correlation between ING4 and JWA yet a negative correlation between ING4 and ILK. Importantly, their concomitant expressions were significantly related to 5-year survival of melanoma patients (P=0.002 and 0.003, respectively).Conclusion:JWA has an important role in ING4-regulated melanoma angiogenesis, and ING4/JWA/ILK are promising prognostic markers and may be used as anti-angiogenic therapeutic targets for melanoma.

C-terminal tensin-like protein is a novel prognostic marker for primary melanoma patients.

Background C-terminal tensin-like protein (Cten) is a focal adhesion protein originally identified as a tumor suppressor in prostate cancer. It has since been found to be overexpressed and function as an oncogene in numerous other cancers, but the expression status of Cten in melanoma is still unknown. Methods Using tissue microarrays containing 562 melanocytic lesions, we evaluated Cten protein expression by immunohistochemistry. The association between Cten expression and patient survival was examined using Kaplan-Meier survival analysis, and univariate and multivariate Cox regression analyses were used to estimate the crude and adjusted hazard ratios. Results Strong Cten expression was detected in 7%, 24%, 41%, and 46% of normal nevi, dysplastic nevi, primary melanoma, and metastatic melanoma samples, respectively, and Cten expression was found to be significantly higher in dysplastic nevi compared to normal nevi (P = 0.046), and in primary melanoma compared to dysplastic nevi (P = 0.003), but no difference was observed between metastatic and primary melanoma. Cten staining also correlated with AJCC stages (P = 0.015) and primary tumor thickness (P = 0.002), with Cten expression being induced in the transition from thin (<1mm) to thick (≥1mm) melanomas. Strong Cten expression was significantly associated with a worse 5-year overall (P = 0.008) and disease-specific survival (P = 0.004) for primary melanoma patients, and multivariate Cox regression analysis revealed that Cten expression was an independent prognostic marker for these patients (P = 0.038 for overall survival; P = 0.021 for disease-specific survival). Conclusion Our findings indicate that induction of Cten protein expression is a relatively early event in melanoma progression, and that Cten has the potential to serve as a prognostic marker for primary melanoma patients.

Increased expression of neuropilin 1 in melanoma progression and its prognostic significance in patients with melanoma

Neuropilin 1 (NRP1), a receptor of vascular endothelial growth factor (VEGF), promotes angiogenesis, tumor growth, tumor invasion and metastasis. However, the function of NRP1 in melanoma progression, as well as the effect of NRP1 expression on the prognosis of patients with melanoma remains unknown. In the present study, NRP1 expression was examined in 460 cases of melanocytic lesions (28 common nevi, 51 dysplastic nevi, 250 primary melanoma and 131 metastatic melanoma) at different stages, using a tissue microarray. The correlation of NRP1 expression with melanoma progression, and its prognostic value in patients with melanoma was examined. In addition, the correlation between matrix metalloproteinase 2 (MMP2) and NRP1 expression in patients with melanoma was analyzed. The results demonstrated that NRP1 expression was significantly increased in primary (56%) and metastatic melanoma (62%), compared with common nevi (11%) and dysplastic nevi (24%). Notably, increased NRP1 expression was correlated with a poorer overall, and disease specific, 10-year survival (P=0.03 and P=0.002, respectively). Multivariate Cox regression analyses indicated that NRP1 is an independent prognostic marker for melanoma. Furthermore, a significant positive correlation between NRP1 and MMP2 expression in melanoma biopsies was observed, and their concomitant expression was closely correlated with melanoma patient survival, further supporting the hypothesis that the expression of NRP1 is associated with melanoma invasion and metastasis. In conclusion, increased NRP1 expression is associated with disease progression and reduced survival in patients with melanoma, and is a promising prognostic molecular marker for this disease.

Reduced expression of SRY-box containing gene 17 correlates with an unfavorable melanoma patient survival

SRY-box containing gene 17 (Sox17), a transcription factor, is considered as an antagonist to canonical Wnt/β‑catenin signaling in several types of malignant tumors. As the influence of Sox17 in the pathogenesis of human melanoma is still unknown, the investigation of Sox17 expression in melanoma is warranted and its prognostic value is of great interest. In the present study, Sox17 expression was examined in 525 cases of melanocytic lesions (33 common acquired nevi, 59 dysplastic nevi, 291 primary melanomas and 142 metastatic melanomas) at different stages by tissue microarray. The correlation of Sox17 expression with melanoma progression and its prognostic value in melanoma patients were examined. We also analyzed the correlation between Sox17 and cyclin-dependent kinase inhibitor p27 expression in 374 melanoma samples. The results showed that Sox17 expression was significantly decreased in primary and metastatic melanoma compared to common acquired nevi and dysplastic nevi (P=2.4x10-17). Furthermore, Sox17 expression was inversely correlated with American Joint Committee on Cancer stage (P=4.6x10-15), thickness (P=0.00004) and ulceration (P=0.03). Notably, reduced Sox17 expression was correlated with a poorer overall and disease-specific 5- and 10-year survival of the patients. Multivariate Cox regression analyses indicated that Sox17 is an independent prognostic marker for melanoma patients. Moreover, we found a significant positive correlation between Sox17 and p27 expression in melanoma biopsies; their concomitant expression was closely correlated with the survival of melanoma patients. Taken together, decreased Sox17 expression is correlated with melanoma progression, an unfavorable survival of melanoma patients and is an independent molecular prognostic factor for melanoma.

Abstract A33: Loss of tumor suppressors KAI1 and p27 identifies a unique subgroup of primary melanoma patients with poor survival

Objective: Melanoma is a highly heterogeneous disease. A certain subgroup of patients with primary melanomas exhibits a greater potential to develop metastatic disease and thus has worse survival. The present study attempts to identify the molecular features shared by metastatic melanomas and the subgroup of primary melanomas, and use the information to assist clinicians to design personalized therapy for primary melanoma patients. Patients and methods: Seven previously reported biomarkers, including BRAF, Dicer, Fbw7, KAI1, MMP2, p27 and Tip60, were investigated. A training cohort with 250 melanoma patients (145 primary melanomas and 105 metastatic melanomas) and an independent cohort with 92 primary melanoma patients were used for the study. Logistic regression analysis was used to identify discriminate biomarkers of metastatic melanoma from primary melanoma. Kaplan-Meier survival and multivariate Cox proportional hazard regression analysis were performed to assess the significance of the molecular signature in the prognosis of melanoma. Results: In the training cohort, we found the loss expression of KAI1 and p27 to be most significant between metastatic and primary melanoma. The primary melanoma patients with loss expression of both KAI1 and p27 had poor 5-year survival in both training cohort (P = 0.002) and independent cohort (P = 0.03). Multivariate Cox regression analysis showed that the KAI1-/p27- signature was an independent factor for disease- specific survival (P = 0.004). More important, compared to KAI1 and p27 as an individual prognostic marker, the KAI1-/p27- signature is more closely associated with melanoma patient survival: the P value is 0.004 for KAI1-/p27-, and 0.044, 0.181 for KAI1 and p27, respectively. Conclusion: Loss of both KAI1 and p27 defines a subgroup of primary melanoma patients with poor prognosis, and the combination signature is better than individual biomarker. The clinical significance of KAI1 and p27 warrants further assessment in prospective clinical trials. Citation Format: Yabin Cheng, Guohong Zhang, Yun Tang, Guangdi Chen, Gholamreza Safaee, Annand Rotte, Magdalena Martinka, Kevin McElwee, Youwen Zhou. Loss of tumor suppressors KAI1 and p27 identifies a unique subgroup of primary melanoma patients with poor survival. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Melanoma: From Biology to Therapy; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(14 Suppl):Abstract nr A33.