Purpose: Immune checkpoint blockade (ICB) therapy induces durable tumor regressions in a minority of patients with cancer. In this study, we aimed to identify kinase inhibitors that were capable of increasing the antimelanoma immunity.Experimental Design: Flow cytometry-based screening was performed to identify kinase inhibitors that can block the IFNg-induced PD-L1 expression in melanoma cells. The pharmacologic activities of regorafenib alone or in combination with immunotherapy in vitro and in vivo were determined. The mechanisms of regorafenib were explored and analyzed in melanoma patients treated with or without anti-PD-1 using The Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) datasets.Results: Through screening of a kinase inhibitor library, we found approximately 20 agents that caused more than half reduction of cell surface PD-L1 level, and regorafenib was one of the most potent agents. Furthermore, our results showed that regorafenib, in vitro and in vivo, strongly promoted the antitumor efficacy when combined with IFNg or ICB. By targeting the RET-Src axis, regorafenib potently inhibited JAK1/2-STAT1 and MAPK signaling and subsequently attenuated the IFNg-induced PD-L1 and IDO1 expression without affecting MHC-I expression much. Moreover, RET and Src cohigh expression was an independent unfavorable prognosis factor in melanoma patients with or without ICB through inhibiting the antitumor immune response.Conclusions: Our data unveiled a new mechanism of alleviating IFNg-induced PD-L1 and IDO1 expression and provided a rationale to explore a novel combination of ICB with regorafenib clinically, especially in melanoma with RET/Src axis activation.

Autophagy is a degradative pathway that delivers cellular components to the lysosome for degradation. The role of autophagy in cell differentiation is poorly understood. Here we show that CaMKII can directly phosphorylate Beclin 1 at Ser90 to promote K63-linked ubiquitination of Beclin 1 and activation of autophagy. Meanwhile, CaMKII can also promote K63-linked ubiquitination of inhibitor of differentiation 1/2 (Id-1/2) by catalyzing phosphorylation of Id proteins and recruiting TRAF-6. Ubiquitinated Id-1/Id-2 can then bind to p62 and be transported to autolysosomes for degradation. Id degradation promotes the differentiation of neuroblastoma cells and reduces the proportion of stem-like cells. Our study proposes a mechanism by which autophagic degradation of Id proteins can regulate cell differentiation. This suggests that targeting of CaMKII and the regulation of autophagic degradation of Id may be an effective therapeutic strategy to induce cell differentiation in neuroblastoma.

Rationale: Nasopharyngeal carcinoma (NPC) is the most frequent head and neck tumor in South China. The presence of cancer stem cells (CSCs) in NPC contributes to tumor maintenance and therapeutic resistance, while the ability of CSCs to escape from the apoptosis pathway may render them the resistant property to the therapies. Inhibitor of apoptosis proteins family proteins (IAPs), which are overexpressed in nasopharyngeal carcinoma stem cells, may play an important role in maintaining nasopharyngeal cancer stem cell properties. Here, we develop a novel CSC-targeting strategy to treat NPC through inhibiting IAPs.Methods: Human NPC S-18 and S-26 cell lines were used as the model system in vitro and in vivo. Fluorescence activated cell sorting (FACS) assay was used to detect nasopharyngeal SP cells and CD44+ cells. The characteristics of CSCs were defined by sphere suspension culture, colony formation assay and cell migration. The role of XIAP on the regulation of Sox2 protein stability and ERK1-mediated phosphorylation of Sox2 signaling pathway were analyzed using immunoblotting, immunoprecipitation, immunofluorescence, phosphorylation mass spectrometry, siRNA silencing and plasmid overexpression. The correlation between XIAP and Sox2 in NPC biopsies and their role in prognosis was performed by immunohistochemistry. APG-1387 or chemotherapies-induced cell death and apoptosis in S-18 and S-26 were determined by WST, immunoblotting and flow cytometry assay.Results: IAPs, especially X chromosome-linked IAP (XIAP), were elevated in CSCs of NPC, and these proteins were critically involved in the maintenance of CSCs properties by enhancing the stability of Sox2. Mechanistically, ERK1 kinase promoted autophagic degradation of Sox2 via phosphorylation of Sox2 at Ser251 and further SUMOylation of Sox2 at Lys245 in non-CSCs. However, XIAP blocked autophagic degradation of Sox2 by inhibiting ERK1 activation in CSCs. Additionally, XIAP was positively correlated with Sox2 expression in NPC tissues, which were associated with NPC progression. Finally, we discovered that a novel antagonist of IAPs, APG-1387, exerted antitumor effect on CSCs. Also, the combination of APG-1387 with CDDP /5-FU has a synergistic effect on NPC.Conclusion: Our study highlights the importance of IAPs in the maintenance of CSCs in NPC. Thus, XIAP is a promising therapeutic target in CSCs and suggests that NPC patients may benefit from a combination treatment of APG-1387 with conventional chemotherapy.

BACKGROUNDTo develop and validate a nomogram based on log of odds between the number of positive lymph node and the number of negative lymph node (LODDS) in predicting the overall survival (OS) and cancer specific survival (CSS) for epithelial ovarian cancer (EOC) patients.MATERIALS AND METHODSA total of 10,692 post-operative EOC patients diagnosed between 2004 and 2013 were obtained from the Surveillance, Epidemiology, and End Results (SEER) database and randomly divided into training (n = 7,021) and validation (n = 3,671) cohorts. Multiple clinical pathological parameters were assessed and compared with outcomes. Parameters significantly correlating with outcomes were used to build a nomogram. Bootstrap validation was subsequently used to assess the predictive value of the model.RESULTSIn the training set, age at diagnosis, race, marital status, tumor location, stage, grade and LODDS were correlated significantly with outcome in both the univariate and multivariate analyses and were used to develop a nomogram. The nomogram demonstrated good accuracy in predicting OS and CSS, with a bootstrap-corrected concordance index of 0.757 (95% CI, 0.746-0.768) for OS and 0.770 (95% CI, 0.759-0.782) for CSS. Notably, in this population our model performed favorably compared to the currently utilized Federation of Gynecology and Obstetrics (FIGO) model, with concordance indices of 0.699 (95% CI, 0.688-0.710, P < 0.05) and 0.719 (95% CI, 0.709- 0.730, P < 0.05) for OS and CSS, respectively. Using our nomogram in the validation cohort, the C-indices were 0.757 (95% CI, 0.741-0.773, P < 0.05, compared to FIGO) for OS and 0.762 (95% CI, 0.746-0.779, P < 0.05, compared to FIGO) for CSS.CONCLUSIONSLODDS works as an independent prognostic factor for predicting survival in patients with EOC regardless of the tumor stage. By incorporating LODDS, our nomogram may be superior to the currently utilized FIGO staging system in predicting OS and CSS among post-operative EOC patients.

Protein tyrosine kinase 6 (PTK6) is a non-receptor tyrosine kinase and works as an oncogene in various cancers. Recently, PTK6 has been used as a therapeutic target for breast cancer patients in a clinical study. However, the prognostic value of PTK6 in bladder cancer (BC) remains vague. Therefore, we retrieved 3 independent investigations of Oncomine database and found that PTK6 is highly expressed in BC tissues compared with corresponding normal controls. Similar results were also observed in clinical specimens at both mRNA and protein levels. Immunohistochemical analysis indicated that PTK6 overexpression was highly related to the T classification, N classification, grade, recurrence, and poor prognosis of BC patients. Furthermore, we demonstrated that when PTK6 expression was knocked down by siRNAs, cell proliferation and migration were considerably inhibited in BC cell lines T24 and EJ. By these approaches, we are intended to elucidate PTK6 may be a reliable therapeutic target in BC and might benefit from PTK6 inhibitors in the future.