Glioblastoma（GBM） is the most malignant type of primary brain tumor, and the main treatment includes maximum surgical resection, followed by chemotherapy and radiotherapy.DNA-damaging chemotherapeutic drugs and radiation can cause widespread cellular senescence, a phenomenon called therapy-induced senescence（TIS）. While TIS may exert tumor-suppressive effects in the early stage of treatment, chemoradiotherapy eventually leads to the accumulation of senescent cells in both tumor and normal tissues, resulting in unfavorable clinical outcomes. This review summarized the research progress of TIS in the treatment of GBM, with a focus on the mechanisms by which senescent cells promote tumorigenesis through reprogramming and reshaping the tumor microenvironment. We also prospected the challenges and opportunities of senolytics, a novel anti-senescence therapy, in the treatment of GBM, hoping to help improve the outcome of GBM treatment and promote further research in basic and clinical application in GBM treatment.

Purpose: This study was based on hepatocellular carcinoma (HCC) patients of early-stage to explore the diagnostic capability and possible production causes of anti-GNAS autoantibody. Methods: We evaluated the frequency of anti-GNAS autoantibody in sera from patients with early-stage HCC by enzyme-linked immunosorbent assay (ELISA) and the expression of GNAS protein in early-stage HCC tissues by immunohistochemistry. Western blotting (WB) and real-time polymerase chain reaction (RT-PCR) were utilized to examine the expressions of GNAS protein and mRNA in cell lines. GEO and International Cancer Genome Consortium (ICGC) databases were inquired to explore mRNA expression and mutation of GNAS in HCC tissues. Results: The positive rates of anti-GNAS autoantibody in HCC patients at clinical stage I (78.1 %) and clinical stage II (57.1 %) were all significantly higher than that in healthy control (20 %). There was also a significant difference in GNAS protein expression between HCC and its adjacent normal liver tissues. The results from WB and RT-PCR showed a significant difference at the mRNA level but no statistical difference at the protein level between HCC and normal liver cell lines. The difference in mRNA level between HCC and adjacent normal liver tissues was verified to be significant. Furthermore, the ICGC database demonstrated a 10.6 % mutation frequency for GNAS in HCC patients. Conclusion: The coordination of elevated anti-GNAS autoantibody, high expression of GNAS in the mRNA and protein levels in HCC, and high frequency of GNAS mutation indicates that anti-GNAS autoantibody may be used as an early indicator of HCC.

Esophageal squamous cell carcinoma is a severe malignancy for its high mortality and poor prognosis. Mainstay chemotherapies cause serious side effects for their ways of inducing cell death. Oridonin is the main bioactive constituent from natural plants that has anticancer ability and weak side effects. The proteomics method is efficient to understand the anticancer mechanism. However, proteins identified by proteomics aimed at understanding oridonin's anticancer mechanism is seldom overlapped by different groups. This study used proteomics based on two-dimensional electrophoresis sodium dodecyl sulfate-polyacrylamide gel electrophoresis (2-DE SDS-PAGE) integrated with mass spectrometry and Gene Set Enrichment Analysis (GSEA) to understand the anticancer mechanism of oridonin on esophageal squamous cell carcinoma (ESCC). The results showed that oridonin induced ESCC cell death via apoptosis by decreasing the protein expression of LASP1 and PDLIM1.

The chemokine receptor CXCR3 is functionally pleiotropic, not only recruiting immune cells to the inflamed liver but also mediating the pathological process of cholestatic liver injury (CLI). However, the mechanism of its involvement in the CLI remains unclear. Both alpha-naphthylisothiocyanate (ANIT) and triptolide are hepatotoxicants that induce CLI by bile acid (BA) dysregulation, inflammation, and endoplasmic reticulum (ER)/oxidative stress. Through molecular docking, CXCR3 is a potential target of ANIT and triptolide. Therefore, this study aimed to investigate the role of CXCR3 in ANIT- and triptolide-induced CLI and to explore the underlying mechanisms. Wild-type mice and CXCR3-deficient mice were administered with ANIT or triptolide to compare CLI, BA profile, hepatic recruitment of IFN-γ/IL-4/IL-17+CD4+T cells, IFN-γ/IL-4/IL-17+iNKT cells and IFN-γ/IL-4+NK cells, and the expression of ER/oxidative stress pathway. The results showed that CXCR3 deficiency ameliorated ANIT- and triptolide-induced CLI. CXCR3 deficiency alleviated ANIT-induced dysregulated BA metabolism, which decreased the recruitment of IFN-γ+NK cells and IL-4+NK cells to the liver and inhibited ER stress. After triptolide administration, CXCR3 deficiency ameliorated dysregulation of BA metabolism, which reduced the migration of IL-4+iNKT cells and IL-17+iNKT cells and reduced oxidative stress through inhibition of Egr1 expression and AKT phosphorylation. Our findings suggest a detrimental role of CXCR3 in ANIT- and triptolide-induced CLI, providing a promising therapeutic target and introducing novel mechanisms for understanding cholestatic liver diseases.

BACKGROUND:As the main component of oral contraceptives (OCs), ethinylestradiol (EE) has been widely applied as a model drug to induce murine intrahepatic cholestasis. The clinical counterpart of EE-induced cholestasis includes women who are taking OCs, sex hormone replacement therapy, and susceptible pregnant women. Taking intrahepatic cholestasis of pregnancy (ICP) as an example, ICP consumes the medical system due to its high-risk fetal burden and the impotency of ursodeoxycholic acid in reducing adverse perinatal outcomes.AIM:To explore the mechanisms and therapeutic strategies of EE-induced cholestasis based on the liver immune microenvironment.METHODS:Male C57BL/6J mice or invariant natural killer T (iNKT) cell deficiency (Jα18-/- mice) were administered with EE (10 mg/kg, subcutaneous) for 14 d.RESULTS:Both Th1 and Th2 cytokines produced by NKT cells increased in the liver skewing toward a Th1 bias. The expression of the chemokine/chemokine receptor Cxcr6/Cxcl16, toll-like receptors, Ras/Rad, and PI3K/Bad signaling was upregulated after EE administration. EE also influenced bile acid synthase Cyp7a1, Cyp8b1, and tight junctions ZO-1 and Occludin, which might be associated with EE-induced cholestasis. iNKT cell deficiency (Jα18-/- mice) robustly alleviated cholestatic liver damage and lowered the expression of the abovementioned signaling pathways.CONCLUSION:Hepatic NKT cells play a pathogenic role in EE-induced intrahepatic cholestasis. Our research improves the understanding of intrahepatic cholestasis by revealing the hepatic immune microenvironment and also provides a potential clinical treatment by regulating iNKT cells.

Hepatocellular carcinoma (HCC) exerts huge effects on the health burden of the world because of its high mortality and poor prognosis. HCC is often clinically detected late in patients. If HCC could be detected and treated earlier, the survival rate of patients will be greatly improved. Therefore, identifying specific biomarkers is urgent and important for HCC. The liver is also recognized as an immune organ. The occurrence of HCC is related to exacerbation of immune tolerance and/or immunosurveillance escape. The host immune system plays an important role in the recognition and targeting of tumor cells in cancer immunotherapy, as can be seen from the clinical success of immune checkpoint inhibitors and chimeric antigen receptor (CAR) T cells. Thus, there is a pressing medical need to discover immunodiagnostic biomarkers specific to HCC for understanding the pathological mechanisms of HCC, especially for immunotherapy targets. We have reviewed the existing literature to summarize the immunodiagnostic markers of HCC, including autoantibodies against tumor-associated antigens (TAAs) and exosomes, to provide new insights into HCC and early detection of this deadly cancer.

Acute myeloid leukemia (AML) is a hematological malignancy derived from immature myeloid cells, which have the characteristics of abnormal proliferation and differentiation. Glycolysis has been a popular topic of research in recent years, with increasing uptake and consumption of glucose. The present study aimed to investigate the glycolysis of tumor cells in patients with AML; in particular, how programmed cell death 1 ligand 1 (PD‑L1) regulates tumor cells glycolysis using real time PCR (RT‑PCR), western blotting and flow cytometry. PD‑L1 high expression predicted poor outcome in patients with AML in the public database Gene Expression Profiling Interactive Analysis. PD‑L1 expression was decreased in the samples from patients with AML with complete remission compared to that in patients with relapsed or refractory AML. In AML cell lines, glycolysis‑associated genes ALDOA, PGK1, LDHA and HK2 were highly expressed in a PD‑L1 high‑expressed cell line. Overexpressed PD‑L1 enhanced glucose consumption and the extracellular acidification rate, accompanied by decreased apoptosis and accumulation of cells in the S phase. In contrast, the apoptosis rate of tumor cells and the percentage of cells in the S phase were significantly increased following PD‑L1 knockdown in the THP1 cell line. HK2 and LDHA expression decreased after AML tumor cells were treated with Akt inhibitor or rapamycin. In addition, the PD‑L1‑overexpressed cell line (PD‑L1‑OV) MOLM‑13 exhibited rapid tumor progression. Glycolysis‑associated genes were highly expressed in tumor tissues of PD‑L1‑OV MOLM‑13, with increased Ki67. Based on these findings, PD‑L1 may be considered as a suitable marker for prognosis and treatment in the clinical setting.

p62/IMP2 is an oncofetal protein that was first reported as a tumor-associated antigen in hepatocellular carcinoma (HCC). In our previous studies, we demonstrated a high frequency of p62/IMP2 autoantibodies appearing in various types of cancer. Therefore, we hypothesize that p62/IMP2 plays an important role in the progression of HCC, although the mechanism remains to be explored. In this study, we evaluated the expression of p62/IMP2 protein both in human tissues and liver cancer cell lines by immunohistochemistry and western blotting analysis and found that p62/IMP2 protein is overexpressed in human HCC tissue in comparison to normal human liver tissue. To explore the role that p62/IMP2 plays in HCC, p62/IMP2 was knocked out in two p62/IMP2-positive liver cancer cell lines (SNU449 and HepG2). Due to the low expression level of p62/IMP2 in SNU449, we overexpressed p62/IMP2 in this cell line. We subsequently demonstrated that high expression of p62/IMP2 in both cell lines can promote cell migration and invasion abilities in vitro by activating the Wnt/β-catenin pathway. We also used the Wnt/β-catenin pathway inhibitor, XAV 939, and a phosphoproteome assay to confirm our findings. Conclusion: Our results suggest that p62/IMP2 is an essential regulator of Wnt signaling pathways and plays an important role in HCC progression and metastasis.

Abstract Esophageal Cancer estimated leading deaths are more than 400,000 every year in worldwide, while 80% due to esophageal squamous cell carcinoma. In worldwide, esophageal cancer leading death ranks in sixth among cancer leading causes of death, while the 5-year survival rate of esophageal cancer is 19%. Oridonin, a tetracyclic diterpenes compound, has been indicated the anti-tumor activity for digestive system tumors. To assess the anticancer activity of oridonin on esophageal cancer, we performed cell proliferation experiment of oridonin on esophageal squamous cell carcinoma cell line TE-8 using SRB assay. We found that IC50 of oridonin on TE-8 treated for 72h is 3.0μM, which is better than positive control Gemcitabine treatment on TE-8 for 72h (5.7 μM), as well as TE-8 cells shrunk and floated with increasing concentration on oridonin. At the same time, oridonin induced a dose-dependent inhibition of cell colony formation of esophageal cancer cell line. Colony numbers of TE-8 cell line treated with 1μM and 2μM oridonin are 111 and 54, respectively, which are significantly different (p<0.05). Accompanying with marked morphological changes, flow cytometer results showed that oridonin triggered sub-G0/G1 cell cycle arrest in a dose-dependent manner. After treatment by 10μM and 20μM oridonin for 24h, percentage of sub-G0/G1 cells were 25.5 and 55.3%, respectively, which are significantly different (p<0.05). The apoptosis results showed that oridonin treatment led to the accumulation of cells in early-(Annexin V+/PI-) and late-stage (Annexin V+/PI+) apoptosis in a dose-dependent manner. Citation Format: Xiaojun Zhang, Mengtao Xing, Yangcheng Ma, Xiao Wang, Jianying Zhang. Oridonin inhibits cell growth of esophageal carcinoma cells via induction of apoptosis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1854.

Abstract Although the incidence rate of lung cancer has declined over the past few years, it remains the leading cause of cancer death worldwide. The Cancer Genome Atlas has provided invaluable resources for cancer research. The development of next generation sequencing technology and evolution of data analysis tools make it possible to interpret the underlying mechanism of oncogenesis. In current study, normalized FPKM data from 572 samples derived from patients with lung adenocarcinoma (LUAD) from TCGA was used to do weighted gene co-expression network analysis (WGCNA) to identify gene modules associated with lung adenocarcinoma. Raw counts data was used to identify differentially expressed genes in LUAD by using DESeq2 in R (version 3.4.2). WGCNA results shows that magenta (r = 0.47, p <0.001)), black (r = 0.40, p <0.001)) and yellow (r = -0.88, p <0.001) modules are top 3 modules that associated with LUAD and cancer stage. Gene Ontology over-representation analysis shows that genes in magenta modules are involved with transcript elongation; genes in black modules are enriched in cell meiosis related biological processes, including chromosome segregation, nuclear division, DNA replication and mitotic nuclear division et al; genes in yellow module are enriched in angiogenesis, leukocyte migration and hemostasis. It is noteworthy that genes in magenta and black modules are upregulated and in yellow modules are downregulated. GOLM1 and MUC4 in magenta module play certain roles in cancer development. Several genes in MEGA family, including MAGEA3, MAGEA6, MEGEA16 and MEGEA1, and IGF2BP1 in black module have been reported to be overexpressed in various kinds of tumors. However, whether these in silicon discovered genes can be used to distinguish LUAD from controls still needs to be further evaluated. Keywords: lung adenocarcinoma; TCGA; gene expression analysis; bioinformatics; cancer biomarker Citation Format: Jianxiang Shi, Mengtao Xing, Chenglin Luo, Xiao Wang, Liping Dai, Jianying Zhang. Revisiting TCGA data identifies key genes in lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3292.

Abstract Insulin- like growth factor 2 mRNA binding proteins (IMPs) Family contains three members: IMP1, IMP2 and IMP3. IMPs are oncofetal proteins, expressed during embryogenesis and lost in most tissues in adults. Overexpressed IMPs have been reported in various types of cancers. They share similar protein structure in mammals and have similar functions: to bind specific mRNAs, stabilize and extend their half-life. IMP2 was first reported as a tumor-associated antigen in HCC. However, only a few research was done on what a role of p62/IMP2 plays on HCC progression. Previous studies show that IMP2 is overexpressed in HCC tissues and in several human HCC cell lines. Western blotting was performed for different HCC cells has a relatively low expression in SNU449 and higher expression in HepG2 and Hep3b. Stable IMP2 overexpression SNU449 cell line was made by IMP2 cDNA plasmid transfection; stable IMP2 knockout Hep3b and HepG2 cell lines was made by IMP2 Crispr-Cas9 KO plasmid transfection. Wound healing assay has shown that overexpression of p62/IMP2 can significantly enhance cell migration ability in SNU449 cells. On the contrary, the depletion of p62/IMP2 can reduce the migration ability in HepG2 and Hep3b cells (p<0.05). Furthermore, we analyzed HCC gene expression data from TCGA. Median count of IMP2 was used to separate HCC expression data into high and low IMP2 expression group to conduct differential gene expression analysis. Gene Ontology over-representation analysis shows differentially expressed genes (adjusted p<0.05) mainly enriched in membrane part and plasma membrane. The expression of several genes involved in EMT regulation such as CTNNB1, SMAD2, SNAI2, AKT2 shows significant difference (adjust p<0.05). Western blotting assay was performed to show whether IMP2 is involved in EMT regulation. Wnt/beta-catenin pathway and TGF-beta/SMAD pathway were both activated and further upregulated SNAIL, which is an E-cadherin inhibitor. Western blotting results showed that E-cadherin was inhibited in IMP2 overexpressed SNU449 cells, which is supportive to our hypothesis. Citation Format: Mengtao Xing, Jianxiang Shi, Jitian Li, Beibei Chen, Giulio Francia, Jianying Zhang. P62/IMP2 promotes metastasis in hepatocellular carcinoma (HCC) by regulating EMT in different signaling pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2027.

The driving roles of fusion genes during tumorigenesis have been recognized for decades, with efficacies demonstrated in clinical diagnosis and targeted therapy. With advances in sequencing technologies and computational biology, a surge in the identification of fusion genes has been witnessed during the past decade. The discovery and presence of splicing based fusions in normal tissues have challenged our canonical conceptions on fusion genes and offered us novel medical opportunities. The specificity of fusion genes to neoplastic tissues and their diverse functionalities during carcinogenesis foster them as promising tools in the battle against cancer. It is time to re-visit and comb through our cutting-edge knowledge on fusion genes to accelerate clinical translation of these internal markers. Urged as such, we are encouraged to categorize fusion events according to mechanisms leading to their generation, oncological consequences and clinical implications, offer insights on fusion occurrence across tumors from the system level, highlight feasible practices in fusion-related pharmaceutical development, and identify understudied yet important niches that may lead future research trend in this field.

Abstract Hepatocellular carcinoma (HCC) is one of the most common type of cancer and the third leading cause of cancer related death in worldwide. Insulin-like growth factor 2 mRNA binding protein 2 (p62/IMP2/IGF2BP2) was originally reported as an autoantigen in HCC which belongs to IMPs family. The IMPs family contains three members: IMP1, IMP2 and IMP3, which share similar protein structure. IMPs are oncofetal proteins, which are expressed during embryogenesis and lost in most tissues in adults. Therefore, p62/IMP2 can be used as a potential biomarker for early diagnosis of HCC. We have evaluated the expression of p62/IMP2 protein both in human tissues and HCC cell lines by immunohistochemistry and western blotting analysis, and found that p62/IMP2 protein is overexpressed in human HCC tissues compared with normal human liver tissues (score= 10.30, n=40 vs. score =5.23, n=30, p<0.05). The expression of p62/IMP2 was scored by immune-staining intensity and positively immune-staining cell numbers. At the same time, we found that p62/IMP2 was also overexpressed in several HCC cell lines. Stable p62/IMP2 overexpression and depletion SNU449 human HCC cell line was made by using p62/IMP2 cDNA transfection and lentivirus mediated knockdown approaches, and it was further confirmed by western blotting analysis. Wound healing assay has shown that overexpression of p62/IMP2 can significantly enhance the cell migration ability in SNU449 cells. On the contrary, the depletion of p62/IMP2 can reduce the migration ability in SNU449 cells (p<0.05). Moreover,MTT assay has also shown p62/IMP2 overexpressed cells may have a higher rate of proliferation at 72h(p<0.05). In addition, we have observed that overexpression of p62/IMP2 can significant inhibit the expression of E-cadherin in western blotting analysis. In summary, our data would suggest that overexpression p62/IMP2 may promote metastasis via inhibition of E-cadherin expression in HCC. Citation Format: Mengtao Xing, Pei Li, Jitian Li, Jianxiang shi, Liping Dai, Giulio Francia, Jianying Zhang. Overexpression of P62/IMP2 promotes metastasis in hepatocellular carcinoma via inhibition of E-cadherin expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4850. doi:10.1158/1538-7445.AM2017-4850

We evaluated autoantibodies against nine tumor-associated antigens, including p62, p16, Koc, p53, Cyclin B1, Cyclin E, Survivin, HCC1, and RalA as serological markers in lung cancer. Enzyme-linked immunosorbent assay (ELISA) was used to detect autoantibodies in sera from 50 lung cancer patients and 42 normal controls. Then, four tumor-associated antigens of higher values were selected and validated in sera from validation group. Western blot and serum absorption test were used to confirm positive findings from ELISA. When cutoff values were set as mean optical density values plus 3 standard deviation of normal controls, the positive rate of autoantibodies against four tumor-associated antigens (Survivin, Cyclin B1, HCC1, and p53) reached 32%, 20%, 22%, and 18%, with area under the curve values of 0.653, 0.767, 0.622, and 0.623 in sera from 50 lung cancer, respectively (all p < 0.05). Results from the validation group confirmed the results. When lung cancer patients were divided by their clinicopathological characteristics into different subgroups, we have found that serum anti-Cyclin B1 and anti-HCC1 autoantibodies increased in stages 1, 2, and 3 lung cancer; anti-Survivin autoantibodies increased in stages 2 and 3 lung cancer; and anti-p53 autoantibody only increased in stage 1 when compared with their corresponding levels in controls (all p < 0.05). Serum anti-Cyclin B1 and anti-Survivin autoantibodies increased with disease histological grade 2 and 3 (both p < 0.05). And higher serum level of anti-p53 autoantibodies is positively associated with tumor size. Parallel utilization of these four anti-tumor-associated antigens (any positive) can increase sensitivity to 65.0% at 100% specificity with area under the curve of 0.908 ( p < 0.001) in lung cancer detection in validation group. Our results suggest that autoantibodies against these four tumor-associated antigens have higher values in lung cancer detection, and serum anti-Cyclin-B1 has the potential to serve as novel non-invasive biomarkers in early-stage lung cancer.

In this study, enzyme-linked immunosorbent assay has been used to examine the frequencies of serum autoantibodies against two candidate tumor-associated antigens intensively selected from the Human Protein Atlas database, in combination with 13 tumor-associated antigens available from our lab in sera from 44 OC patients and 50 normal healthy controls. Conventional evaluation (mean + 3SD as the cutoff value to determine a positive reactivity), receiver operating characteristic curve analyses, and classification tree analysis were further used to evaluate the diagnostic performance of autoantibodies against these tumor-associated antigens (anti-tumor-associated antigens) in ovarian cancer. For single anti-tumor-associated antigen, when the cutoff values were set as mean + 3SD of normal healthy controls, NPM1, MDM2, PLAT, p53, and c-Myc could achieve sensitivity higher than 20% at 98% specificity. Combinational utilization of autoantibodies against MDM2, PLAT, NPM1, 14-3-3 Zeta, p53, and RalA achieved the optimal diagnostic performance with 72.7% sensitivity at 96% specificity. Receiver operating characteristic curve analysis showed that the area under the receiver operating characteristic curves of autoantibodies against c-Myc, NPM1, MDM2, p16, p53, and 14-3-3 Zeta were greater than 0.80. This indicated that these tumor-associated antigens held high potential to serve as diagnostic biomarkers in ovarian cancer detection. Decision tree analysis indicated that anti-c-Myc held high potential in the detection of ovarian cancer. Further studies are warranted to validate the diagnostic performance of these anti-tumor-associated antigens with high area under the receiver operating characteristic curve, including autoantibodies against c-Myc, MDM2, PLAT, NPM1, 14-3-3 Zeta, p53, and RalA.

BACKGROUND:The prostate-specific antigen (PSA) testing has been widely implemented for the early detection and management of prostate cancer (PCa). However, the lack of specificity has led to overdiagnosis, resulting in many possibly unnecessary biopsies and overtreatment. Therefore, novel serological biomarkers with high sensitivity and specificity are of vital importance needed to complement PSA testing in the early diagnosis and effective management of PCa. This is particularly critical in the context of PCa health disparities, where early detection and management could help reduce the disproportionately high PCa mortality observed in African-American men. Previous studies have demonstrated that sera from patients with PCa contain autoantibodies that react with tumor-associated antigens (TAAs). METHODS:The serological proteome analysis (SERPA) approach was used to identify tumor-associated antigens (TAAs) of PCa. In evaluation study, the level of anti-NPM1 antibody was examined in sera from test cohort, validation cohort, as well as European-American (EA) and African-American (AA) men with PCa by using immunoassay. RESULTS:Nucleophosmin 1 (NPM1) as a 33 kDa TAA in PCa was identified and characterized by SERPA approach. Anti-NPM1 antibody level in PCa was higher than in benign prostatic hyperplasia (BPH) patients and healthy individuals. Receiver operating characteristic (ROC) curve analysis showed similar high diagnostic value for PCa in the test cohort (area under the curve (AUC):0.860) and validation cohort (AUC: 0.822) to differentiate from normal individuals and BPH. Interestingly, AUC values were significantly higher for AA PCa patients. When considering concurrent serum measurements of anti-NPM1 antibody and PSA, 97.1% PCa patients at early stage were identified correctly, while 69.2% BPH patients who had elevated PSA levels were found to be anti-NPM1 negative. Additionally, anti-NPM1 antibody levels in PCa patients at early stage significantly increased after surgery treatment. CONCLUSION:This intriguing data suggested that NPM1 can elicit autoantibody response in PCa and might be a potential biomarker for the immunodiagnosis and prognosis of PCa, and for supplementing PSA testing in distinguishing PCa from BPH. Prostate 76:1375-1386, 2016. © 2016 Wiley Periodicals, Inc.

Autoantibodies against intracellular tumor-associated antigens (TAAs) are commonly found in human cancers. In this study, we characterized the serum autoantibody response to the RalA, Ras-like GTPase, in patients with prostate cancer (PCa). The autoantibodies were detected by immunofluorescence assay in PCa cell lines, ELISA, and immunoblotting in 339 serum samples from patients with PCa and benign prostatic hyperplasia (BPH), and in normal human sera (NHS). The expression of RalA in prostate tumor tissues was evaluated by immunohistochemistry (IHC) in tumor microarrays. The autoantibody level to RalA (median) in NHS was significantly lower than in PCa (0.053 vs 0.138; P < 0.001) and BPH (0.053 vs 0.132; P < 0.005) groups. The circulating anti-RalA autoantibody could distinguish PCa patients from normal individuals with the area under the receiver operating characteristic (ROC) curve (AUC) performing at 0.861, with sensitivity of 52.9% and specificity of 91.0%. Elevation in serum immunoreactivity was observed in PCa patients after radical prostatectomy. The combined use of both anti-RalA autoantibody and PSA showed a significantly higher discriminatory ability compared with either of those markers alone. RalA protein expression was detected by IHC in 85.3% of tumor tissues from PCa patients, but without significant difference compared to BPH or normal control tissues. Together, our study shows the additional benefits of anti-RalA autoantibody as a potential serological biomarker for PCa, particularly in patients with normal PSA, and further demonstrate the utility of biomarker combinations in the immunodiagnosis of PCa.