Personalized neoantigen vaccine enhances the therapeutic efficacy of bevacizumab and anti-PD-1 antibody in advanced non-small cell lung cancer.

Clinically, a considerable number of non-small cell lung cancer (NSCLC) patients are unable to receive or resist chemotherapy, and the efficacy of non-chemotherapy treatment strategies based on anti-angiogenic agents combined with immune checkpoint blockade is still unsatisfactory. Neoantigen vaccine, based on personalized tumor DNA mutations, could elicit tumor specific T cell infiltration into the tumor site, exerting potent anti-tumor efficacy. Here, we evaluated the feasibility and safety of a new antitumor strategy by adding neoantigen vaccine to the regimen of bevacizumab and anti-PD-1 antibody. Firstly, 7 novel immunogenic neoantigen peptides were identified and developed for neoantigen vaccine (LLCvac), which can elicit strong antitumor immune response in vivo. Then, in orthotopic lung cancer model, LLCvac further combining with bevacizumab and anti-PD-1 antibody exerted a stronger antitumor effect, exhibiting significant decrease of tumor volume without obvious toxicity. Furthermore, tumor immune microenvironment assessment also showed that the proportion of neoantigen-specific T cells in blood could be induced dramatically by the combined therapy. And a large amount of neoantigen-specific Ki67-positive CD8+ T cells were found in tumor tissues, which infiltrated tumor tissues effectively to kill tumor cells expressing identified neoantigens. Overall, these results suggested that this combined therapy could safely induce robust antitumor efficacy, serving as an effective chemotherapy-free strategy for NSCLC treatment.

Bulk and single-cell transcriptome reveal the immuno-prognostic subtypes and tumour microenvironment heterogeneity in HCC.

BACKGROUND & AIMS: Accumulating evidences suggest tumour microenvironment (TME) profoundly influence clinical outcome in hepatocellular carcinoma (HCC). Existing immune subtypes are susceptible to batch effects, and integrative analysis of bulk and single-cell transcriptome is helpful to recognize immune subtypes and TME in HCC. METHODS: Based on the relative expression ordering (REO) of 1259 immune-related genes, an immuno-prognostic signature was developed and validated in 907 HCC samples from five bulk transcriptomic cohorts, including 72 in-house samples. The machine learning models based on subtype-specific gene pairs with stable REOs were constructed to jointly predict immuno-prognostic subtypes in single-cell RNA-seq data and validated in another single-cell data. Then, cancer characteristics, immune landscape, underlying mechanism and therapeutic benefits between subtypes were analysed. RESULTS: An immune-related signature with 29 gene pairs stratified HCC samples individually into two risk subgroups (C1 and C2), which was an independent prognostic factor for overall survival. The machine learning models verified the immune subtypes from five bulk cohorts to two single-cell transcriptomic data. Integrative analysis revealed that C1 had poorer outcomes, higher CNV burden and malignant scores, higher sensitivity to sorafenib, and exhibited an immunosuppressive phenotype with more regulators, e.g., myeloid-derived suppressor cells (MDSCs), Mø_SPP1, while C2 was characterized with better outcomes, higher metabolism, more benefit from immunotherapy, and displayed active immune with more effectors, e.g., tumour infiltrating lymphocyte and dendritic cell. Moreover, both two single-cell data revealed the crosstalk of SPP1-related L-R pairs between cancer and immune cells, especially SPP1-CD44, might lead to immunosuppression in C1. CONCLUSIONS: The REO-based immuno-prognostic subtypes were conducive to individualized prognosis prediction and treatment options for HCC. This study paved the way for understanding TME heterogeneity between immuno-prognostic subtypes of HCC.

Potential oral VEGFR2 inhibitors: Treatment of wet age-related macular degeneration.

Wet age-related macular degeneration (w-AMD) is one of the leading causes of vision loss in industrialized countries. A large body of evidence suggests that inhibitors targeting VEGFR2 may be effective in the treatment of w-AMD. The identification of an oral VEGFR2 inhibitor for the treatment of w-AMD provides an opportunity for a route of administration other than intravitreal injection. While screening potent VEGFR2 inhibitors at the enzyme and cellular levels, ensuring the safety of the compounds was our primary strategy for screening optimal compounds. Finally, compound 16 was identified, exhibiting enhanced inhibition of VEGFR2 enzyme and proliferation of BaF3-TEL-VEGFR2 cells compared to Vorolanib. Compound 16 had a weak inhibitory effect on human Ether-a-go-go-related gene (hERG) channel currents, showing a cardiac safety profile similar to Vorolanib. Compound 16 showed no significant toxicity to human liver cell LX-2, indicating a liver safety profile similar to Vorolanib. The water solubility of compound 16 was found to be higher than that of Vorolanib when tested at pH = 7.4. In addition, compound 16 was found to inhibit VEGFR2 phosphorylation in human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner by WB assay. Furthermore, the in vitro preliminary evaluation of the drug-like properties of compound 16 showed remarkable plasma stability and moderate liver microsomal stability. Based on in vivo pharmacokinetic studies in ICR mice, compound 16 exhibited acceptable oral bioavailability (F = 20.2 %). Overall, these findings provide evidence that compound 16 is a leading potential oral drug candidate for w-AMD.

Design and synthesis of hemicyanine-based NIRF probe for detecting Aß aggregates in Alzheimer's disease.

Alzheimer's disease (AD), a progressive neurodegenerative disorder, has garnered increased attention due to its substantial economic burden and the escalating global aging phenomenon. Amyloid-ß deposition is a key pathogenic marker observed in the brains of Alzheimer's sufferers. Based on real-time, safe, low-cost, and commonly used, near-infrared fluorescence (NIRF) imaging technology have become an essential technique for the detection of AD in recent years. In this work, NIRF probes with hemicyanine structure were designed, synthesized and evaluated for imaging Aß aggregates in the brain. We use the hemicyanine structure as the parent nucleus to enhance the probe's optical properties. The introduction of PEG chain is to improve the probe's brain dynamice properties, and the alkyl chain on the N atom is to enhance the fluorescence intensity of the probe after binding to the Aß aggregates as much as possible. Among these probes, Z2, Z3, Z6, X3, X6 and T1 showed excellent optical properties and high affinity to Aß aggregates (Kd = 24.31 âˆ¼ 59.60 nM). In vitro brain section staining and in vivo NIRF imaging demonstrated that X6 exhibited superior discrimination between Tg mice and WT mice, and X6 has the best brain clearance rate. As a result, X6 was identified as the optimal probe. Furthermore, the docking theory calculation results aided in describing X6's binding behavior with Aß aggregates. As a high-affinity, high-selectivity, safe and effective probe of targeting Aß aggregates, X6 is a promising NIRF probe for in vivo detection of Aß aggregates in the AD brain.

Mechanisms of portal vein tumour thrombus formation and development in patients with hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common and aggressive human malignancies worldwide. Portal vein tumour thrombus (PVTT) is considered one of most fearful complications of HCC and is strongly associated with a poor prognosis. Clarification of the mechanisms underlying the formation and development of PVTT is crucial for developing novel therapeutic strategies for HCC patients. Several studies have been made to uncover that tumour microenvironment, stem cells, abnormal gene expression and non-coding RNAs deregulation are associated with PVTT in patients with HCC in the last decade. However, the exact molecular mechanisms of PVTT in patients with HCC are still largely unknown. In the present review, we briefly summarized the molecular mechanisms underlying the formation and development of PVTT in HCC.

Design, synthesis, and evaluation of N-methyl-propargylamine derivates as isoform-selective monoamine oxidases inhibitors for the treatment of nervous system diseases.

A novel series of N-methyl-propargylamine derivates were designed, synthesized, and evaluated as isoform-selective monoamine oxidases (MAO) inhibitors for the treatment of nervous system diseases. The in vitro studies showed some of the compounds exhibited considerable MAO-A selective inhibitory activity (IC50 of 14.86-17.16 nM), while some of the others exhibited great MAO-B selective inhibitory activity (IC50 of 4.37-17.00 nM). Further studies revealed that compounds A2 （IC50 against MAO-A: 17.16 ± 1.17 nM） and A5 (IC50 against MAO-B: 17.00 ± 1.10 nM) had significant abilities to protect PC12 cells from H2O2-induced apoptosis and reactive oxygen species (ROS) production. The parallel artificial membrane permeability assay showed A2 and A5 would be potent to cross the blood-brain barrier. The results indicated that A2 showed potential use in the therapy of MAO-A related diseases, such as depression and anxiety; while A5 exhibited promising ability in the treatment of MAO-B related diseases, such as Alzheimer's disease and Parkinson's disease.

Personalized neoantigen vaccine prevents postoperative recurrence in hepatocellular carcinoma patients with vascular invasion.

BACKGROUND: Clinically, prophylactic anti-recurrence treatments for hepatocellular carcinoma (HCC) patients after radical surgery are extremely limited. Neoantigen based vaccine can generate robust anti-tumor immune response in several solid tumors but whether it could induce anti-tumor immune response in HCC and serve as a safe and effective prophylactic strategy for preventing postoperative HCC recurrence still remain largely unclear. METHODS: Personalized neoantigen vaccine was designed and immunized for 10 HCC patients with high risk of postoperative recurrence in a prime-boost schedule. The safety and immune response were assessed through adverse events, tissue sequencing, ELISpot, TCR sequencing. The clinical response was evaluated by recurrence-free survival (RFS) and personalized circulating tumor DNA (ctDNA) sequencing. RESULTS: In the 10 enrolled patients, no obvious adverse events were observed during neoantigen vaccinations. Until the deadline of clinical trial, 8 of 10 patients were confirmed with clinical relapse by imaging, the other 2 patients remained relapse-free. From receiving first neoantigen vaccination, the median RFS of 10 patients were 7.4 months. Among 7 patients received all planned neoantigen vaccinations, 5 of them demonstrated neoantigen-induced T cell responses and have significantly longer RFS after radical surgery than other 5 patients without responsive neoantigens or only with prime vaccination and propensity scores matching control patients (p = 0.035). Moreover, tracking personalized neoantigen mutations in ctDNA could provide real-time evaluation of clinical response in HCC patients during neoantigen vaccination and follow up. CONCLUSION: Personalized neoantigen vaccine is proved as a safe, feasible and effective strategy for HCC anti-recurrence, and its progression could be sensitively monitored by corresponding neoantigen mutations in ctDNA, and thus provided solid information for individualized medicine in HCC. TRIAL REGISTRATION: This study was registered at Chinese Clinical Trial Registry; Registration number: ChiCTR1900020990 .

Accurate transcriptome assembly by Nanopore RNA sequencing reveals novel functional transcripts in hepatocellular carcinoma.

The long reads of Nanopore sequencing permit accurate transcript assembly and ease in discovering novel transcripts with potentially important functions in cancers. The wide adoption of Nanopore sequencing for transcript quantification, however, is largely limited by high costs. To address this issue, we developed a bioinformatics software, NovelQuant, that can specifically quantify long-read-assembled novel transcripts with short-read sequencing data. Nanopore Direct RNA Sequencing was carried out on three hepatocellular carcinoma (HCC) patients' tumor, matched portal vein tumor thrombus, and peritumor to reconstruct the HCC transcriptome. Then, based on the reconstructed transcriptome, NovelQuant was applied on Illumina RNA sequencing data of 59 HCC patients' tumor and paired peritumor to quantify novel transcripts. Our further analysis revealed 361 novel transcripts dysregulated in HCC and that 101 of them were significantly associated with prognosis. There were 19 novel prognostic transcripts predicted to be long noncoding RNAs (lncRNAs), and some of them had regulatory targets that were reported to be associated with HCC. Additionally, 42 novel prognostic transcripts were predicted to be protein-coding mRNAs, and many of them could be involved in xenobiotic metabolism. Moreover, the tumor-suppressive roles of two representative novel prognostic transcripts, CDO1-novel (lncRNA) and CYP2A6-novel (protein-coding mRNA), were further functionally validated during HCC progression. Overall, the current study shows a possibility of combining long- and short-read sequencing to explore functionally important novel transcripts in HCC with accuracy and cost-efficiency, which expands the pool of molecular biomarkers that could enhance our understanding of the molecular mechanisms of HCC.

Development of Rofecoxib-Based Fluorescent Probes and Investigations on Their Solvatochromism, AIE Activity, Mechanochromism, and COX-2-Targeted Bioimaging.

Cyclooxygenase-2 (COX-2) fluorescent probes are promising tools for early diagnosis of cancer. Traditionally, COX-2 probes were designed by connecting two parts, a fluorophore and a COX-2 binding unit, via a flexible linker. Herein, a new class of COX-2-specific fluorescent probes have been developed via one-step modification from rofecoxib by an integrative approach to combine the binding unit and the fluorophore into one. Among them, several new rofecoxib analogues not only exhibited still potent COX-2 binding ability but also exhibited attractive fluorescence properties, such as tunable blue-red emission, solvatochromism, aggression-induced emission behavior, and mechanochromism. Notably, the emission of 2a16 can be switched between green-yellow in the crystalline state and red-orange in the amorphous state by grinding and fuming treatments. Furthermore, the highly fluorescent compound 2a16 (Φf = 0.94 in powder) displayed a much stronger fluorescence imaging of COX-2 in HeLa cancer cells overexpressing COX-2 than RAW264.7 normal cells with a minimal expression of COX-2. Most importantly, 2a16 can light up human cancer tissues from adjacent normal tissues with a much brighter fluorescence by targeting the COX-2 enzyme. These results demonstrated the potential of 2a16 as a new red fluorescent probe for human cancer imaging in clinical applications.

Discovery of novel ID2 antagonists from pharmacophore-based virtual screening as potential therapeutics for glioma.

Glioma, especially the most aggressive type glioblastoma multiforme, is a malignant cancer of the central nervous system with a poor prognosis. Traditional treatments are mainly surgery combined with radiotherapy and chemotherapy, which is still far from satisfactory. Therefore, it is of great clinical significance to find new therapeutic agents. Serving as an inhibitor of differentiation, protein ID2 (inhibitor of DNA binding 2) plays an important role in neurogenesis, neovascularization and malignant development of gliomas. It has been shown that ID2 affects the malignant progression of gliomas through different mechanisms. In this study, a pharmacophore-based virtual screening was carried out and 16 hit compounds were purchased for pharmacological evaluations on their ID2 inhibitory activities. Based on the cytotoxicity of these small-molecule compounds, two compounds were shown to effectively inhibit the viability of glioma cells in the micromolar range. Among them, AK-778-XXMU was chosen for further study due to its better solubility in water. A SPR (Surface Plasma Resonance) assay proved the high affinity between AK-778-XXMU and ID2 protein with the KD value as 129 nM. The plausible binding mode of ID2 was studied by molecular docking and it was found to match AGX51 very well in the same binding site. Subsequently, the cancer-suppressing potency of the compound was characterized both in vitro and in vivo. The data demonstrated that compound AK-778-XXMU is a potent ID2 antagonist which has the potential to be developed as a therapeutic agent against glioma.

Achieving Molecular Fluorescent Conversion from Aggregation-Caused Quenching to Aggregation-Induced Emission by Positional Isomerization.

In this work, we synthesized a pair of positional isomers by attaching a small electron-donating pyrrolidinyl group at ortho- and para-positions of a conjugated core. These isomers exhibited totally different fluorescent properties. PDB2 exhibited obvious aggregation-induced emission properties. In contrast, PDB4 showed the traditional aggregation-caused quenching effect. Their different fluorescent properties were investigated by absorption spectroscopy, fluorescence spectroscopy, density functional theory calculations and single-crystal structural analysis. These results indicated that the substituent position of the pyrrolidinyl groups affects the twisted degree of the isomers, which further induces different molecular packing modes, thus resulting in different fluorescent properties of these two isomers. This molecular design concept provided a new accurate strategy for designing new aggregation-induced emission luminogens.

Profiling of hepatocellular carcinoma neoantigens reveals immune microenvironment and clonal evolution related patterns.

OBJECTIVE: Neoantigens derived from tumor-specific genomic alterations have demonstrated great potential for immunotherapeutic interventions in cancers. However, the comprehensive profile of hepatocellular carcinoma (HCC) neoantigens and their complex interplay with immune microenvironment and tumor evolution have not been fully addressed. METHODS: Here we integrated whole exome sequencing data, transcriptome sequencing data and clinical information of 72 primary HCC patients to characterize the HCC neoantigen profile, and systematically explored its interactions with tumor clonal evolution, driver mutations and immune microenvironments. RESULTS: We observed that higher somatic mutation/neoantigen load was associated with better clinical outcomes and HCC patients could be further divided into two subgroups with distinct prognosis based on their neoantigen expression patterns. HCC subgroup with neoantigen expression probability high (NEP-H) showed more aggressive pathologic features including increased incidence of tumor thrombus (P=0.038), higher recurrence rate (P=0.029), more inclined to lack tumor capsule (P=0.026) and with more microsatellite instability sites (P=0.006). In addition, NEP-H subgroup was also characterized by higher chance to be involved in tumor clonal evolution [odds ratio (OR)=46.7, P<0.001]. Gene set enrichment analysis revealed that upregulation of MYC and its targets could suppress immune responses, leading to elevated neoantigen expression proportion in tumor cells. Furthermore, we discovered an immune escape mechanism that tumors could become more inconspicuous by evolving subclones with less immunogenicity. We observed that smaller clonal mutation clusters with higher immunogenicity in tumor were more likely to involve in clonal evolution. Based on identified neoantigen profiles, we also discovered series of neoantigenic hotspot genes, which could serve as potential actionable targets in future. CONCLUSIONS: Our results revealed the landscape of HCC neoantigens and discovered two clinically relevant subgroups with distinct neoantigen expression patterns, suggesting the neoantigen expression should be fully considered in future immunotherapeutic interventions.

Tissue and serum metabolomic phenotyping for diagnosis and prognosis of hepatocellular carcinoma.

More than two-thirds of patients with hepatocellular carcinoma (HCC) cannot receive curative therapy and have poor survival due to late diagnosis and few prognostic directions. In our study, nontargeted and targeted metabolomics analyses were conducted by liquid chromatography-mass spectrometry to characterize metabolic features of HCC and identify diagnostic and prognostic biomarker candidate incorporating liver tissue and serum metabolites. A total of 552 subjects, including 432 with liver tissue and 120 with serum specimens, were recruited in China. In the discovery cohort, a series of 138 metabolites were identified to discriminate HCC tissues from matched nontumor tissues. Retinol presented with the highest area under the curve (AUC) of 0.991 and associated with Edmondson grade. In the validation cohort, all metabolites in retinol metabolism pathway were examined and the levels of retinol and retinal in tumor tissue and serum decreased in the order of normal to cirrhosis to HCC of Edmondson Grades I to IV. Retinol and retinal levels could also differentiate between HCC and cirrhosis, with AUCs of 0.996 and 0.994, respectively, in tissue and 0.812 and 0.744, respectively, in serum. The AUC of the combined retinol and retinal panel in serum was 0.852. Univariate and multivariate Cox regression identified this panel as an independent predictor for HCC and showed that low expression of retinol and retinal correlated with decreased survival time. In conclusion, the retinol metabolic signature had considerable diagnostic and prognostic value for identifying HCC patients who would benefit from prompt therapy and optimal prognostic direction.

Long-Term Survival Outcomes After Liver Resection for Binodular Hepatocellular Carcinoma: A Multicenter Cohort Study.

BACKGROUND: The long-term prognosis after liver resection for multinodular (≥3 nodules) hepatocellular carcinoma (HCC) is generally considered to be unfavorable. However, the role of liver resection for binodular HCC is less investigated. SUBJECTS, MATERIALS, AND METHODS: From a multicenter database, consecutive patients who underwent curative-intent liver resection for binodular HCC and without macrovascular invasion between 2003 and 2015 were retrospectively reviewed. Patients' clinical variables as well as perioperative and long-term survival outcomes were analyzed. Univariable and multivariable analyses were performed to identify the risk factors associated with overall survival (OS) and recurrence-free survival (RFS) after curative resection. RESULTS: Of 263 enrolled patients, the perioperative 30-day mortality and morbidity rates were 1.5% and 28.5%. The 1-, 3-, and 5-year OS and RFS rates were 81.5%, 52.4%, and 39.1% and 57.1%, 35.8%, and 26.6%, respectively. Multivariable Cox-regression analyses identified preoperative alpha-fetoprotein level >400 µg/L, tumor size with a sum of two nodules >8 cm, tumor size ratio of large/small nodule >1.5 (asymmetrical proportion), unilateral hemiliver distribution of two nodules, distance of ≤3 cm between two nodules, and microvascular invasion in any nodule as independent risk factors associated with decreased OS and RFS. CONCLUSION: Liver resection was safe and feasible in patients with binodular HCC, with acceptable perioperative and long-term outcomes. Sum of two tumor sizes, size ratio and distribution, and distance between two nodules were independent risk factors associated with long-term survival outcomes after surgery. These results may guide clinicians to make individualized surgical decisions and estimate long-term prognosis for these patients. IMPLICATIONS FOR PRACTICE: Liver resection was safe and feasible in patients with binodular hepatocellular carcinoma, with acceptable perioperative and long-term outcomes. The sum of two tumor sizes, the size ratio and distribution of the two nodules, and the distance between two nodules were independent risk factors associated with long-term overall survival and recurrence-free survival after liver resection. The results of this study may guide clinicians to make individualized surgical decisions, estimate long-term prognosis, and plan recurrence surveillance and adjuvant therapy for these patients.

The impact of resection margin and microvascular invasion on long-term prognosis after curative resection of hepatocellular carcinoma: a multi-institutional study.

BACKGROUND: The resection margin (RM) status and microscopic vascular invasion (MVI) are known prognostic factors for hepatocellular carcinoma (HCC). An enhanced understanding of their impact on long-term prognosis is required to improve oncological outcomes. METHODS: Using multi-institutional data, the different impact of the RM status (narrow, <1 cm, or wide, ≥1 cm) and MVI (positive or negative) on overall survival (OS) and recurrence-free survival (RFS) after curative liver resection of solitary HCC without macrovascular invasion was analyzed. RESULTS: In 801 patients, 306 (38%) had a narrow RM and 352 (44%) had positive MVI. The median OS and RFS were 109.8 and 74.8 months in patients with wide RM & negative MVI, 93.5 and 53.1 months with wide RM & positive MVI, 79.2 and 41.6 months with narrow RM & negative MVI, and 69.2 and 37.5 months with narrow RM & positive MVI (both P < 0.01). On multivariable analyses, narrow RM & positive MVI had the highest hazard ratio with reduced OS and RFS (HR 2.96, 95% CI 2.11-4.17, and HR 3.15, 95% CI, 2.09-4.67, respectively). CONCLUSIONS: Concomitant having narrow RM and positive MVI increases the risks of postoperative death and recurrence by about 2-fold in patients with solitary HCC.

Clonal evolution in long-term follow-up patients with hepatocellular carcinoma.

To investigate tumor clonal evolution in hepatocellular carcinoma (HCC), we collected 31 tumor samples,16 peritumor samples and matched PBMCs from 11 long-term follow-up patients with HCC. Whole-exome sequencing was performed to obtain SNVs and CNVs for each sample. An average of 652.2 somatic mutations were identified in each patient and the mean percentage of nonubiquitous tumor mutations was 63.7% (range, 0.7%-100%), reflecting the variety of tumor heterogeneity. Further analysis of clonal evolution was conducted based on mutation clustering results and revealed that different clonal evolution patterns indeed existed in single and multifocal HCC while these patterns were significantly correlated to patients' clinical course. These patterns clearly demonstrated different mechanisms of tumor recurrence. During tumor clonal evolution, potential therapeutic targets also emerged and vanished dynamically. Moreover, mutation analysis revealed that the contribution of mutational signature was correlated with clonal evolution history. Target sequencing of follow-up plasma samples also confirmed that ctDNA level could dynamically reflect tumor clonal/subclonal burden. By investigating clonal evolution in HCC patients, our analysis revealed that different patterns indeed existed during HCC progression and proposed a novel strategy for identifying the origin of recurrent tumor as well as optimizing treatment selection.

CDK13 RNA Over-Editing Mediated by ADAR1 Associates with Poor Prognosis of Hepatocellular Carcinoma Patients.

BACKGROUND/AIMS: Aberrant RNA editing, mediated by adenosine deaminases acting on RNA (ADAR), serves as a post-transcriptional event participating in tumorigenesis and prognosis. However, the RNA editing profiles during HCC progression and their clinical correlations remain unclear. METHODS: Multiple tissue samples were collected from an advanced HCC patient. RNA-seq was performed to obtain the RNA editing profiles for each sample. Two RNA editing sites from CDK13 were further validated in 60 HCC patients; and their potential regulatory mechanisms were investigated. RESULTS: In-depth analysis of the RNA-seq data revealed a significant number of editing sites (632-816) in coding regions for each tissue sample, showing branched evolution during tumorigenesis and metastasis. Two editing sites (Q103R and K96R) in CDK13 showed significant over-editing in tumor, and these phenomenon were validated in 60 HCC patients. Furthermore, the clinicopathological analysis revealed that these CDK13 over-editing sites were positively associated with TNM, PVTT and poor prognosis. In addition, the editing level of these sites were significantly correlated with the expression of ADAR1. Loss of function assays further proved that these CDK13 over-editing sites were mediated by ADAR1 in HCC cells. CONCLUSIONS: CDK13 RNA over-editing sites mediated by ADAR1 may serve as novel cancer driver events in HCC progression.

High Expressions of CUL4A and TP53 in Colorectal Cancer Predict Poor Survival.

BACKGROUND/AIMS: Cullin 4A (CUL4A) is vital in cell survival, development, growth and cell cycle, it plays an important role in chaperone-mediated ubiquitination and interacts with TP53 in carcinogenesis. However, the clinicopathologic significance of CUL4A expression in colorectal cancer is unknown; in particular, the prognostic value of CUL4A combined with TP53 expression has not been explored. METHODS: We analyzed the expression of CUL4A in both public database (Oncomine) and 180 cases of colorectal cancer and paired normal tissues by real-time polymerase chain reaction and western blotting. Colony formation, wound healing, migration and invasion assays and tumorigenesis in nude mice were used to explore the function of CUL4A in CRC proliferation and metastasis in vitro and in vivo. Markers of epithelial to mesenchymal transition (EMT) were evaluated by western blotting. Immunohistochemistry (IHC) was used to analyse the relationship between CUL4A expression and E-cadherin expression. RESULTS: CUL4A and TP53 protein expression was significantly higher in cancerous tissues compared to normal tissues. Significant correlation between CUL4A and TP53 expression was observed. CUL4A expression was an independent prognostic factor for overall survival (OS) and disease-free survival (DFS). Interestingly, patients with tumors that had both CUL4A overexpression and mutant TP53 protein accumulation relapsed and died within a significantly short period after surgery (P < 0.001). Multivariate analysis showed that patients with both CUL4A+ and TP53+ positive tumors had extremely poor OS and DFS. Knockdown of CUL4A by a short interfering RNA (siRNA) significantly suppressed the progression of EMT, proliferation, migration, and invasion of colon cancer cells in vitro and tumor growth in vivo. ZEB1 silencing blocked CUL4A-driven these processes. CONCLUSION: CUL4A expression correlated positively with the prognosis of colorectal cancer. Mechanistically, ZEB1 was confirmed to mediate the function of CUL4A in regulating the EMT. The assessment of both CUL4A and mutant TP53 expression will be helpful in predicting colon cancer prognosis.

Protein induced by vitamin K absence or antagonist-II versus alpha-fetoprotein in the diagnosis of hepatocellular carcinoma: A systematic review with meta-analysis.

BACKGROUND: As a promising biomarker of hepatocellular carcinoma (HCC), protein induced by vitamin K absence or antagonist-II (PIVKA-II) has been studied extensively. However, its diagnostic capability varies across HCC studies. This study aimed to compare the performance of PIVKA-II with alpha-fetoprotein (AFP) in the diagnosis of HCC. DATA SOURCES: A systematic literature search was conducted to identify the studies from MEDLINE, Embase and Cochrane Library Databases, which were published up to December 20, 2017 to compare the diagnostic capability of PIVKA-II and AFP for HCC. The data were pooled using random effects model. Pooled sensitivity and specificity were calculated. Summary receiver operating characteristic curve (ROC) was employed to evaluate the diagnostic accuracy of each marker. RESULTS: Thirty-one studies were included. The pooled sensitivity (95% CI) of PIVKA-II and AFP was 0.66 (0.65-0.68) and 0.66 (0.65-0.67), respectively in diagnosis of HCC; and the corresponding pooled specificity (95% CI) was 0.89 (0.88-0.90) and 0.84 (0.83-0.85), respectively. The area under the ROC curve (AUC) of PIVKA-II and AFP was 0.856 (0.817-0.895) and 0.770 (0.728-0.811), respectively. Subgroup analysis showed that PIVKA-II was superior to AFP in terms of the AUC for both small HCC (< 3 cm) [0.863 (0.825-0.901) vs 0.717 (0.658-0.776)] and large HCC (≥ 3 cm) [0.854 (0.811-0.897) vs 0.729 (0.682-0.776)]; for American [0.926 (0.897-0.955) vs 0.698 (0.594-0.662)], European [0.772 (0.743-0.801) vs 0.628 (0.594-0.662)], Asian [0.838 (0.812-0.864) vs 0.785 (0.764-0.806)] and African [0.812 (0.794-0.840) vs 0.721 (0.675-0.767)] HCC patients; and for HBV-related [0.909 (0.866-0.951) vs 0.714 (0.673-0.755)] and mixed-etiology [0.847 (0.821-0.873) vs 0.794 (0.772-0.816)] HCC. CONCLUSION: This meta-analysis indicates that PIVKA-II is better than AFP in terms of the accuracy for diagnosing HCC, regardless of tumor size, patient ethnic group, or HCC etiology.

Deletions at SLC18A1 increased the risk of CRC and lower SLC18A1 expression associated with poor CRC outcome.

Copy number variations (CNVs) contribute to the development of colorectal cancer (CRC). We conducted a two-stage association study to identify CNV risk loci for CRC. We performed a gene-based rare CNV study on 694 sporadic CRC and 1641 controls using Illumina Human-OmniExpress-12v1.0 BeadChips, and further replicated in 934 CRC cases and 2680 controls for risk CNVs by using TaqMan Copy Number Assay. Tumor buddings, cancer cells in the center of primary tumor and normal intestinal epithelial cells were captured using laser capture microdissection (LCM) and were assayed using AffymetrixGeneChip® Human Genome U133 Plus 2.0 Array. In addition, The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus data were assessed for the effects of risk CNVs. We found that germline deletions affecting the last six exons of SLC18A1 significantly associated with CRC with a combined P value of 6.4 × 10-5 by a two-stage analysis. Both in TCGA CRC RNA seq dataset and GDS4382, SLC18A1 was significantly down regulated in CRC tissues than in paired normal tissues (N = 32 and 17 pairs, P = 0.004 and 0.009, respectively). In LCM samples, similar observations were obtained that the expression levels of SLC18A1 in the tumor buddings, cancer cells in the center of primary tumor, and stroma of both tumor budding and cancer cells were lower than normal intestinal epithelial and stromal cells (fold change = 0.17-0.62, 0.12-0.57 and 0.37-0.68, respectively). In summary, the germline deletions at SLC18A1 contributed to the development of CRC. The role of SLC18A1 required further exploration.