Hepatocellular carcinoma (HCC) is one of the most deadly cancers worldwide and has no effective treatment, yet the molecular basis of hepatocarcinogenesis remains largely unknown. Here we report ...findings from a whole-genome sequencing (WGS) study of 88 matched HCC tumor/normal pairs, 81 of which are Hepatitis B virus (HBV) positive, seeking to identify genetically altered genes and pathways implicated in HBV-associated HCC. We find beta-catenin to be the most frequently mutated oncogene (15.9%) and TP53 the most frequently mutated tumor suppressor (35.2%). The Wnt/beta-catenin and JAK/STAT pathways, altered in 62.5% and 45.5% of cases, respectively, are likely to act as two major oncogenic drivers in HCC. This study also identifies several prevalent and potentially actionable mutations, including activating mutations of Janus kinase 1 (JAK1), in 9.1% of patients and provides a path toward therapeutic intervention of the disease.
Abstract
Chronic hypoxia is a common inducer of end-stage cardiovascular disease. In cells under hypoxia, the hypoxia-inducible factor-1 (HIF-1) plays a vital role in regulating downstream target ...genes. However, the mechanism of hypoxia in cardiomyocytes is still unclear. In this study, we aimed to identify novel downstream epigenetic targets of HIF-1α in cardiomyocytes under hypoxia. H9c2 cells were exposed to hypoxia condition, and quantitative real-time PCR analysis was performed to evaluate the expression of miR-20b-5p. The results indicated that the expression of miR-20b-5p was down-regulated in H9c2 cells under low oxygen condition. Meanwhile, HIF-1α overexpression further down-regulated the miR-20b-5p expression in H9c2 cells transfected with HIF-1α plasmids. In addition, Annexin-V-FITC/PI flow cytometry analysis suggested that overexpression of miR-20b-5p attenuated cell apoptosis under hypoxia condition in H9c2 cells. Western blot analysis showed that the hypoxia apparently increased Bax and cleaved-caspase-3, but decreased Bcl-2 expression in H9c2 cells, indicating that hypoxia-induced NF-κB signaling pathway activation is mediated by miR-20b-5p. Hypoxia-induced H9c2 cell apoptosis was reduced after HIF-1α knockdown as shown by the flow cytometry analysis. In conclusion, we identified that miR-20b-5p plays an important role in mediating cardiomyocytes apoptosis under hypoxia, which is mediated by the HIF-1/NF-κB signaling pathway.
Objectives
The mechanisms underlying the effects of Toll‐like receptor 9 (TLR9) and autophagy on rheumatoid arthritis (RA)‐aggravated periodontitis are unclear. We aimed to explore a novel target, ...cathepsin K (Ctsk)‐mediated TLR9‐related autophagy, during the progress of periodontitis with RA.
Materials and Methods
DBA/J1 mouse model of periodontitis with RA was created by local colonization of Porphyromonas gingivalis (Pg) and injection of collagen. The expression of Ctsk was inhibited by adeno‐associated virus (AAV). Micro‐CT, immunohistochemistry (IHC), Western blot and quantitative real‐time polymerase chain reaction (qRT‐PCR) were used to detect the expression of TLR9‐related autophagy in periodontitis with RA. Small interfering RNA (siRNA) and CpG oligodeoxynucleotides (CpG ODN) were applied in macrophages. Western blot, immunofluorescence (IF) and qRT‐PCR were used to verify the in vivo results.
Results
RA can promote periodontitis bone destruction in the lesion area, while inhibiting Ctsk could effectively alleviate this effect. The infiltration of macrophages, TLR9, autophagy proteins (TFEB and LC3) and inflammatory cytokines increased in the periodontitis‐with‐RA group and was reduced by the inhibition of Ctsk in the periodontal region. Macrophage stimulation confirmed the in vivo results. With the activation of TLR9 by CpG ODN, inhibition of Ctsk could suppress both TLR9 downstream signalling proteins and autophagy‐related proteins.
Conclusions
This study advanced a novel role for Ctsk in TLR9 and autophagy to explain the interaction between periodontitis and RA.
An unbiased scanning methodology using ultra high-performance liquid chromatography coupled with high-resolution mass spectrometry was used to bank data and plasma samples for comparing the data ...generated at different dates. This method was applied to bank the data generated earlier in animal samples and then to compare the exposure to metabolites in animal versus human for safety assessment. With neither authentic standards nor prior knowledge of the identities and structures of metabolites, full scans for precursor ions and all ion fragments (AIF) were employed with a generic gradient LC method to analyze plasma samples at positive and negative polarity, respectively. In a total of 22 tested drugs and metabolites, 21 analytes were detected using this unbiased scanning method except that naproxen was not detected due to low sensitivity at negative polarity and interference at positive polarity; and 4′- or 5-hydroxy diclofenac was not separated by a generic UPLC method. Statistical analysis of the peak area ratios of the analytes versus the internal standard in five repetitive analyses over approximately 1 year demonstrated that the analysis variation was significantly different from sample instability. The confidence limits for comparing the exposure using peak area ratio of metabolites in animal plasma versus human plasma measured over approximately 1 year apart were comparable to the analysis undertaken side by side on the same days. These statistical analysis results showed it was feasible to compare data generated at different dates with neither authentic standards nor prior knowledge of the analytes.
Abstract
Background: PF-06647020, an ADC comprising a humanized monoclonal antibody against PTK7, a cleavable valine-citrulline linker, and an auristatin payload, is being investigated in an ongoing ...Phase I clinical trial in pts with advanced solid tumors. We hypothesized that response to a PTK7-directed ADC would correlate with PTK7 expression in the pts tumor. We report preliminary baseline PTK7 expression in pts tumors and clinical response using a novel CLIA validated laboratory developed test (LDT).
Methods: We developed and validated an LDT comprising an anti-PTK7 immunohistochemistry assay with digital tissue analysis using Flagship Biosciences Inc’s cTA® platform to detect membrane-associated PTK7 in formalin-fixed paraffin embedded (FFPE) epithelial malignancies of ovary, breast and lung. Tumor samples were immunolabeled with the LDT and image analysis was applied to derive a digital H-score. Specifically the digital image annotation included desired regions of analysis (ROA) and excluded regions not of interest. The cTA® platform identified each cell in a given ROA and quantified an overall H-score for malignant epithelial cell membranes immunolabeled for PTK7. Non-small cell lung cancer (NSCLC) pts with H-scores ≥ 56.8 were eligible for enrollment whereas ovarian cancer (OVCA) pts were enrolled unselected. Triple negative breast cancer (TNBC) pts were initially enrolled unselected, but after review a requirement for H-scores ≥ 92.8 was implemented for enrollment.
Results: PTK7 IHC was performed on baseline FFPE samples from 69 pts and digital H-scores were generated (Table 1). An additional 7 pt samples were not amenable to analysis with the cTA® platform.
Conclusions: A novel CLIA validated LDT was developed and implemented to assess PTK7 baseline levels in FFPE tumors from pts treated with PF-06647020. Clinical responses tended to correlate with H-scores that were higher than the mean for each tumor type.
Table 1.PTK7 digital H-scores in Phase I dose expansionNumber of SamplesMinimum H-scoreMaximum H-scoreMean H-scoreMean H-score of Non-respondersMean H-score of RespondersOVCA31120089.280.2111.2NSCLC1993287167.8159.2200.3TNBC1953258159.2147.0224.3
Citation Format: Amy Jackson-Fisher, Navi Mehra, Roberto Gianani, Pamela Whalen, Pamela Vizcarra, Shibing Deng, Stephen McComish, Xiaohua Xin, Eric L. Powell. Protein tyrosine kinase 7 (PTK7) biomarker analysis in patients (pts) treated with PF-06647020, a PTK7 antibody-drug conjugate (ADC), in a phase I dose expansion study abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4035.
Abstract Carrier dynamics detection in different dimensions (space, time, and energy) with high resolutions plays a pivotal role in the development of modern semiconductor devices, especially in ...low‐dimensional, high‐speed, and ultrasensitive devices. Here, a femtosecond electron‐based versatile microscopy is reported that combines scanning ultrafast electron microscopy (SUEM) imaging and time‐resolved cathodoluminescence (TRCL) detection, which allows for visualizing and decoupling different dynamic processes of carriers involved in surface and bulk in semiconductors with unprecedented spatiotemporal and energetic resolutions. The achieved spatial resolution is better than 10 nm, and the temporal resolutions for SUEM imaging and TRCL detection are ≈500 fs and ≈4.5 ps, respectively, representing state‐of‐the‐art performance. To demonstrate its unique capability, the surface and bulk carrier dynamics involved in n‐type gallium arsenide (GaAs) are directly tracked and distinguished. It is revealed, in real time and space, that hot carrier cooling, defect trapping, and interband‐/defect‐assisted radiative recombination in the energy domain result in ordinal super‐diffusion, localization, and sub‐diffusion of carriers at the surface, elucidating the crucial role of surface states on carrier dynamics. The study not only gives a comprehensive physical picture of carrier dynamics in GaAs, but also provides a powerful platform for exploring complex carrier dynamics in semiconductors for promoting their device performance.
Previous studies on the influence of a rural/urban setting on the prevalence of cardiovascular disease risk factors in children have not sufficiently controlled for socioeconomic status, race, ...gender, and perhaps, may not have included a representative sample of rural and urban children. This study compared the cardiovascular disease risk factors and rate of obesity of children living in rural and urban settings. It also determined the magnitude of the effect of the rural/urban setting on cardiovascular disease risk factors and obesity when controlling for race, socioeconomic status, and gender. The subjects were 2,113 third- and fourth-grade children; 962 from an urban setting and 1,151 from a rural setting. Height, weight, skinfolds, resting blood pressure, and total cholesterol levels were measured. Aerobic power (pVO2max) was estimated from cycle ergometry. Physical activity and smoking history were obtained from a questionnaire. Clustering analyses using adjustment for sample error indicated that total cholesterol, blood pressure, smoking, and physical activity levels of rural and urban children were not different (P>0.10); however, body mass index and sum of skinfolds was greater for rural youth (P<0.004). Logistic regression indicated that rural children had a 54.7 percent increased risk of obesity (P=0.0001). This study's results indicate that, in children, a rural setting is associated with obesity, but not with the major risk factors associated with cardiovascular disease.
Abstract
Background Intrinsic breast cancer molecular subtyping (IBCMS) provides significant prognostic information for patients (pts) with breast cancer (BC) treated with chemotherapy, human ...epidermal growth factor receptor 2 (HER2) targeted therapies, and endocrine therapies (ETs). Classifying tumors into intrinsic subtypes to determine optimal treatment is often applied using PAM50, commercially known as Prosigna. Meanwhile, Absolute Assignment of Breast Cancer Intrinsic Molecular Subtypes (AIMS) computational method was trained to predict PAM50-based IBCMS. As the PAM50 algorithm was developed to capture the major subtypes in a general pt population, clinicopathologic distribution of the study cohort and technology platform calibration should be considered in IBCMS analyses. This study compared different next-generation sequencing technologies and methodologic approaches of PAM50 on tumor samples from 2 randomized trials of postmenopausal women with estrogen receptor-positive (ER+)/HER2-negative (HER2-) BC. Methods PALOMA-2 is a double-blind, randomized study of first-line palbociclib (PAL) + letrozole (LET) for ER+/HER2- advanced BC (ABC). Tumor samples from consented pts were subtyped using the validated RUO PAM50 assay (ruoProsigna, NanoString); results were compared with published subtype results using AIMS on EdgeSeq Oncology Biomarker Panel (HTG Molecular Diagnostics). PALLET is a phase 2, randomized trial of PAL+LET as neoadjuvant therapy in pts with ER+ HER2- BC. Baseline frozen tumor biopsies underwent whole transcriptome mRNA-sequencing (mRNA-seq). IBCMS was performed using AIMS; PAM50 subtyping was performed on data normalised with subgroup-specific gene centering and microarray-RNA-sequencing calibration. Results In PALOMA-2, 222 pts had both ruoProsigna and AIMS data; an overall 54% agreement rate between methods was observed, with 46% (56/121) of Luminal B (LumB) subtype by ruoProsigna assigned as Luminal A (LumA) by AIMS and 67% (6/9) of basal-like by ruoProsigna as HER2-enriched (HER2-E) by AIMS (Table 1). In PALLET, 224 pts had mRNA-seq data; a 69% agreement between the two approaches (AIMS and PAM50) was observed, with only 4% (2/49) of LumB assigned as LumA by AIMS but 17% (26/156) and 16% (25/156) of LumA considered LumB or normal-like by AIMS, respectively. Progression-free survival (PFS) by ruoProsigna-derived subtype in PALOMA-2 showed that PAL+LET benefited all pts but those with a basal-like subtype (Table 2). With AIMS, PAL+LET provided a PFS benefit in pts with LumA and LumB subtypes, but was less effective in the HER2-E subtype. Conclusion Intrinsic subtyping has potential clinical utility. PAL+ET should be considered for ER+/HER2- ABC, except possibly in pts with a basal-like tumor, consistent with previous reports. A standardized clinical PAM50 assay and bioinformatics approach should be used as discrepancies in gene expression platforms and algorithms lead to different results and could misguide treatment decisions. Clinical trial identification: Pfizer (NCT01740427)
Table 1.Intrinsic Subtyping by IBCMS MethodsPALOMA-2PALLETMethodruoProsignaPAM50 mRNAseqAIMS BasalHER2LumALumBGrand TotalBasalHER2LumALumBNormalGrand TotalBasal-like, n (%)1 (11)NANANA13 (75)0001 (8)4HER2-E, n (%)6 (67)6 (30)6 (8)13 (10)311 (25)3 (100)8 (5)6 (12)1 (8)19LumA, n (%)NA2 (10)60 (83)56 (46)1180097 (62)2 (4)099LumB, n (%)2 (22)12 (60)3 (4)52 (43)690026 (17)41 (84)067Normal-like, n (%)NANA3 (4)NA30025 (16)010 (83)35Grand Total9 (100)20 (100)72 (100)121 (100)2224 (100)3 (100)156 (100)49 (100)12 (100)224NA=Not available
Table 2.Median PFS statistics by subtype in PALOMA-2PAL+LET PFS, monthsPBO+LET PFS, monthsHazard Ratio(95% CI)P ValueruoProsignaBasal-like8.2 (n=5)3.6 (n=4)0.39 (0.09-1.77)0.206HER2-E11.0 (n=12)5.1 (n=8)0.41 (0.15-1.11)0.071LumA37.2 (n=52)13.6 (n=20)0.42 (0.21-0.84)0.011LumB27.6 (n=79)13.8 (n=42)0.63 (0.40-1.00)0.049AIMSBasal-likeNANANANAHER2-E16.4 (n=21)8.4 (n=10)0.82 (0.32-2.1)0.684LumA30.6 (n=84)16.5 (n=34)0.56 (0.33-0.95)0.029LumB19.3 (n=41)8.8 (n=28)0.39 (0.23-0.67)<0.001NA=Not available; PBO=placebo
Citation Format: Maggie Cheang, Mitch Dowsett, Mothaffar Rimawi, Stephen Johnston, Samuel Jacobs, Judith Bliss, Katherine Pogue-Geile, Lucy Kilburn, Zhou Zhu, Eugene F. Schuster, Hui Xiao, Lisa Swaim, Shibing Deng, Dongrui R. Lu, Eric Gauthier, Jennifer Tursi, Dennis J. Slamon, Hope S. Rugo, Richard S. Finn, Yuan Liu. Impact of using cross-platform gene expression profiling technologies and computational methods for intrinsic breast cancer subtyping in PALOMA-2 and PALLET abstract. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PD2-07.
Abstract
One of the main challenges in immuno-oncology is to quantify different types of immune cells in the tumor microenvironment, as such data can greatly facilitate our understanding of the ...mechanism of action of, and response to cancer immunotherapies. Traditionally immune profiling is performed by immunohistochemistry or flow cytometry experiments using surface markers specific to each immune cell type. However, both approaches suffer from practical limitations such as reagent availability and real world specimen conditions. Recently several computational deconvolution approaches using expression profiles of the bulk tumor tissue samples have been developed. This alternative approach utilizes existing database of purified immune cell gene signatures and relies on a rigorous mathematical framework of signal deconvolution. One of such algorithms, called CIBERSORT (Newman et al. 2015), has been demonstrated to perform robustly in deconvoluting the relative fractions of 22 human leukocyte subsets in solid tumor tissue samples, and benchmarked favorably against FACS analysis.
In this work, we performed immune phenotyping by the CIBERSORT-based expression deconvolution on a cohort of 100 gastric cancer (GC) patients. We found that the fractions of activated CD4 memory T cells, rather than CD8+ T cell, are most significantly correlated with tumor neo-antigen load, whereas the latter is most significantly associated with the expression of a “Cytolytic Activity” metagene and an interferon gamma signature. There is stronger pre-existing host immune response against the MSI and EBV subtypes of GCs mediated by either cytotoxic T cells or Natural Killer cells. Lastly, we found a high M2/M1 tumor associated Macrophages (TAMs) ratio is strongly associated with poor GC prognosis, corroborating recent reports on the role of a TAM and stromal response in gastric cancer angiogenesis.
Taken together, we demonstrated in this work that critical information about the tumor infiltrating immune cells can be gleaned from the computational deconvolution of bulk tumor gene expression profiling data. The comprehensiveness and cost-effectiveness of this approach can complement other immune profiling techniques in characterizing a wide variety of tumor specimens under various treatment conditions. We foresee its application in characterizing pharmacodynamics, understanding immunological mechanism of action and monitoring treatment response and disease progression for cancer immunotherapies and their combinations in both pre-clinical and clinical settings.
Citation Format: Kai Wang, Hoicheong Siu, Shibing Deng, Jadwiga R. Bienkowska, Suet Yi Leung. Characterizing the immunogenicity of gastric cancer by transcriptomic expression based immune phenotyping. abstract. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4143.
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