scholarly journals Integrative Proteomic Characterization of Trace FFPE Samples in Early-Stage Gastrointestinal Cancer

2021 ◽  
Author(s):  
Lingling Li ◽  
Hui Liu ◽  
Yan Li ◽  
Chunmei Guo ◽  
Bing Wang ◽  
...  
Keyword(s):  
Gastrointestinal Cancer ◽  
Immune Cell ◽  
Early Stage ◽  
Tissue Samples ◽  
Early Stage Cancer ◽  
Relationship Analysis ◽  
Ffpe Samples ◽  
Single Trace ◽  
Distinct Separation

Abstract Background The surveillance and therapy of early-stage cancer would be better for patients’ prognosis. However, the extreme trace amount of tissue samples in different stages have limited in portraying the characterization of early-stage cancer. Therefore, we focused on and presented comprehensive proteomic and phosphoproproteomic profiling of the trace FFPE samples from early-stage gastrointestinal cancer, and then explored the potential biomarkers of early-stage gastrointestinal cancer. Methods In this study, a quantitative proteomic method with chromatography with mass spectrometry (LC-MS/MS) was used to analyse the proteomic difference between the trace early-stage esophageal squamous cell carcinoma (EESCC) and early-stage duodenum adenocarcinoma cancer (EDAC). Results We identified ~6,000 proteins and >10,000 phosphosites in single trace FFPE samples. The distinct separation of EESCC and EDAC illustrated the functions of cell cycle (RB1 T373, EGFR T693) in EESCC, and the positive impacts of apoptosis, metabolic processes (MTOR and MTOR S1261) in EDAC. Furthermore, we deconvoluted the immune infiltration of early-stage gastrointestinal cancer, in which higher immune cell signatures were detected in EDAC, and showed the specific cytokines in EESCC and EDAC. We performed kinases-substates relationship analysis and elucidated the specific proteomic kinase characterization of EESCC and EDAC, and proposed the medicative effects and corresponding drugs for EESCC and EDAC at the clinic.Conclusion We disclosed the specific immune characterization of the early-stage gastrointestinal cancer, and presented potential makers of EESCC (EGFR, PDGFRB, CDK4, WEE1) and EDAC (MTOR, MAP2K1, MAPK3). This study represents a major stepping stone towards investigating the carcinogenesis mechanism of gastrointestinal cancer, and providing a rich resource for medicative strategy in the clinic.

scholarly journals De novo characterization of cell-free DNA fragmentation hotspots boosts the power for early detection and localization of multi-cancer

2020 ◽  
Author(s):  
Xionghui Zhou ◽  
Yaping Liu
Keyword(s):  
Dna Fragmentation ◽  
De Novo ◽  
Early Stage ◽  
Fine Scale ◽  
Cell Free Dna ◽  
Early Stage Cancer ◽  
Free Dna ◽  
Cancer Types ◽  
Fragmentation Patterns

AbstractThe global variation of cell-free DNA fragmentation patterns is a promising biomarker for cancer diagnosis. However, the characterization of its hotspots and aberrations in early-stage cancer at the fine-scale is still poorly understood. Here, we developed an approach to de novo characterize genome-wide cell-free DNA fragmentation hotspots by integrating both fragment coverage and size from whole-genome sequencing. These hotspots are highly enriched in regulatory elements, such as promoters, and hematopoietic-specific enhancers. Surprisingly, half of the high-confident hotspots are still largely protected by the nucleosome and located near repeats, named inaccessible hotspots, which suggests the unknown origin of cell-free DNA fragmentation. In early-stage cancer, we observed the increases of fragmentation level at these inaccessible hotspots from microsatellite repeats and the decreases of fragmentation level at accessible hotspots near promoter regions, mostly with the silenced biological processes from peripheral immune cells and enriched in CTCF insulators. We identified the fragmentation hotspots from 298 cancer samples across 8 different cancer types (92% in stage I to III), 103 benign samples, and 247 healthy samples. The fine-scale fragmentation level at most variable hotspots showed cancer-specific fragmentation patterns across multiple cancer types and non-cancer controls. Therefore, with the fine-scale fragmentation signals alone in a machine learning model, we achieved 42% to 93% sensitivity at 100% specificity in different early-stage cancer. In cancer positive cases, we further localized cancer to a small number of anatomic sites with a median of 85% accuracy. The results here highlight the significance to characterize the fine-scale cell-free DNA fragmentation hotspot as a novel molecular marker for the screening of early-stage cancer that requires both high sensitivity and ultra-high specificity.

Is microsatellite instability a negative prognostic factor in early stage endometrial cancer patients?

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. e17107-e17107
Author(s):  
Sahin Lacin ◽  
Gozde Elif Tasar ◽  
Alp Usubutun ◽  
Zafer Arik ◽  
Yusuf Karakas ◽  
...  
Keyword(s):  
Endometrial Cancer ◽  
Microsatellite Instability ◽  
Immune Cell ◽  
Early Stage ◽  
Menopausal Status ◽  
Survival Rates ◽  
Disease Stage ◽  
Biological Behaviour ◽  
Tissue Samples ◽  
Early Stage Disease

e17107 Background: Endometrial cancer(EC) is a heterogeneous disease with diverse histological features and biological behaviour. Microsatellite instability (MSI) which occurs in cancerous tissue secondary to mismatch repair (MMR) defect of hereditary or somatic origin is a well-known feature further diversifying genetic tumor landscape. In this study, we aim to investigate the relationship between MSI and prognosis in patients with early stage EC. Methods: The patients diagnosed with EC between 2004 and 2017 were retrospectively analysed in the study. The demographic characteristics, disease stage, menopausal status, and clinical and laboratory values were noted. MLH1, MSH2, MSH6 and PMS2 antibodies were used to detect microsatellite status within the tumor tissue samples. Results: The mean age of the 93 patients and menopause was 60.6 ± 9.8 and 49.68 ± 4.5 years , respectively. The median follow-up period was 28 (1-110) months. At the time of diagnosis, number of the patients with stage IA, IB, II, III, and IV were 5, 64, 8, 14, and 2, respectively. Forty three patients were grad 1, 23 patients grad 2, and 27 patients were grade 3. In terms of microsatellite status, 59 patients (63.4%) were microsatellite stable (MSS) and 34 patients (36.6%) were microsatellite instable(MSI). There was no significant association between MSI and tumor stage or grade (X2 = 1.97 p = 0.74 and X2= 3.2, p = 0.19, respectively). There was a significant relationship between peritumoral lymphocyte infiltration rate (pTIL) and MSI. PTIL ratio was higher in tumoral tissues with MSI, whereas pTIL was low in MSS tumor tissues (X2= 28.6, p < 0.0001). Patients with MSI tended to have poorer survival than patients with MSS irrespective of disease stage; mOS rates were 76 and 91 months, respectively (p = 0.086). However, patients with early stage disease and MSI had significantly poorer survial in comparison to patients with similar disease stage and MSS: Overall survival rates for patients with MSI and MSS were 75.8 and 94.7 months, respectively (Log-rank p = 0.048). Conclusions: There is a limited number of studies assessing the association between MSI and clinical outcomes in EC. Our study hints at the presence of potential relationship between MSI and pognosis in patients with MSI. Similar to previous studies performed with various types of tumors we found that EC with MSI attract more immune cells to tumor microenvironments. Interestingly, patients with MSI tended to have poorer prognosis despite augmented immune-cell inftration.

A new approach for immuno-oncology biomarker discovery: High-plex, spatial protein profiling based on NanoString digital quantification.

2017 ◽  
Vol 35 (7_suppl) ◽  
pp. 27-27 ◽  
Author(s):  
David Lee ◽  
Yan Liang ◽  
Chris Merritt ◽  
Fiona Pakiam ◽  
Giang Ong ◽  
...  
Keyword(s):  
Immune Response ◽  
Single Cell ◽  
Spatial Resolution ◽  
Biomarker Discovery ◽  
Immune Cell ◽  
Light Exposure ◽  
Protein Detection ◽  
Protein Profiling ◽  
Ffpe Samples

27 Background: The immune response to cancer is shaped by the abundance and localization of immune cell populations, their activation status, and expression of immunomodulatory factors. To detect proteins at high multiplex with spatial resolution, NanoString optical barcoding technology was used to digitally profile protein expression in formalin fixed paraffin embedded (FFPE) samples and compared to traditional IHC. Methods: NanoString has enabled digital profiling of immuno-oncology protein targets, including immune cell markers and checkpoint proteins. Protein detection is enabled via primary antibodies (Abs) which are attached via a UV cleavable linker to DNA indexing oligos. FFPE samples are stained with a multiplex cocktail of labeled Abs, then DNA oligos are released by UV light exposure. Liberated oligos are then hybridized to optical barcodes for quantitation on a NanoString instrument. This technique enables quantitative, multiplex protein detection over 5 logs of dynamic range. IHC and NanoString spatial protein profiling were performed on alternating sections from blocks of tonsil, melanoma, and colorectal cancer. Sections were fluorescently labeled with panCK, Ki67, and Syto-83 to visualize morphology, and regions of interest (ROI) were selected for profiling using up to 30 oligo-tagged Abs. Results: NanoString counts strongly correlated with quantification of CD3, CD4, CD8, PanCK, Ki67, PD-1, and PD-L1 derived from IHC. To explore limits of detection, ROIs of 1, 2, 4, or 8 cells were profiled. Digital counts were detected above background at the single cell level and linearly correlated with cell count. Furthermore, quantification with two cell lineage markers and double positive cells were demonstrated using a 30-plex Ab cocktail. Conclusions: Multiplex, digital protein profiling with spatial resolution will enable deep characterization of immune responses in tumors. Additionally, single cell profiling enables inter-cellular characterization of variations in immune response. The strong correlation of NanoString data to IHC indicates the feasibility to spatially profile multiple key proteins with minimal consumption of patient tissue.

scholarly journals Reclassification of Kidney Clear Cell Carcinoma Based on Immune Cell Gene-Related DNA CpG Pairs

Biomedicines ◽  
2021 ◽  
Vol 9 (2) ◽  
pp. 215
Author(s):  
Qizhan Luo ◽  
Thomas-Alexander Vögeli
Keyword(s):  
Cell Carcinoma ◽  
Immune Cell ◽  
Clear Cell ◽  
Clear Cell Carcinoma ◽  
Pairwise Comparison ◽  
Clinicopathological Features ◽  
Immune Checkpoints ◽  
Potential Mechanism ◽  
Tissue Samples

Background: A new method was developed based on the relative ranking of gene expression level, overcoming the flaw of the batch effect, and having reliable results in various studies. In the current study, we defined the two methylation sites as a pair. The methylation level in a specific sample was subject to pairwise comparison to calculate a score for each CpGs-pair. The score was defined as a CpGs-pair score. If the first immune-related CpG value was higher than the second one in a specific CpGs-pair, the output score of this immune-related CpGs-pair was 1; otherwise, the output score was 0. This study aimed to construct a new classification of Kidney Clear Cell Carcinoma (KIRC) based on DNA CpGs (methylation sites) pairs. Methods: In this study, the biomarkers of 28 kinds of immune infiltration cells and corresponding methylation sites were acquired. The methylation data were compared between KIRC and normal tissue samples, and differentially methylated sites (DMSs) were obtained. Then, DNA CpGs-pairs were obtained according to the pairs of DMSs. In total, 441 DNA CpGs-pairs were utilized to construct a classification using unsupervised clustering analysis. We also analyzed the potential mechanism and therapy of different subtypes, and validated them in a testing set. Results: The classification of KIRC contained three subgroups. The clinicopathological features were different across three subgroups. The distribution of immune cells, immune checkpoints and immune-related mechanisms were significantly different across the three clusters. The mutation and copy number variation (CNV) were also different. The clinicopathological features and potential mechanism in the testing dataset were consistent with those in the training set. Conclusions: Our findings provide a new accurate and stable classification for developing personalized treatments for the new specific subtypes.

scholarly journals MBRS-46. CHARTING NEOPLASTIC AND IMMUNE CELL HETEROGENEITY IN HUMAN AND GEM MODELS OF MEDULLOBLASTOMA USING scRNAseq

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii406-iii406
Author(s):  
Andrew Donson ◽  
Kent Riemondy ◽  
Sujatha Venkataraman ◽  
Ahmed Gilani ◽  
Bridget Sanford ◽  
...  
Keyword(s):  
Single Cell ◽  
Rna Sequencing ◽  
Immune Cell ◽  
Genetically Engineered ◽  
Cellular Heterogeneity ◽  
Cell Heterogeneity ◽  
Transcriptomic Data ◽  
Single Cell Rna Sequencing ◽  
Transcript Profiles

Abstract We explored cellular heterogeneity in medulloblastoma using single-cell RNA sequencing (scRNAseq), immunohistochemistry and deconvolution of bulk transcriptomic data. Over 45,000 cells from 31 patients from all main subgroups of medulloblastoma (2 WNT, 10 SHH, 9 GP3, 11 GP4 and 1 GP3/4) were clustered using Harmony alignment to identify conserved subpopulations. Each subgroup contained subpopulations exhibiting mitotic, undifferentiated and neuronal differentiated transcript profiles, corroborating other recent medulloblastoma scRNAseq studies. The magnitude of our present study builds on the findings of existing studies, providing further characterization of conserved neoplastic subpopulations, including identification of a photoreceptor-differentiated subpopulation that was predominantly, but not exclusively, found in GP3 medulloblastoma. Deconvolution of MAGIC transcriptomic cohort data showed that neoplastic subpopulations are associated with major and minor subgroup subdivisions, for example, photoreceptor subpopulation cells are more abundant in GP3-alpha. In both GP3 and GP4, higher proportions of undifferentiated subpopulations is associated with shorter survival and conversely, differentiated subpopulation is associated with longer survival. This scRNAseq dataset also afforded unique insights into the immune landscape of medulloblastoma, and revealed an M2-polarized myeloid subpopulation that was restricted to SHH medulloblastoma. Additionally, we performed scRNAseq on 16,000 cells from genetically engineered mouse (GEM) models of GP3 and SHH medulloblastoma. These models showed a level of fidelity with corresponding human subgroup-specific neoplastic and immune subpopulations. Collectively, our findings advance our understanding of the neoplastic and immune landscape of the main medulloblastoma subgroups in both humans and GEM models.

scholarly journals Panel of serum miRNAs as potential non-invasive biomarkers for pancreatic ductal adenocarcinoma

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Imteyaz Ahmad Khan ◽  
Safoora Rashid ◽  
Nidhi Singh ◽  
Sumaira Rashid ◽  
Vishwajeet Singh ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Pancreatic Ductal Adenocarcinoma ◽  
Early Stage ◽  
Ductal Adenocarcinoma ◽  
Next Generation ◽  
Serum Samples ◽  
Tissue Samples ◽  
Non Invasive ◽  
Specific Symptoms ◽  
Generation Sequencing

AbstractEarly-stage diagnosis of pancreatic ductal adenocarcinoma (PDAC) is difficult due to non-specific symptoms. Circulating miRNAs in body fluids have been emerging as potential non-invasive biomarkers for diagnosis of many cancers. Thus, this study aimed to assess a panel of miRNAs for their ability to differentiate PDAC from chronic pancreatitis (CP), a benign inflammatory condition of the pancreas. Next-generation sequencing was performed to identify miRNAs present in 60 FFPE tissue samples (27 PDAC, 23 CP and 10 normal pancreatic tissues). Four up-regulated miRNAs (miR-215-5p, miR-122-5p, miR-192-5p, and miR-181a-2-3p) and four down-regulated miRNAs (miR-30b-5p, miR-216b-5p, miR-320b, and miR-214-5p) in PDAC compared to CP were selected based on next-generation sequencing results. The levels of these 8 differentially expressed miRNAs were measured by qRT-PCR in 125 serum samples (50 PDAC, 50 CP, and 25 healthy controls (HC)). The results showed significant upregulation of miR-215-5p, miR-122-5p, and miR-192-5p in PDAC serum samples. In contrast, levels of miR-30b-5p and miR-320b were significantly lower in PDAC as compared to CP and HC. ROC analysis showed that these 5 miRNAs can distinguish PDAC from both CP and HC. Hence, this panel can serve as a non-invasive biomarker for the early detection of PDAC.

scholarly journals Genes associated with inflammation and bone remodeling are highly expressed in the bone of patients with the early-stage cam-type femoroacetabular impingement

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Guanying Gao ◽  
Ruiqi Wu ◽  
Rongge Liu ◽  
Jianquan Wang ◽  
Yingfang Ao ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bone Tissue ◽  
Bone Remodeling ◽  
Femoroacetabular Impingement ◽  
Early Stage ◽  
Control Group ◽  
Tissue Samples ◽  
Expression Levels ◽  
Normal Bone ◽  
Impingement Zone

Abstract Background Recent studies have shown high expression levels of certain inflammatory, anabolic, and catabolic genes in the articular cartilage from the impingement zone of the hips with femoroacetabular impingement (FAI), representing an increased metabolic state. Nevertheless, little is known about the molecular properties of bone tissue from the impingement zone of hips with FAI. Methods Bone tissue samples from patients with early-stage cam-type FAI were collected during hip arthroscopy for treatment of cam-type FAI. Control bone tissue samples were collected from six patients who underwent total hip replacement because of a femoral neck fracture. Quantitative real-time polymerase chain reaction (PCR) was performed to determine the gene expression associated with inflammation and bone remodeling. The differences in the gene expression in bone tissues from the patients with early-stage cam-type FAI were also evaluated based on clinical parameters. Results In all, 12 patients with early-stage cam-type FAI and six patients in the control group were included in this study. Compared to the control samples, the bone tissue samples from patients with FAI showed higher expression levels of interleukin-6 (IL-6), alkaline phosphatase (ALP), receptor activator of nuclear factor-kB ligand (RANKL), and osteoprotegerin (OPG) (P < 0.05). IL-1 expression was detected only in the control group. On the other hand, there was no significant difference in IL-8 expression between the patients with FAI and the control group. The patients with FAI having a body mass index (BMI) of >24 kg/m2 showed higher ALP expression (P < 0.05). Further, the expression of IL-6 and ALP was higher in the patients with FAI in whom the lateral center-edge angle was >30° (P < 0.05). Conclusions Our results indicated the metabolic condition of bone tissues in patients with early-stage cam-type FAI differed from that of normal bone in the femoral head-neck junction. The expression levels of the genes associated with inflammation and bone remodeling were higher in the bone tissue of patients with early-stage cam-type FAI than in the patients with normal bone tissue.

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