Normal Peripheral Tissue Research Articles

14 Natural killer cells have impaired cytotoxicity in advanced renal cell carcinoma

Abstract Background Natural killer (NK) cells are key mediators of anti-tumor activity in multiple cancers, including clear cell renal cell carcinoma (ccRCC). However, the phenotype and function of NK cells across different disease stages of ccRCC is incompletely characterized. Methods Single-cell RNA (scRNA)-sequencing (10x Genomics) data from tumor and adjacent normal kidney from ccRCC was analyzed at multiple clinical stages (localized – stage I/II/III; and advanced – stage IV). Graph-based clustering and lineage markers were used to identify distinct NK cell populations. Differential gene expression analysis was performed to further investigate NK cell phenotype and to derive a gene expression signature (GES). Gene signatures from NK cell subclusters of interest were used to interrogate bulk transcriptomic datasets and expression with clinical outcomes. Tumor-infiltrating NK cell function (cytokine production and cytotoxicity) was assessed by isolation of live NK cells from ccRCC tissue, co-culture with K562 target cells, and measurement of cytokine (IFNgamma) and cytotoxicity (CD107a) markers by flow cytometry. Results Single-cell transcriptomic data were analyzed from 13 patients with ccRCC (tumor and normal kidney), resulting in 21,139 high-quality NK cells. Patients with advanced/metastatic RCC had a lower proportion of NK cells versus total immune cells in the tumor microenvironment compared to normal kidney (p=0.036) and localized ccRCC (p=0.088). Clustering analysis revealed 6 distinct NK cell subsets. The C0.Bright-like NK cell cluster was significantly enriched in advanced ccRCC compared to localized ccRCC (p=0.048) and normal kidney (p=0.0059), expressed markers of tissue residency (ZNF683, ITGA1, CD9, ITGAE), and had decreased expression of cytotoxicity genes (GZMB, PRF1). A GES signature composed of genes upregulated in the dysfunctional C0.Bright-like NK cell cluster was used to interrogate bulk RNA-sequencing data from The Cancer Genome Atlas clear cell cohort, providing validation in a large, independent cohort, of an increase in this dysfunctional NK subset in advanced ccRCC compared to localized ccRCC (p=0.00039) and normal tissue (p=5.3e-05). To functionally confirm the decreased cytotoxicity of this dysfunctional NK population, CD45+CD56+CD3- NK cells were isolated from advanced ccRCC, and co-cultured with K562 target cells. Consistent with the transcriptomic phenotype, tumor-resident CD49a+CD9+ NK cells had cytokine production (IFNgamma) but decreased markers of cytotoxicity (CD107a) compared to CD49a-CD9- NK cells. By contrast, CD49a+CD9+ NK cells isolated from peripheral blood or normal kidney tissue maintained cytotoxic activity. Conclusions Among patients with ccRCC, a single-cell transcriptomic analysis revealed heterogeneous NK cell populations. A dysfunctional tumor resident NK cell phenotype was enriched among patients with advanced disease in ccRCC, and functionally confirmed to have diminished cytotoxicity. This study therefore provides insight into a new axis of immune dysfunction – decreased NK-mediated cytotoxicity – in advanced ccRCC. An improved understanding of NK cell dysfunction within ccRCC may provide a foundation for therapeutic restoration of NK-mediated anti-tumor activity in ccRCC. DOD CDMRP Funding: yes

Expression of Intragenic LCAL4 Long Non-Coding RNAs as a Potential Diagnostic and Prognostic Marker in Female Breast Cancer.

In breast cancer early detection is associated with reduced mortality and it is essential to identify new biomarkers for early detection and appropriate management of cancer patients with the best response to treatment. Long non-coding RNAs (LncRNAs) have attracted much attention as potential diagnostic, prognostic, or predictive biomarkers due to their high specificity, easy access to non-invasive methods, and their aberrant expression under various pathological and physiological conditions. Have attracted the aim of this study is to investigate the expression profile of intragenic non-coding LncRNAs LCAL4 as a biomarker as potential diagnostic and prognostic biomarkers in cancer. In this research, 62 tissue samples were obtained from patients undergoing therapeutic surgery in Khatam al-Anbia Hospital and the normal peripheral tissue that was removed for prevention was used as a control by Real-time PCR method. The expression pattern of LCAL4 long non-coding RNA gene is significantly different between two groups of healthy control samples and samples obtained from patients with different breast cancer subtypes, Also its expression between samples obtained from different subgroups and different stages showed significant differences. The studied LncRNAs can act as a factor to identify tumor tissue from healthy tissue, and the diagnosis of cancer grades can be different depending on the type of LncRNA. These results can be proposed in the introduction of LncRNA LCAL4 as a new marker in the diagnosis of breast cancer. In addition, by interpreting the results, it can be concluded that these LncRNAs can be considered as influential factors in the process of breast cancer.

Glucose and Oxygen Levels Modulate the Pore-Forming Effects of Cholesterol-Dependent Cytolysin Pneumolysin from Streptococcus pneumoniae.

A major Streptococcus pneumoniae pathogenic factor is the cholesterol-dependent cytolysin pneumolysin, binding membrane cholesterol and producing permanent lytic or transient pores. During brain infections, vascular damage with variable ischemia occurs. The role of ischemia on pneumolysin's pore-forming capacity remains unknown. In acute brain slice cultures and primary cultured glia, we studied acute toxin lysis (via propidium iodide staining and LDH release) and transient pore formation (by analyzing increases in the intracellular calcium). We analyzed normal peripheral tissue glucose conditions (80 mg%), normal brain glucose levels (20 mg%), and brain hypoglycemic conditions (3 mg%), in combinations either with normoxia (8% oxygen) or hypoxia (2% oxygen). At 80 mg% glucose, hypoxia enhanced cytolysis via pneumolysin. At 20 mg% glucose, hypoxia did not affect cell lysis, but impaired calcium restoration after non-lytic pore formation. Only at 3 mg% glucose, during normoxia, did pneumolysin produce stronger lysis. In hypoglycemic (3 mg% glucose) conditions, pneumolysin caused a milder calcium increase, but restoration was missing. Microglia bound more pneumolysin than astrocytes and demonstrated generally stronger calcium elevation. Thus, our work demonstrated that the toxin pore-forming capacity in cells continuously diminishes when oxygen is reduced, overlapping with a continuously reduced ability of cells to maintain homeostasis of the calcium influx once oxygen and glucose are reduced.

Evaluation of Potential miR-302, miR-132, miR-205, and miR-126 as Biomarkers for Laryngeal Squamous Cell Carcinoma

Background: Laryngeal tumor is the most commonly identified malignancy in head and neck cancers, containing 2.4% of all cancers, and is the 11th most prevalent and fatal cancer in the world. Previous research has revealed that microRNAs (miRNAs) have a significant role in the progression of laryngeal tumors. Additionally, research focuses on the association with the clinical function of miRNA in malignancy. Objectives: Based on previous research, miR-132, miR-302, miR-126, and miR-205 have potential roles in the tumor development of the LSCC. Then, in the current study, we pointed to study if there are any significant alters in the expression of microRNAs and whether they have any significant possibility as a diagnostic or prognostic factor for laryngeal squamous cell carcinoma. Methods: From 30 LSCC patients, we have collected tumor and marginal healthy tissues through laryngectomy. Total RNA extraction from the marginal healthy and malignancy tissues was done. Next, we control the quality of the RNAs and, then, synthesize cDNA. Lastly, the expression of the miRNAs was evaluated by qPCR. The expression level of genes and their association with the patient’s clinicopathological features were studied, using related statistical tests. Results: miR-132 (P = 0.0001, fold change: 1.28) and miR-302 (P = 0.0001, fold change: 1.o5) are significantly up-regulated in tumor tissues compared to normal peripheral tissues. Furthermore, we revealed that miR-205 (P < 0.0001, fold change: 0.241) and miR-126 (P = 0.0001, fold change: 0.251) are low expressed in LSCC cancer tissue. Between these miRNAs, gust expression of the miR-205 had no relationship with clinical-pathological characteristics in malignancy tissue. Conclusions: The results of the current study showed the possibility of miR-302, miR-132, miR-126, and miR-205 as diagnostic or prognostic factors in LSCC.

Regulation of PD-L1 Expression by Nuclear Receptors.

The suppression of excessive immune responses is necessary to prevent injury to the body, but it also allows cancer cells to escape immune responses and proliferate. Programmed cell death 1 (PD-1) is a co-inhibitory molecule that is present on T cells and is the receptor for programmed cell death ligand 1 (PD-L1). The binding of PD-1 to PD-L1 leads to the inhibition of the T cell receptor signaling cascade. PD-L1 has been found to be expressed in many types of cancers, such as lung, ovarian, and breast cancer, as well as glioblastoma. Furthermore, PD-L1 mRNA is widely expressed in normal peripheral tissues including the heart, skeletal muscle, placenta, lungs, thymus, spleen, kidney, and liver. The expression of PD-L1 is upregulated by proinflammatory cytokines and growth factors via a number of transcription factors. In addition, various nuclear receptors, such as androgen receptor, estrogen receptor, peroxisome-proliferator-activated receptor γ, and retinoic-acid-related orphan receptor γ, also regulate the expression of PD-L1. This review will focus on the current knowledge of the regulation of PD-L1 expression by nuclear receptors.

POTENTIAL OF microRNAs AS A BIOMARKER FOR LARYNGEAL CANCER

Objective: Laryngeal malignancy known as the most frequently diagnosed cancer between head and neck tumor, covering 2.4% of all tumors and is the 11th most occurrence and deadliest tumor in the world. Research has shown that microRNAs (miRNAs) play a considerable role in the development of laryngeal malignancy. Moreover, studies also focus on the relationship between clinical relevance and miRNA deregulation in cancer illness. According to recent evidence, miR-205, miR-126, miR-302, and miR-132 have possible roles in the carcinogenesis process of the LSCC. Therefore, in this research, we aimed to investigate whether there are any considerable changes in the microRNAs expression level and also if they have any remarkable potential as a prognostic or diagnostic biomarker for laryngeal squamous cell carcinoma (LSCC). Methods: Thirty LSCC patients were included in this research, and tumor tissues and healthy normal tissues were collected during laryngectomy. Total RNA extraction from the normal and cancer tissues was accomplished and after the quality control, cDNA was synthesized, and finally, target miRNAs expression was determined by qPCR. The expression level of genes and their relationship with the patient’s clinicopathological characteristics were analyzed using appropriate statistical tests. Results: According to the study results, miR-302 (fold change: 1.o5, P-value = 0.0001) and miR-132 (fold change: 1.28, P-value = 0.0001), is considerably overexpressed in cancer tissues compare to normal peripheral tissues. We also found that miR-126 (fold change: 0.251, P-value = 0.0001) and miR-205 (fold change: 0.241, P-value<0.0001) is down-regulated in LSCC tumor tissue. Among these microRNAs, only mir-205 expression had no association with clinical-pathological features in tumor tissue. Conclusion: The results of this research revealed the potential of the miR-302, miR-132, miR-126, and miR-205 as prognostic or diagnostic biomarkers in LSCC.

High Expression of Heat Shock Protein Family D Member 1 Predicts Poor Prognosis of Esophageal Cancer.

BackgroundHeat shock protein family D (Hsp60) member 1 (HSPD1) has been reported as a potential survival-related biomarker in some cancers. However, the correlation between HSPD1 expression with prognosis and clinical features of esophageal cancer (EC) is poorly understood. Our research aimed to explore the clinical and prognostic significance of HSPD1 expression in EC patients.MethodsIn our study, HSPD1 expression was detected by immunochemistry in 87 EC tissue specimens and 20 normal cancerous peripheral tissue specimens. Meanwhile, we also analyzed the expression of HSPD1 in EC by The Cancer Genome Atlas (TCGA) database. Then Chi-squared and Fisher’s exact tests and Wilcoxon signed-rank test and logistic regression models were separately used to test the correlation between clinical characteristics and HSPD1 expression in our and TCGA cohort. Moreover, we evaluated the value of HSPD1 in prognosis by Kaplan-Meier curves and Cox analysis. Finally, gene set enrichment analysis (GSEA) was performed using the data accessed from TCGA.ResultsThe results showed that HSPD1 was overexpressed in EC, and the expression was related to histological type, histological grade, N classification, and clinical stage. Moreover, Kaplan-Meier curves and Cox analysis indicated that high expression of HSPD1 correlated with poor prognosis, and HSPD1 was an independent risk factor for EC. GSEA identified pathways involved in cysteine and methionine metabolism, spliceosome, selenoamino acid metabolism, mismatch repair, RNA degration, DNA replication, and cell cycle as differentially enriched in ECs with high HSPD1 expression.ConclusionsOur results suggest that HSPD1 is expressed at high levels in EC, and has potential to be used as a novel biomarker for the prognosis of patients with EC.

DNA methylation patterns at and beyond the histological margin of early-stage invasive lung adenocarcinoma radiologically manifested as pure ground-glass opacity

BackgroundEarly-stage lung cancers radiologically manifested as ground-glass opacities (GGOs) have been increasingly identified, among which pure GGO (pGGO) has a good prognosis after local resection. However, the optimal surgical margin is still under debate. Precancerous lesions exist in tumor-adjacent tissues beyond the histological margin. However, potential precancerous epigenetic variation patterns beyond the histological margin of pGGO are yet to be discovered and described.ResultsA genome-wide high-resolution DNA methylation analysis was performed on samples collected from 15 pGGO at tumor core (TC), tumor edge (TE), para-tumor tissues at the 5 mm, 10 mm, 15 mm, 20 mm beyond the tumor, and peripheral normal (PN) tissue. TC and TE were tested with the same genetic alterations, which were also observed in histologically normal tissue at 5 mm in two patients with lower mutation allele frequency. According to the difference of methylation profiles between PN samples, 2284 methylation haplotype blocks (MHBs), 1657 differentially methylated CpG sites (DMCs), and 713 differentially methylated regions (DMRs) were identified using reduced representation bisulfite sequencing (RRBS). Two different patterns of methylation markers were observed: Steep (S) markers sharply changed at 5 mm beyond the histological margin, and Gradual (G) markers changed gradually from TC to PN. S markers composed 86.2% of the tumor-related methylation markers, and G markers composed the other 13.8%. S-marker-associated genes enriched in GO terms that were related to the hallmarks of cancer, and G-markers-associated genes enriched in pathways of stem cell pluripotency and transcriptional misregulation in cancer. Significant difference in DNA methylation score was observed between peripheral normal tissue and tumor-adjacent tissues 5 mm further from the histological margin (p < 0.001 in MHB markers). DNA methylation score at and beyond 10 mm from histological margin is not significantly different from peripheral normal tissues (p > 0.05 in all markers).ConclusionsAccording to the methylation pattern observed in our study, it was implied that methylation alterations were not significantly different between tissues at or beyond P10 and distal normal tissues. This finding explained for the excellent prognosis from radical resections with surgical margins of more than 15 mm. The inclusion of epigenetic characteristics into surgical margin analysis may yield a more sensitive and accurate assessment of remnant cancerous and precancerous cells in the surgical margins.

Exploring the diagnostic potential of adding T2 dependence in diffusion-weighted MR imaging of the prostate.

Magnetic resonance imaging (MRI) is essential in the detection and staging of prostate cancer. However, improved tools to distinguish between low-risk and high-risk cancer are needed in order to select the appropriate treatment. To investigate the diagnostic potential of signal fractions estimated from a two-component model using combined T2- and diffusion-weighted imaging (T2-DWI). 62 patients with prostate cancer and 14 patients with benign prostatic hyperplasia (BPH) underwent combined T2-DWI (TE = 55 and 73 ms, b-values = 50 and 700 s/mm2) following clinical suspicion of cancer, providing a set of 4 measurements per voxel. Cancer was confirmed in post-MRI biopsy, and regions of interest (ROIs) were delineated based on radiology reporting. Signal fractions of the slow component (SFslow) of the proposed two-component model were calculated from a model fit with 2 free parameters, and compared to conventional bi- and mono-exponential apparent diffusion coefficient (ADC) models. All three models showed a significant difference (p<0.0001) between peripheral zone (PZ) tumor and normal tissue ROIs, but not between non-PZ tumor and BPH ROIs. The area under the receiver operating characteristics curve distinguishing tumor from prostate voxels was 0.956, 0.949 and 0.949 for the two-component, bi-exponential and mono-exponential models, respectively. The corresponding Spearman correlation coefficients between tumor values and Gleason Grade Group were fair (0.370, 0.499 and -0.490), but not significant. Signal fraction estimates from a two-component model based on combined T2-DWI can differentiate between tumor and normal prostate tissue and show potential for prostate cancer diagnosis. The model performed similarly to conventional diffusion models.

Streptococcus australis and Ralstonia pickettii as Major Microbiota in Mesotheliomas.

The microbiota has been reported to be correlated with carcinogenesis and cancer progression. However, its involvement in the pathology of mesothelioma remains unknown. In this study, we aimed to identify mesothelioma-specific microbiota using resected or biopsied mesothelioma samples. Eight mesothelioma tissue samples were analyzed via polymerase chain reaction (PCR) amplification and 16S rRNA gene sequencing. The operational taxonomic units (OTUs) of the effective tags were analyzed in order to determine the taxon composition of each sample. For the three patients who underwent extra pleural pneumonectomy, normal peripheral lung tissues adjacent to the tumor were also included, and the same analysis was performed. In total, 61 OTUs were identified in the tumor and lung tissues, which were classified into 36 species. Streptococcus australis and Ralstonia pickettii were identified as abundant species in almost all tumor and lung samples. Streptococcus australis and Ralstonia pickettii were found to comprise mesothelioma-specific microbiota involved in tumor progression; thus, they could serve as targets for the prevention of mesothelioma.

Activation of MAT2A-RIP1 signaling axis reprograms monocytes in gastric cancer

BackgroundThe activation of tumor-associated macrophages (TAMs) facilitates the progression of gastric cancer (GC). Cell metabolism reprogramming has been shown to play a vital role in the polarization of TAMs. However,...

Cybersecurity in Brain-Computer Interfaces: RFID-based design-theoretical framework

Abstract Brain Computer Interface BCI is a real-time communication system that connects the brain and external devices. The BCI system can directly convert the information sent by the brain into commands that can drive external devices and replace human limbs or language organs to achieve hu-man communication with the outside world and the external environment Control. In other words, the BCI system can replace the normal peripheral nerve and muscle tissue to achieve communication between the human and the computer or between the human and the external environment. The objective of this paper is to assist the network security for the BCI applications to identify brain activities in a secure real-time mode. To achieve this, we proposed the design of a Radio Frequency Identification RFID-based system having semi− active RFID tags placed outside the brain on the scalp transmit the collected brain activities wirelessly to a devise SC (Scanner Controller) consist of mini-reader and timer integrated together for every patient. Additionally, the paper implemented a novel system prototype interface called BCI Identification System (BCIIS) to assist the patient in the identification process. Given the benefits of RFID, we believed that if the idea is adopted and implemented by industry, it could enhance and provide secure BCI applications.

Cytochemically Demonstrable B-Glucuronidase Activity in Normal and Neoplastic Human Lymphoid Cells

Mononuclear cell suspensions were prepared from 40 normal peripheral blood and lymphoid tissue specimens and 42 neoplastic specimens obtained from patients with malignant lymphoma and lymphocytic leukemia. These suspensions were analyzed for la antigens, surface immunoglobulin (Slg), sheep erythrocyte (E) rosette formation and, in some instances, acid alpha-naphthyl acetate esterase (ANAE) activity. The results of these studies were correlated with the expression of cytochemically demonstrable BG activity. The percentage of BG+ lymphocytes was found to be comparable, within 10%, to the percentage of E+ (T) cells in the majority of normal, non-neoplastic peripheral blood, tonsil, spleen, and lymph node specimens examined. Occasionally, the percentage of E+ cells exceeded the percentage of BG+ cells by 20% or more, suggesting the presence of an E+BG- T cell subpopulation. BG+ B lymphocytes were only demonstrated in 1 of 40 non-neoplastic lymphoid specimens. The neoplastic B cells in each of 14 B cell (la+Slg+E-) lymphomas were BG-. However, a variable proportion of the neoplastic cells isolated from 6 cases of B cell chronic lymphocytic leukemia and neoplastic plasma cells isolated from 7 cases of multiple myeloma expressed BG activity. Thus, it appears that both normal and neoplastic BG- and BG+ B lymphocyte populations exist; the latter may be related to a state of activation or a stage of B cell differentiation. The neoplastic cells isolated from 4 T cell (la-Slg-E+) malignancies were BG+ while those isolated from 3 T cell malignancies were BG-. The variable expression of BG activity by T cell malignancies may be related to T cell differentiation. Investigation of BG expression by T cell derived malignancies may prove useful in sorting out T cell phenotypes.

Evaluation of extracellular signal regulated kinase 1 expression in premalignant lesions of oral cavity

Background: Most oral squamous cell carcinomas are preceded by premalignant lesions like leukoplakia, erythroplakia etc. Identification of these lesions by molecular markers and intervention at an early stage can reduce mortality, morbidity and cost of treatment associated with oral cancer. Extracellular signal regulated kinase 1 (ERK 1) is one of the various markers involved in carcinogenesis.Methods: We studied the expression of ERK 1 in twenty-five oral premalignant lesion biopsies using immunohistochemistry (IHC) and western blot analysis.Results: Ninety-two percent of premalignant lesions showed positive expression of ERK 1 on IHC and the same were confirmed by western blot analysis. ERK 1 expression was increased in premalignant lesions as compared to peripheral normal tissue but the difference between them was not statistically significant. On comparing the level of expression of peripheral normal and premalignant tissue on IHC, moderate and high level of expressions were significantly higher in premalignant lesions (24%) than peripheral normal tissue (12%).Conclusions: There was an increased expression of ERK 1 in oral premalignant lesions as compared to peripheral normal tissue, although, this difference was not statistically significant.

Matrix metalloproteinase MMP1 and MMP9 genes expression in breast cancer tissue

Nowadays, Breast cancer is recognized as the most common cancer among women worldwide. Matrix metalloproteinases (MMPs) are a group of endopeptidase enzymes that can degrade extracellular matrix proteins and increase invasion and metastasis. The purpose of this study was to evaluate the expression of MMP1 and MMP9 genes in breast cancer tissue. In the present case-control study, 90 breast tissue sample, includes 45 breast cancer tissue samples as case group and 45 healthy breast tissue samples as control group were studied. After RNA extraction and cDNA synthesis, MMP1 and MMP9 genes mRNA expression levels were evaluated using Real-Time PCR. In the present study, a significant increase was observed in the expression levels of MMP1 and MMP9 genes mRNA expression in breast cancer tissues compared to normal peripheral tissues. The mRNA expression levels of MMP1 and MMP9 genes in breast cancer tissue were increased by 2.5 and 2.8 fold, respectively. Due to the increased expression of MMP1 and MMP9 genes in breast cancer tissue samples, these proteins can be used as diagnostic markers and novel therapeutic targets. However, further studies with larger sample sizes and on other MMPs are recommended.

Photothermal Therapy Combined with Light-Induced Generation of Alkyl Radicals for Enhanced Efficacy of Tumor Treatment

Photothermal therapy (PTT) shows great potential in tumor treatment, but the harsh photothermal heating might give rise to irreversible damage to peripheral normal tissues. Combining PTT with the a...

Distribution-Free Multisample Tests Based on Optimal Matchings With Applications to Single Cell Genomics

In this article, we propose a nonparametric graphical test based on optimal matching, for assessing the equality of multiple unknown multivariate probability distributions. Our procedure pools the data from the different classes to create a graph based on the minimum non-bipartite matching, and then utilizes the number of edges connecting data points from different classes to examine the closeness between the distributions. The proposed test is exactly distribution-free (the null distribution does not depend on the distribution of the data) and can be efficiently applied to multivariate as well as non-Euclidean data, whenever the inter-point distances are well-defined. We show that the test is universally consistent, and prove a distributional limit theorem for the test statistic under general alternatives. Through simulation studies, we demonstrate its superior performance against other common and well-known multisample tests. The method is applied to single cell transcriptomics data obtained from the peripheral blood, cancer tissue, and tumor-adjacent normal tissue of human subjects with hepatocellular carcinoma and non-small-cell lung cancer. Our method unveils patterns in how biochemical metabolic pathways are altered across immune cells in a cancer setting, depending on the tissue location. All of the methods described herein are implemented in the R package multicross. Supplementary materials for this article are available online.

Clinical Characteristics and Mutation Analyses of Ovarian Sertoli-Leydig Cell Tumors.

There are limited studies on Sertoli-Leydig cell tumors (SLCTs) and no data in the population of Chinese patients with SLCTs from the genetic level. In addition, previous studies on SLCTs have focused exclusively on mutations in the DICER1 gene and no data exists on the genetic landscape of SLCTs. Patients with moderately or poorly differentiated SLCTs who underwent surgical resection between January 2012 and October 2018 in our institution were recruited. Whole exome sequencing was performed on formalin-fixed, paraffin-embedded tumor tissue and peripheral blood or normal tissue samples. Seventeen patients were recruited with 19 tumor samples. The rate of tumor-associated germline mutations was 6 of 17 (35.3%), and that of DICER1 germline mutations was 4 of 17 (23.5%). Regarding clinical relapse, patients with germline tumor-associated mutations had significantly poorer prognosis than those without (p = .007), and those with germline DICER1 mutations were relatively more likely to exhibit clinical relapse, although not to a significant degree (p = .069). Regarding somatic mutations, firstly, the subclone evolution analysis demonstrated that the two tumors on the contralateral ovary were primary tumors, respectively. Secondly, somatic mutations were most commonly found in CDC27 (10/19, 52.6%), DICER1 (4/19, 21.1%), and MUC22 (4/19, 21.1%). And the analysis of cancer cell fractions showed that DICER1 mutations were correlated with tumorigenesis of SLCTs. The rates of germline and somatic DICER1 mutations were higher in patients who were younger than 18 years than those in older patients (p = .022 and p = .001, respectively). Our study indicates that genetic testing may have important clinical significance for patients with SLCTs, particularly for younger patients. Bilateral ovarian Sertoli-Leydig cell tumors were verified to be primary tumors from the genetic perspective. The rates of germline and somatic DICER1 mutations were 4 of 17 (23.5%) and 4 of 19 (21.1%), respectively. The rates of germline and somatic DICER1 mutations were higher in patients who were younger than 18 years than those in older patients (p = .022 and p = .001, respectively).

Neoantigen screening identifies EGFR L858R mutation immunogenicity in 1862 Chinese NSCLC patients.

e21512 Background: Therapeutic vaccines targeting mutation-derived neoantigens prime T-cell responses and deliver long-term clinical benefit to patients. Last year we reported EGFR L858R mutation could be a possible target for individual-independent vaccine in 799 Chinese patients with non-small cell lung cancer (NSCLC). Here we verified the findings and explored possible immunological mechanisms in a larger sample size of 1862 Chinese NSCLC. Peripheral blood or normal tissue was used as control. Methods: DNA sequencing data was acquired using our targeted 1021-gene panel and HLA-I genotyping was determined based on DNA sequencing by OptiType v1.0. The peptide-HLA binding affinity was predicted with netMHCpan v4.0. Neoantigen was identified if the IC50 MT &lt; 500 nM and IC50 WT &gt; 500 nM, furthermore, IC50 MT &lt; 50 nM was considered as strong-binders. Results: EGFR (50.05%) was the most prevalent mutated gene, with 22.61% and 13.16% harboring L858R mutations and E746_A750del respectively. HLA typing showed HLA-A*11:01(42.59%) was the top one allele and HLA-A*33:03(12.94%) ranked 12th. The combination of EGFR L858R and HLA-A*33:03 had both relatively strongest binding affinity (IC50 MT = 22.9 nM and IC50 WT = 12734.0 nM, 2.93%) and highest shared frequency (2.93%). In terms of mechanism, we examined 7 of 1862 (0.37%) B2M gene mutation and 1092 of 1731 (63.1%) HLA-I LOH (loss of heterozygosity) in our population, but didn’t see any statistical difference between L858R subtype and other position mutations in EGFR. We also calculated 26 immune cell types’ signature scores from Chinese CHOICE study and found L858R subtype was associated with elevated level of pDC (KW-test p = 0.02) and lower level of neutrophils signature(KW-test p = 0.07) compared with exon 19 deletion and other uncommon mutations of EGFR. Conclusions: (1) Targeting EGFR L858R mutation in patients with HLA-A*33:03 allele appeared to be valuable for the neoantigen-based vaccine designed for Chinese NSCLC patients. (2) The specific immunogenicity of L858R seemed to be little influenced by B2M mutation and HLA-I LOH which were important factors for neoantigen presentation. (3) High pDC and low neutrophils infiltration might contribute to the inhibitive microenvironment associated with L858R mutation.

Aberrant Expression of miR-103, miR-184, miR-378, miR-497 and miR-506 in Tumor Tissue from Patients with Oral Squamous Cell Carcinoma Regulates the Clinical Picture of the Patients.

Background:This study aimed to evaluate the expression patterns of miR-103, miR-184, miR-378, miR497 and in squamous cell carcinoma (SCC) of the tongue and to be compared with normal peripheral tissues. Methods:Tumor and marginal tissues were obtained from 50 patients with OSCC. After RNA extraction, expression level of miR-103, miR-184, miR-378, miR497, and miR506 was estimated using SYBR green master mix and real-time quantitative PCR. Results:It was observed that, there was no detectable difference in expression level of miR-103 between tumoral and marginal tissues. However, expression level of miR-184, and miR-378 showed significant increase in tumor tissue samples compared to marginal tissue samples. MiR-497 and miR-506 demonstrated considerable decrease in tumoral cells in comparison with peripheral tissues. Moreover, the expression level of miRNAs was associated with clinicopathological features of the patients. Conclusions:Our data indicated that miR-184, miR-378, miR-497, and miR-506 can be used as a potential diagnostic and prognostic biomarker in OSCC. Nevertheless, more studies are needed to confirm this claim.