mRNA Profiles Research Articles

LncRNA and mRNA expression characteristic and bioinformatic analysis in myocardium of diabetic cardiomyopathy mice

BackgroundDiabetic cardiomyopathy (DCM) is becoming a very well-known clinical entity and leads to increased heart failure in diabetic patients. Long non-coding RNAs (LncRNAs) play an important role in the pathogenesis of DCM. In the present study, the expression profiles of lncRNAs and mRNAs were illuminated in myocardium from DCM mice, with purpose of exploring probable pathological processes of DCM involved by differentially expressed genes in order to provide a new direction for the future researches of DCM.ResultsThe results showed that a total of 93 differentially expressed lncRNA transcripts and 881 mRNA transcripts were aberrantly expressed in db/db mice compared with the controls. The top 6 differentially expressed lncRNAs like up-regulated Hmga1b, Gm8909, Gm50252 and down-regulated Msantd4, 4933413J09Rik, Gm41414 have not yet been reported in DCM. The lncRNAs-mRNAs co-expression network analysis showed that LncRNA 2610507I01Rik, 2310015A16Rik, Gm10503, A930015D03Rik and Gm48483 were the most relevant to differentially expressed mRNAs.ConclusionOur results showed that db/db DCM mice exist differentially expressed lncRNAs and mRNAs in hearts. These differentially expressed lncRNAs may be involved in the pathological process of cardiomyocyte apoptosis and fibrosis in DCM.

Construction of the underlying circRNA-miRNA-mRNA regulatory network and a new diagnostic model in ulcerative colitis by bioinformatics analysis.

Circular RNAs (circRNAs) are involved in the pathogenesis of many diseases through competing endogenous RNA (ceRNA) regulatory mechanisms. To investigate a circRNA-related ceRNA regulatory network and a new predictive model by circRNA to understand the diagnostic mechanism of circRNAs in ulcerative colitis (UC). We obtained gene expression profiles of circRNAs, miRNAs, and mRNAs in UC from the Gene Expression Omnibus dataset. The circRNA-miRNA-mRNA network was constructed based on circRNA-miRNA and miRNA-mRNA interactions. Functional enrichment analysis was performed to identify the biological mechanisms involved in circRNAs. We identified the most relevant differential circRNAs for diagnosing UC and constructed a new predictive nomogram, whose efficacy was tested with the C-index, receiver operating characteristic curve (ROC), and decision curve analysis (DCA). A circRNA-miRNA-mRNA regulatory network was obtained, containing 12 circRNAs, three miRNAs, and 38 mRNAs. Two optimal prognostic-related differentially expressed circRNAs, hsa_circ_0085323 and hsa_circ_0036906, were included to construct a predictive nomogram. The model showed good discrimination, with a C-index of 1(> 0.9, high accuracy). ROC and DCA suggested that the nomogram had a beneficial diagnostic ability. This novel predictive nomogram incorporating hsa_circ_0085323 and hsa_circ_0036906 can be conveniently used to predict the risk of UC. The circRNa-miRNA-mRNA network in UC could be more clinically significant.

Analysis of differentially expressed lncRNAs and mRNAs associated with slow‑transit constipation

Slow transit constipation (STC) is a refractory gastrointestinal disease, accounting for approximately 13 ∼ 37 % of chronic constipation. However, the molecular mechanism of STC remains poorly understood. Herein, this study aims to identify the key mRNAs and lncRNAs associated with STC. To this end, we performed high-throughput RNA sequencing to identify differentially expressed (DE) mRNAs and lncRNAs in the whole-layer sigmoid intestinal tissues from 4 STC patients and 4 non-STC patients. The identified DE lncRNAs and mRNAs were validated through quantitative real-time PCR. Weighted gene co-expression network analysis (WGCNA) and Pearson correlation analysis were conducted to determine the significantly correlated DE mRNA-lncRNA pairs. A total of 1420 DE lncRNAs and 1634 DE mRNAs were identified. Kyoto Encyclopedia of Genes and Genomes analysis of DE mRNAs indicated that these DE mRNAs might be associated with systemic lupus erythematosus, alcoholism, intestinal immune network for IgA production, inflammatory bowel disease, NF-kappa B signaling pathway. WGCNA and Pearson correlation analyses jointly identified 16,577 significantly correlated DE mRNA-lncRNA pairs. Furthermore, lncRNAs LINC00641, LINC02268, LINC03013 were identified as hub lncRNAs. The protein–protein interaction (PPI) network of proteins encoded by DE mRNAs was established, and PPI-based analysis revealed that Interleukin 2(IL2), CD80 molecule (CD80), interleukin-17A (IL-17A) might play significant roles in the development of STC. This study analyzes the expression profiles of lncRNAs and mRNAs associated with STC. Our findings will contribute to further understanding of the molecular mechanism of STC and provide potential diagnostic or therapeutic biomarkers for STC.

Identification of Plasma Exosomes hsa_circ_0001360 and hsa_circ_0000038 as Key Biomarkers of Coronary Heart Disease.

Coronary heart disease (CHD) is the leading cause of death and disability worldwide. Accumulating evidence reveals that atherosclerosis (AS), characterized by systemic, chronic, and multifocal disease, and is the primary pathological basis of cardiovascular diseases, including CHD. However, the molecular underpinnings of CHD are still far from well understood. Our study attempted to identify aberrant plasma exosome-derived circRNAs and key exosomal circRNA biomarkers for CHD. The expression profiles of mRNAs, circRNAs, and lncRNAs in the blood exosomes of CHD patients and healthy controls were obtained from the exoRBase database. The corresponding miRNAs of the differentially expressed mRNAs, circRNAs, and lncRNAs were predicted via ENCORI and the miRcode database. LncRNAs/circRNAs and mRNAs with the cotargeted miRNAs were selected to construct an interaction network. Multiple machine learning algorithms have been used to explore potential biomarkers, followed by verification in patients with CHD using real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR). Based on the cutoff criterion of P < 0.05, we identified 85 differentially expressed circRNAs (4 upregulated and 81 downregulated), 43 differentially expressed lncRNAs (24 upregulated and 19 downregulated), and 312 differentially expressed mRNAs (55 upregulated and 257 downregulated). Functional enrichment analysis revealed that the differentially expressed mRNAs were involved mainly in neutrophil extracellular trap (NET) formation and the nucleotide-binding oligomerization domain- (NOD-) like receptor signaling pathway. Further analysis revealed that the DEGs in the circRNA/lncRNA-miRNA-mRNA interaction network were closely related to lipid and atherosclerotic signaling pathways. Hsa_circ_0001360 and hsa_circ_0000038 were identified as potential biomarkers for CHD based on three machine learning algorithms. The relative expression levels of hsa_circ_0001360 and hsa_circ_0000038 were significantly altered in plasma exosomes from patients with CHD. ROC curve analysis revealed that the areas under the curve (AUCs) were 0.860, 0.870, and 0.940 for hsa_circ_0001360, hsa_circ_0000038, and the two-gene combination, respectively. The circRNA/lncRNA-miRNA-mRNA interaction network might help to elucidate the pathogenesis of CHD. Hsa_circ_0001360 combined with hsa_circ_0000038 might be an important diagnostic biomarker.

Abstract 5249: Macrophages expressing synthetic cytokine receptors reverse IL10-mediated immunosuppression within solid tumors and promote adaptive immunity

Abstract Motivation: In solid tumors, anti-inflammatory cytokines such as IL10 and TGFβ support an immunosuppressive tumor microenvironment (TME) and inhibit anti-tumor immunity. Many therapeutic approaches to overcome immunosuppression have used monoclonal antibodies to block cytokine signaling. Instead of silencing TME-associated cytokines, a more powerful strategy may be to leverage them as disease markers and convert them into anti-tumor signals, using a logic-gated cell-based immunotherapy. Here, we engineered macrophages with synthetic cytokine switch receptors (SR) that convert IL10 or TGFβ into pro-inflammatory signals. Macrophages are homeostatic regulators capable of both infiltrating solid tumors and initiating inflammation, and we harnessed this proficiency using SRs that convert prevalent cytokines in the TME into pro-inflammatory responses for TME modulation. We termed this engineered myeloid cell platform “Engineered Microenvironment Converters” (EM-C) and evaluated its ability to overcome cytokine-mediated immunosuppression in solid tumors. Methods: EM-C were produced by transducing primary macrophages or monocytes with a SR that converts IL10 into an interferon-based signal. The response of EM-C to IL10 was monitored in vitro using phenotypic characterization of surface molecules, measurement of secreted factors, and mRNA profiling. The in vivo activity of EM-C was evaluated using a subcutaneous tumor model in immunocompetent mice. Murine EM-C were administered to tumor-bearing mice, and their ability to modulate the TME was monitored via immunophenotyping, secretome analysis, and single cell transcriptomics. To demonstrate modularity of the EM-C platform, additional SR were designed to convert TGFβ into interferon or toll-like receptor signals. Results: EM-C efficiently sequestered IL10 or TGFβ and upregulated pro-inflammatory markers, cytokines, and pathways in a dose-dependent manner in vitro. EM-C administered to tumor-bearing mice remodeled the TME immune compartment by increasing the abundance of CD8 T cells and reducing the presence of immunosuppressive Tregs. EM-C furthermore augmented the cytokine/chemokine profile of the TME in a manner that correlated with anti-tumor response. TME modulation by EM-C improved tumor control, as compared to non-engineered macrophages, and had additive efficacy with checkpoint blockade. Conclusion: We present a versatile immunotherapy platform that harnesses macrophages as “living converters” to locally augment inflammation in solid tumors. EM-C exhibit a modular ability to locally convert IL10 or TGFβ into pro-inflammatory signals without systemic cytokine antagonism. The EM-C platform enables development of target antigen-agnostic myeloid cell immunotherapies for overcoming immunosuppression in diverse solid tumors. Citation Format: Chris Sloas, Silvia Beghi, Yuhao Huangfu, Rehman Qureshi, Benjamin Schott, Michael Ball, Daniel Blumenthal, Thomas Condamine, Michael Klichinsky, Yumi Ohtani. Macrophages expressing synthetic cytokine receptors reverse IL10-mediated immunosuppression within solid tumors and promote adaptive immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5249.

Abstract 3984: Full-length TCR and BCR repertoire analysis, integrated with mRNA and surface proteins characterization, in a tumor mouse model

Abstract Full-length TCR and BCR analysis with single-cell multiomic assays provides an understanding of immune responses, offering insights into the complete receptor structure and functionality that is crucial for accurately assessing the diversity and specificity of immune repertoires. Studying mouse immune cell heterogeneity can be achieved by simultaneous mRNA and surface protein profiling. However, when coupled with full-length TCR and BCR analysis, this integrated approach provides a more comprehensive understanding of immune responses and cellular function in healthy and diseased conditions. In this study, we used the MC38 colon carcinoma model to study T and B cell heterogeneity and clonotype diversity. We stained single-cell suspensions prepared from bone marrow, thymus, spleen and tumor tissues with 20 AbSeq (CITE-Seq) antibody oligos and sample-multiplexing tags. We then loaded 50,000 isolated T and B cells into two lanes of an 8-lane microwell cartridge for single-cell capture by a single-cell analysis system. To aid in the simultaneous analysis of clonality and gene expression, we carried out library preparation using a full-length TCR/BCR multiomic assay and a mouse immune response panel. The combination of mRNA and protein expression confirmed the elevated expression of exhaustion and activation markers in tumor-infiltrating lymphocytes (TILs). We identified high-frequency clonotypes within TILs and tracked their frequency across other tissues. Tumor-burdened mice splenic T cells showed shared TCR combinations with TILs, which were absent in healthy tissue. The results show the ability to successfully profile T and B cell receptors using a full-length TCR/BCR multiomic assay in a mouse tumor model and the use of transcriptome, protein and TCR/BCR clonal information to investigate immune cell characteristics across multiple samples. For Research Use Only. Not for use in diagnostic or therapeutic procedures. BD, the BD Logo and BD Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2023 BD. All rights reserved. NPM-2536 (v1.0) 1023 Citation Format: Ali Rezvan, Xiaoshan Shi, Moen Sen, Stephanie Widmann, Aaron Tyznik. Full-length TCR and BCR repertoire analysis, integrated with mRNA and surface proteins characterization, in a tumor mouse model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3984.

Abstract 5694: Elucidating areca nut-induced miRNA-mRNA regulatory networks in head and neck cancer pathogenesis

Abstract Areca nut has been considered a high-risk carcinogen for head and neck cancer (HNC) in Southeast Asia, while its molecular effects, especially on the miRNA regulatory roles, remain elusive. This study characterized miRNA-mRNA networks associated with areca nut-related HNC. We integratively analyzed the areca nut-induced mRNA profile and TCGA-HNSC gene set. A total of 2575 genes were identified, with 1971 upregulated and 604 downregulated in cancers. The KEGG pathway analysis revealed that these genes were enriched in the cellular functions related to motility and stress response. Experimental studies confirmed that arecoline promoted cell migration, invasion, and chemo-radioresistance. We further cross-analyzed the miRNAs induced by areca nut and dysregulated in TCGA-HNSC data and defined 71 molecules affected, with 35 upregulation and 36 downregulation. Using the multimiR tool to map potential mRNA-miRNA interactions, we uncovered that these miRNAs were connected to 2173 genes. These molecules participated in several oncogenic mechanisms, such as ErbB and PI3K-Akt signaling, cell adhesion, and DNA damage response. We endeavored to establish miRNA-mRNA interaction networks with the functions of cellular motility and stress response via the Cytoscape analytical method. Our data revealed nexus comprising 41 miRNAs with 83 mRNAs, and 38 miRNAs with 66 mRNAs, respectively, co-regulate these two malignant functions. A key miRNA, miR-410-3p, downregulated in HNC and decreased following areca nut treatment, was studied. Ectopic expression of miR-410-3p attenuated areca nut-induced cell invasion and exerted modulatory effects on its target oncogenes, CRK. Thus, our study delineates a comprehensive view of the miRNA-mRNA interplay altered by areca nut, illustrating a sophisticated mechanism of carcinogenesis and suggesting new avenues for diagnostic and therapeutic strategies in HNC. Citation Format: Hung-Han Huang, Guo-Rung You, Kai-Hsin Lin, Yin-Ju Chen, Ann-Joy Cheng. Elucidating areca nut-induced miRNA-mRNA regulatory networks in head and neck cancer pathogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5694.

Abstract 266: The effects of retinoid agents and retinoic acid receptor genotype on proliferation, differentiation, and stem cells in colorectal cancer

Abstract Understanding how to therapeutically target colonic stem cells (SCs) is important because SC overpopulation drives CRC growth and development. Our goal is to therapeutically sensitize cancer SCs (CSCs) to the differentiation-inducing effects of retinoid agents. We previously found that ALDH specifically marks normal and malignant colonic SCs. We also found that retinoic acid (RA) receptors and other RA signaling components are selectively expressed in ALDH+ SCs, indicating that RA signaling occurs through ALDH+ SCs. Because all-trans retinoic acid (ATRA) is a highly effective treatment for acute promyelocytic leukemia, which is caused by a genetic translocation involving retinoic acid receptor alpha (RARA), we conjecture that the response of RA agents may be effective against solid tumors depending on their pathway genotype. Our bioinformatics analysis on mutations and overexpression of RA signaling component genes in CRC showed that most of the RA pathway genes are altered in human CRCs. Therefore, we hypothesize that the ability of retinoid agents and retinoid metabolism blocking agents to induce differentiation of CSCs depends on the RA pathway genotype of CRC cells.Testing the effect of various retinoid drugs on cell proliferation shows that RA pathway genotype affects the response of RA agents in CRC cell lines. HCT116 and SW480 CRC cell lines, which have RA receptor mutations, displayed resistance to ATRA. In contrast, HT29 CRC cells that have wild-type RA receptors are sensitive to ATRA. All three cell lines however showed similar dose responses to the ATRA metabolizing enzyme, CYP26A1 inhibitor Liarozole. This finding indicates that regardless of RA receptor mutations, inhibition of ATRA metabolism can increase intracellular RA levels in a similar manner on all three cell lines to induce anti-proliferative effects. We used CRISPR-Cas9 gene editing to knock out the RARA gene in HT29 cells and evaluated its response to ATRA compared to HT29 cells. The RARA knockout derived lines showed reduced sensitivity to ATRA treatment and a slower rate of cell division compared to HT29 control cells. Additionally, Nanostring mRNA profiling analysis shows that ATRA treatment of HT29 cells increases expression of RARA, CYP26A1, and KRT20 (differentiation marker); and decreased decreases expression of CSC markers LGR5 and ALDH1A1. Altogether, these findings indicate that upregulated RA signaling can reduce cell proliferation, increase differentiation, and decrease CSC numbers in CRCs. Overall, our findings indicate that colon SCs are regulated by RA signaling mechanisms and dysregulation of RA signaling contributes to the SC overpopulation that drives CRC growth. Thus, therapeutically sensitizing CSCs to differentiation-inducing effects of retinoid agents should provide insight into designing new SC-targeted therapies for CRC. Citation Format: Victoria O. Hunsu, Caroline O. Facey, Bruce M. Boman. The effects of retinoid agents and retinoic acid receptor genotype on proliferation, differentiation, and stem cells in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 266.

Abstract 5562: Altered tumor microenvironment in animal model of concomitant GBM and Alzheimer's pathology

Abstract Glioblastoma (GBM) and Alzheimer’s disease (AD) are two devastating central nervous system diagnoses with no known cure. Both diseases are associated with advanced age. The majority of GBM tumors form in the frontal and temporal lobes, two of the brain regions most impacted by amyloid and tau pathology in AD. Females have a higher risk of AD, while males have a higher risk of GBM. Several large case series have established epidemiological evidence that the two diseases are inversely correlated. Nevertheless, it remains unknown whether presence of subclinical amyloid or tau pathology antagonizes the establishment or progression of GBM. The present study sought to characterize GBM growth, progression, and immune response in the setting of concomitant AD pathology using the syngeneic murine GL261 tumor model. To further understand the tumor cell changes that occur in the presence or absence of AD pathology, we engineered GL261 with a dual-tagged vector consisting of RiboTag (to allow for full-length mRNA profiling) and TurboID (to allow for proteomic profiling). GL261 cells were injected intracranially into APPNL-G-F/NL-G-F - MAPThuman/human, APPNL-G-F/+ - MAPThuman/+, and wildtype C57BL/6J mice. Tumors in both wildtype and AD conditions contained ~50% intratumoral myeloid cells (Iba+, P2RY12-). Differential activation of intratumoral myeloid cells will be assessed with immunohistochemistry. Microgliosis (IBA1, P2RY12) and astrogliosis (GFAP) around the tumor in the setting of Alzheimer’s pathology versus normal conditions will be compared. Invasiveness, proliferation, DNA damage (pH2AX), and apoptosis (cleaved PARP, cleaved Caspase 3) will be compared. Tumor cell-specific changes unique to tumors in the AD microenvironment will be assessed transcriptionally with RiboTag profiling as well as proteomically with TurboID purifications. Our preliminary data suggests that the presence of concomitant AD pathology affects GBM growth and progression as well as the immune response to the tumor, and warrants further investigation. Citation Format: David Nascari, Ryan Eghlimi, Angad Beniwal, Drake Alton, John Fryer, Nhan L. Tran. Altered tumor microenvironment in animal model of concomitant GBM and Alzheimer's pathology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5562.

Abstract 5452: AXIN2 differentially modulates stem cell subpopulations providing a potential therapeutic opportunity for targeting colorectal cancer stem cells

Abstract Constitutively activated WNT-signaling promotes stem cell (SC) overpopulation which drives most colorectal cancer (CRC) development. It is not known, however, how WNT-signaling activity regulates different SC sub-populations. We previously reported that SC population size decreases as AXIN2 is knocked down, and increases when AXIN2 is overexpressed. Changes in cell cycle and apoptotic rate were also observed. Here, we present our recent results on changes between different SC subpopulations (ALDH+ vs. LGR5+) that occur due to AXIN2 modulation. We hypothesize that modulation of AXIN2 promotes differential expansion between different CRC SC subpopulations (ALDH+ and LGR5+). We observed that AXIN2 not only increases LGR5+ SC population size, but also expands the ALDH+ SC population to a greater extent as shown by an increased ALDH+-to-LGR5+ SCs ratio. This indicates that AXIN2, via WNT signaling, regulates SC population size, particularly ALDH+ SCs. Since the ALDH SC marker is also a key component in the retinoic acid (RA) signaling pathway, we surmised that increased AXIN2 would not only lead to elevated ALDH levels, but also upregulate the RA-signaling pathway components in CRC cells. To this end, we conducted NanoString mRNA profiling to study the effects of AXIN2 knockdown and overexpression in HT29 and SW480 CRC cells. The results showed that increased AXIN2 significantly up-regulated several key components of the RA-signaling pathway. Taken together, our results indicate that AXIN2 drives a shift in SC subpopulation sizes favoring the ALDH+ SCs. It also promotes the RA-signaling component expression - a pathway known to regulate SC differentiation. As a potential treatment strategy for CRC, therapeutically inducing high levels of AXIN2 could shift the balance between SC populations towards ALDH+ SCs, which may provide a way to target cancer SCs with retinoids to induce their differentiation. Citation Format: Chi Zhang, Caroline O. Facey, Lynn M. Opdenaker, Bruce M. Boman. AXIN2 differentially modulates stem cell subpopulations providing a potential therapeutic opportunity for targeting colorectal cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5452.

Abstract 3519: Reconstructing a latent representation of gene expression from genomic alterations to improve clinical utility of real-world clinicogenomics data

Abstract Background: Molecularly and clinically well-annotated patient datasets are ideal for studying tumor biology and developing robust machine learning (ML) models for predicting outcome and treatment response. These data however rarely exist in real-world settings or in sufficient quantities within research contexts. Large publicly available datasets like The Cancer Genome Atlas (TCGA) which provid multi-omic profiles for diverse cancer types, have profoundly advanced cancer research and facilitated development of novel therapies and personalized medicines. However, the absence of patient outcome data tied to treatment limits the applicability of these data for understanding and modeling treatment response. Real-world clinicogenomics cohorts, such as the AACR Project GENIE, on the other hand are typically very rich in clinical annotations, including treatment regimens and outcomes measures. These data are however sparsely annotated for patient tumor molecular profiles, rarely exceeding ~100’s of genes profiled. We hypothesized that it would be possible to reconstruct latent tumor mRNA representations from limited genomic and clinical data available in real-world clinicogenomic cohorts, and that these reconstructed expression profiles would be useful for a variety of clinically meaningful downstream applications. Methods: We developed an ML model, called Mut2Ex, to reconstruct tumor gene expression profiles using genetic information available on commercial next generation sequencing panels using a Principle Label Space Transformation (PLST) we adapted to regression problem, along with embeddings from clinical information (OncoTree code, sex and stage) generated by a language model. Mut2Ex was trained on ~1200 cell lines from DepMap representing 26 cancer types, to generate ~2000 reconstructed mRNA gene profiles that were applied to a variety of clinical tasks. We used Mut2Ex to reconstruct mRNA profiles for ~10,000 tumors from TCGA and ~180,000 tumors from AACR Project GENIE. Results: Reconstructed mRNA expression by Mut2Ex was highly correlated with true expression in cell lines (rho = 0.926, [0.924-0.928 95% CI, N = 1184]). Compared to true expression, reconstructed profiles recapitulate sub-clusters within cancer types, PAM50 subtyping in breast tumors, survival signatures in colorectal tumors and multiple oncogenic signatures in a pan-cancer manner. Analysis of reconstructed expression for AACR Project GENIE tumors revealed expected enrichment of known driver genes within expression subtypes and enrichment of oncogenic signatures associated with distinct clinical outcomes in a cancer type specific manner. Conclusions: Our flexible analytic framework for reconstructing gene expression profiles from clinicogenomics data substantially augments the clinical utility and value of data acquired in real-world settings. Citation Format: Maayan Baron, Sunil Kumar, Felicia Kuperwaser, Dillon Tracy, Emily Vucic, Jeff Sherman. Reconstructing a latent representation of gene expression from genomic alterations to improve clinical utility of real-world clinicogenomics data [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3519.

Green synthesis of CuO nanoparticle using Cyperus rotundus and evaluation of their anticancer potentials against lung cancer cells invitro

Copper oxide nanoparticles (CuONps) are known to cause oxidative stress and disrupt normal cellular functions. Due to the negative consequences of chemically manufactured CuONps, the current work focused on the biogenic sol-gel method of CuONps synthesis utilizing Cyperus rotundus as a biocatalyst (CyrCuONps) and the investigation of their anticancer potentials against lung cancer cells. The FTIR and EDAX analysis of synthesized Nps confirms their identification whereas XRD spectrum authorizes the purity of the synthesized Nps. SEM and TEM images show that the CyrCuONps were spherical in shape and had consistent size. The plant extract was shown to have significant antioxidant activity (56.92 %). Cancer cells treated with CyrCuONps exhibited cell shrinking, detachment, membrane blebbing and distorted shape. The MTT experiment confirmed CyrCuONps' cytotoxic capability against cancer cells by significantly lowering cell viability (23.01%) and suppressing cell growth (72.6%). The apoptotic assay revealed the presence of fragmented nuclei with compacted chromatin in treated cells, demonstrating genotoxicity caused by CyrCuONps exposure. In CyrCuO-Nps-treated cells, mRNA profiling indicated a 42% downregulation of mTOR and 56.94% of p13K genes, hence proving the ability of CyrCuONps in causing an aberration in the prognosis of cancer cells. The combined action of different bioactive chemicals in the plant extract and direct toxicities produced by CuONps attributes to their multifaced supremacy in cancer control, and because they are biogenic, they may be considered safe for anticancer treatment in humans.

Nonsense-mediated mRNA decay of mRNAs encoding a signal peptide occurs primarily after mRNA targeting to the endoplasmic reticulum

Translation of mRNAs encoding integral membrane proteins or secreted proteins occurs on the surface of the endoplasmic reticulum (ER). When a nascent signal peptide is synthesized from the mRNAs, the ribosome-nascent chain complex (RNC) is recognized by the signal recognition particle (SRP) and then transported to the surface of the ER. The appropriate targeting of the RNC–SRP complex to the ER is monitored by a quality control pathway, a nuclear cap-binding complex (CBC)-ensured translational repression of RNC–SRP (CENTRE). In this study, using ribosome profiling of CBC-associated and eIF4E-associated mRNAs, we reveal that, at the transcriptomic level, CENTRE is in charge of the translational repression of the CBC–RNC–SRP until the complex is specifically transported to the ER. We also find that CENTRE inhibits the nonsense-mediated mRNA decay (NMD) of mRNAs within the CBC–RNC–SRP. The NMD occurs only after the CBC–RNC–SRP is targeted to the ER and after eIF4E replaces CBC. Our data indicate dual surveillance for properly targeting mRNAs encoding integral membrane or secretory proteins to the ER. CENTRE blocks gene expression at the translation level before the CBC–RNC–SRP delivery to the ER, and NMD monitors mRNA quality after its delivery to the ER.

Transcriptomic sequencing analysis of key long noncoding RNAs and mRNAs expression profiles in postoperative recurrence of hepatocellular carcinoma.

Recurrence is the main cause of death in hepatocellular carcinoma (HCC) patients after liver resection. The long non-coding RNAs (lncRNAs) have been reported participated in progression and prognosis of HCC, however, the vital role of lncRNA in postoperative recurrence of HCC has rarely been systematically identified. RNA-sequencing (RNA-seq) was performed between orthotopic model of HCC and hepatoma postoperative recurrent model to comprehensively analyze the integrated transcriptome expression profiles of lncRNA and mRNA. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) was then conducted to quantify the expression levels of DElncRNAs and their target mRNAs. In our study, 211 lncRNAs (P-value < 0.05) and 1125 mRNAs (P-adjust < 0.05) were significantly differentially expressed (DE) between two groups. Moreover, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that DElncRNAs and DEmRNAs were mainly enriched in lipid metabolism, including Arachidonic acid metabolism, PPAR signaling pathway, Steroid hormone biosynthesis, Linoleic acid metabolism, Inflammatory mediator regulation of TRP channels, and Fatty acid degradation. Furthermore, we constructed lncRNA-mRNA interaction networks and protein-protein interaction (PPI) network, and verified by qRT-PCR, suggesting that increased DEIncRNAs (XLOC_063499 and XLOC_042016) may prevent HCC recurrence after surgery by upregulating on targeted cytochrome P450 (CYP) family genes in the lipid metabolism pathway, such as cyp3a16, cyp3a44, cyp2c39, cyp2c40 and cyp2c68. Overall, Our findings provided new insights for further investigation of biological function in lncRNA related HCC recurrence.

Plasma Circulating mRNA Profile for the Non-Invasive Diagnosis of Colorectal Cancer Using NanoString Technologies.

Colorectal cancer (CRC) is one of the most prevalent cancers and the second leading cause of cancer deaths in developed countries. Early CRC may have no symptoms and symptoms usually appear with more advanced diseases. Regular screening can identify people who are at increased risk of CRC in order to offer earlier treatment. A cost-effective non-invasive platform for the screening and monitoring of CRC patients allows early detection and appropriate treatment of the disease, and the timely application of adjuvant therapy after surgical operation is needed. In this study, a cohort of 71 plasma samples that include 48 colonoscopy- and histopathology-confirmed CRC patients with TNM stages I to IV were recruited between 2017 and 2019. Plasma mRNA profiling was performed in CRC patients using NanoString nCounter. Normalized data were analyzed using a Mann-Whitney U test to determine statistically significant differences between samples from CRC patients and healthy subjects. A multiple-group comparison of clinical phenotypes was performed using the Kruskal-Wallis H test for statistically significant differences between multiple groups. Among the 27 selected circulating mRNA markers, all of them were found to be overexpressed (gene expression fold change > 2) in the plasma of patients from two or more CRC stages. In conclusion, NanoString-based targeted plasma CRC-associated mRNAs circulating the marker panel that can significantly distinguish CRC patients from a healthy population were developed for the non-invasive diagnosis of CRC using peripheral blood samples.

Improving hematopoietic differentiation from human induced pluripotent stem cells by the modulation of Hippo signaling with a diarylheptanoid derivative

BackgroundThe diarylheptanoid ASPP 049 has improved the quality of adult hematopoietic stem cell (HSC) expansion ex vivo through long-term reconstitution in animal models. However, its effect on hematopoietic regeneration from human induced pluripotent stem cells (hiPSCs) is unknown.MethodWe utilized a defined cocktail of cytokines without serum or feeder followed by the supplementation of ASPP 049 to produce hematopoietic stem/progenitor cells (HSPCs). Flow cytometry and trypan blue exclusion analysis were used to identify nonadherent and adherent cells. Nonadherent cells were harvested to investigate the effect of ASPP 049 on multipotency using LTC-IC and CFU assays. Subsequently, the mechanism of action was explored through transcriptomic profiles, which were validated by qRT-PCR, immunoblotting, and immunofluorescence analysis.ResultThe supplementation of ASPP 049 increased the number of phenotypically defined primitive HSPCs (CD34+CD45+CD90+) two-fold relative to seeded hiPSC colonies, indicating enhanced HSC derivation from hiPSCs. Under ASPP 049-supplemented conditions, we observed elevated HSPC niches, including CD144+CD73− hemogenic- and CD144+CD73+ vascular-endothelial progenitors, during HSC differentiation. Moreover, harvested ASPP 049-treated cells exhibited improved self-renewal and a significantly larger proportion of different blood cell colonies with unbiased lineages, indicating enhanced HSC stemness properties. Transcriptomics and KEGG analysis of sorted CD34+CD45+ cells-related mRNA profiles revealed that the Hippo signaling pathway is the most significant in responding to WWTR1/TAZ, which correlates with the validation of the protein expression. Interestingly, ASPP 049-supplemented HSPCs upregulated 11 genes similarly to umbilical cord blood-derived HSPCs.ConclusionThese findings suggest that ASPP 049 can improve HSC-generating protocols with proliferative potentials, self-renewal ability, unbiased differentiation, and a definable mechanism of action for the clinical perspective of hematopoietic regenerative medicine.Graphical abstract

A genome-wide study of gastric intramucosal neoplasia based on somatic copy number alterations, gene mutations, and mRNA expression patterns.

We performed comprehensive analyses of somatic copy number alterations (SCNAs) and gene expression profiles of gastric intramucosal neoplasia (IMN) using array-based methods in 97 intestinal-type IMNs, including 39 low-grade dysplasias (LGDs), 37 high-grade dysplasias (HGDs), and 26 intramucosal carcinomas (IMCs) with stromal invasion of the lamina propria to identify the molecular mechanism of IMN. In addition, we examined gene mutations using gene panel analyses. We used cluster analyses for exclusion of arbitrariness to identify SCNA patterns and expression profiles. IMNs were classified into two distinct subgroups (subgroups 1 and 2) based on SCNA patterns. Subgroup 1 showed a genomic stable pattern due to the low frequency of SCNAs, whereas subgroup 2 exhibited a chromosomal instability pattern due to the high frequencies of SCNAs and TP53 mutations. Interestingly, although the frequencies of LGD and HGD were significantly higher in subgroup 1 than in subgroup 2, IMC was commonly found in both types. Although the expression profiles of specific mRNAs could be used to categorise subgroups 1 and 2, no clinicopathological findings correlated with either subgroup. We examined signalling pathways specific to subgroups 1 and 2 to identify the association of each subgroup with signalling pathways based on gene ontology tree visualisation: subgroups 1 and 2 were associated with haem metabolism and chromosomal instability, respectively. These findings reveal a comprehensive genomic landscape that highlights the molecular complexity of IMNs and provide a road map to facilitate our understanding of gastric IMNs.

Differential mRNA profiles reveal the potential roles of genes involved in lactate stimulation in mouse macrophages

Lactate is a glycolysis end product, and its levels are markedly associated with disease severity, morbidity, and mortality in sepsis. It modulates key functions of immune cells, including macrophages. In this investigation, transcriptomic analysis was performed using lactic acid, sodium lactate, and hydrochloric acid-stimulated mouse bone marrow-derived macrophages (iBMDM), respectively, to identify lactate-associated signaling pathways. After 24 h of stimulation, 896 differentially expressed genes (DEG) indicated were up-regulation, whereas 792 were down-regulated in the lactic acid group, in the sodium lactate group, 128 DEG were up-regulated, and 41 were down-regulated, and in the hydrochloric acid group, 499 DEG were up-regulated, and 285 were down-regulated. Subsequently, clinical samples were used to further verify the eight genes with significant differences, among which Tssk6, Ypel4, Elovl3, Trp53inp1, and Cfp were differentially expressed in patients with high lactic acid, indicating their possible involvement in lactic acid-induced inflammation and various physiological diseases caused by sepsis. However, elongation of very long chain fatty acids protein 3 (Elovl3) was negatively correlated with lactic acid content in patients. The results of this study provide a necessary reference for better understanding the transcriptomic changes caused by lactic acid and explain the potential role of high lactic acid in the regulation of macrophages in sepsis.

Malignancy-associated ischemic stroke: Implications for diagnostic and therapeutic workup.

Patients with malignancies have an increased risk of suffering ischemic stroke via several mechanisms such as coagulation dysfunction and other malignancy-related effects as well as iatrogenic causes. Moreover, stroke can be the first sign of an occult malignancy, termed as malignancy-associated ischemic stroke (MAS). Therefore, timely diagnostic assessment and targeted management of this complex clinical situation are critical. Patients with both stroke and malignancy have atypical ages, risk factors, and often exhibit malignancy-related symptoms and multiple lesions on neuroimaging. New biomarkers such as eicosapentaenoic acid and blood mRNA profiles may help in distinguishing MAS from other strokes. In terms of treatment, malignancy should not be considered a contraindication, given comparable rates of recanalization and complications between stroke patients with or without malignancies. In this review, we summarize the latest developments in diagnosing and managing MAS, especially stroke with occult malignancies, and provide new recommendations from recently emerged clinical evidence for diagnostic and therapeutic workup strategies.

Combined analysis of mRNA and miRNA transcriptomes reveals the regulatory mechanism of Xanthomonas arboricola pv pruni resistance in Prunus persica

BackgroundPeach bacterial shot hole, caused by Xanthomonas arboricola pv pruni (Xap), is a global bacterial disease that poses a threat to the yield and quality of cultivated peach trees (Prunus persica).ResultsThis study compared the mRNA and miRNA profiles of two peach varieties, ‘Yanbao’ (resistant) and ‘Yingzui’ (susceptible), after inoculation with Xap to identify miRNAs and target genes associated with peach tree resistance. mRNA sequencing results revealed that in the S0-vs-S3 comparison group, 1574 genes were upregulated and 3975 genes were downregulated. In the R0-vs-R3 comparison group, 1575 genes were upregulated and 3726 genes were downregulated. Through miRNA sequencing, a total of 112 known miRNAs belonging to 70 miRNA families and 111 new miRNAs were identified. Notably, some miRNAs were exclusively expressed in either resistant or susceptible varieties. Additionally, 59 miRNAs were downregulated and 69 miRNAs were upregulated in the R0-vs-R3 comparison group, while 46 miRNAs were downregulated and 52 miRNAs were upregulated in the S0-vs-S3 comparison group. Joint analysis of mRNA and miRNA identified 79 relationship pairs in the S0-vs-S3 comparison group, consisting of 48 miRNAs and 51 target genes. In the R0-vs-R3 comparison group, there were 58 relationship pairs, comprising 28 miRNAs and 20 target genes. Several target genes related to resistance, such as SPL6, TIFY6B, and Prupe.4G041800_v2.0.a1 (PPO), were identified through literature reports and GO/KEGG enrichment analysis.ConclusionIn conclusion, this study discovered several candidate genes involved in peach tree resistance by analyzing differential expression of mRNA and miRNA. These findings provide valuable insights into the mechanisms underlying resistance to Xap in peach trees.