Gene Expression Patterns In Cells Research Articles

Immunotherapy and pan-apoptotic characterization of the tumor microenvironment in gastric cancer (STAD): a single-cell multidimensional analysis

BackgroundThe aim of this study was to elucidate the critical role of autophagy-related gene aggregation in gastric cancer tumor microenvironment cells and to investigate their major roles in cellular functions. In particular, the expression of these genes in tumor-associated fibroblast subtypes was scrutinized in an attempt to explain their cell-subpopulation-specific roles in cell–cell communication and regulation of cellular functions.MethodsIn this study, single-cell RNA sequencing data were first analyzed in multiple steps, including data preprocessing, cell clustering, and cell classification. Cell subpopulations and gene expression patterns were identified and analyzed using unsupervised non-negative matrix factorization (NMF) techniques. The dynamic expression of autophagy-related gene aggregates in various cell types was deciphered by pseudotime trajectory analysis (PTA). Intercellular communication analysis was performed using the CellChat R software package, revealing the intricate communication patterns and exchange of key signaling molecules between cell subpopulations, and SCENIC analysis was used to identify gene regulatory networks and reveal the mechanisms behind cellular heterogeneity.ResultCell subpopulations associated with pan-apoptosis were identified by NMF decomposition and SCENIC analysis. Cell–cell communication analysis revealed intricate communication patterns and exchange of key signaling molecules between cell subpopulations. Dynamic expression of autophagy-related genes aggregated in the pseudotemporal trajectory of STAD was observed by PTA. In the fibroblast subtype, different ligand-receptor interactions and their key roles in immunomodulation were observed.ConclusionBy deeply analyzing and comparing gene expression patterns within cellular subpopulations and intercellular communication, this study provides new insights into the role of pan-apoptosis-related genes in regulating immune responses and cellular functions in gastric cancer. These findings pave the way for further exploration of the role of these genes in tumorigenesis and immune regulation, as well as laying the foundation for potential therapeutic strategies.

Gene expression levels in cumulus cells are correlated with developmental competence of bovine oocytes

The generation of mammalian embryos by in vitro culture is hampered by the failure of many of the embryos to develop to the blastocyst stage. This problem occurs even when cumulus–oocyte complexes (COCs) with good morphology are visually selected and used for culture. Because cumulus cells are important for oocyte maturation and subsequent embryo development, here we compared gene expression patterns in cumulus cells of COCs that developed in vitro to the blastocyst stage with those of COCs that failed to develop. Cumulus cells were aspirated from bovine COCs selected for in vitro culture. Oocyte developmental competence was evaluated by screening for cleavage and development to the blastocyst stage. The collected cumulus cells were used to quantify mRNA levels of FSH receptor (FSHR), insulin-like growth factor-1 receptor (IGF-1R), anti-Müllerian hormone (AMH), AMH receptor II (AMHRII), epidermal growth factor receptor (EGFR), estrogen receptor β (ERβ), B cell lymphoma/leukemia-2 associated X (Bax), and cysteine-aspartic acid protease-3 (Caspase-3). We found that the expression levels of FSHR, IGF-1R, AMH, and EGFR were higher in cumulus cells from COCs that developed to blastocysts as compared with those that failed to develop, whereas expression levels of Bax and Caspase-3 were lower in cumulus cells of COCs that matured to the blastocyst stage. Positive correlations were found between FSHR and IGF-1R expression (r = 0.59) and between ERβ and EGFR expression (r = 0.43) in cumulus cells from COCs that developed to the blastocyst stage. Our findings indicate that gene expression levels in cumulus cells are correlated with the developmental competence of bovine oocytes. Measurement of gene expression in cumulus cells therefore offers a non-invasive means of predicting oocyte developmental competence.

Epigenomic, transcriptomic and proteomic characterizations of reference samples.

A variety of newly developed next-generation sequencing technologies are making their way rapidly into the research and clinical applications, for which accuracy and cross-lab reproducibility are critical, and reference standards are much needed. Our previous multicenter studies under the SEQC-2 umbrella using a breast cancer cell line with paired B-cell line have produced a large amount of different genomic data including whole genome sequencing (Illumina, PacBio, Nanopore), HiC, and scRNA-seq with detailed analyses on somatic mutations, single-nucleotide variations (SNVs), and structural variations (SVs). However, there is still a lack of well-characterized reference materials which include epigenomic and proteomic data. Here we further performed ATAC-seq, Methyl-seq, RNA-seq, and proteomic analyses and provided a comprehensive catalog of the epigenomic landscape, which overlapped with the transcriptomes and proteomes for the two cell lines. We identified >7,700 peptide isoforms, where the majority (95%) of the genes had a single peptide isoform. Protein expression of the transcripts overlapping CGIs were much higher than the protein expression of the non-CGI transcripts in both cell lines. We further demonstrated the evidence that certain SNVs were incorporated into mutated peptides. We observed that open chromatin regions had low methylation which were largely regulated by CG density, where CG-rich regions had more accessible chromatin, low methylation, and higher gene and protein expression. The CG-poor regions had higher repressive epigenetic regulations (higher DNA methylation) and less open chromatin, resulting in a cell line specific methylation and gene expression patterns. Our studies provide well-defined reference materials consisting of two cell lines with genomic, epigenomic, transcriptomic, scRNA-seq and proteomic characterizations which can serve as standards for validating and benchmarking not only on various omics assays, but also on bioinformatics methods. It will be a valuable resource for both research and clinical communities.

Transcriptome analysis of the effect of HERV-K env gene knockout in ovarian cancer cell lines.

Human endogenous retroviruses (HERVs) have been implicated in the pathogenesis of various diseases, particularly cancers. Previous investigations from our group demonstrated that targeted knockout (KO) of the HERV-K env gene led to a significant reduction in tumorigenic attributes, including proliferation, migration, and invasion of ovarian cancer cells. In this study, we aimed to elucidate the impact of HERV-K env KO on gene expression in ovarian cancer cell lines through comparative RNA sequencing (RNA-Seq) analysis with two distinct HERV-K env KO ovarian cancer cell lines, SKOV3 and OVCAR3. HERV-K env gene KO was achieved in SKOV3 and OVCAR3 ovarian cancer cell lines using the CRISPR-Cas9 system. Next-generation mRNA sequencing was employed to assess the gene expression profiles of both mock and HERV-K env KO ovarian cancer cells. Furthermore, comprehensive analyses involving gene ontology and pathway assessments were conducted. Transcriptome analysis revealed that 23 differentially expressed genes (DEGs) were upregulated and 17 DEGs were downregulated in SKOV3 cells. In OVCAR3 cells, 198 DEGs were upregulated, and 17 DEGs were downregulated. Notably, 53 DEGs exhibited statistically significant differences among the 1,612 DEGs identified. Our findings indicate that HERV-K env gene KO exerts a profound influence on gene expression patterns in OVCAR3 cells, while genetic alterations in expression were relatively modest in SKOV3 cells. Nevertheless, genes ND1, ND2, and CYTB displayed a common increase in expression, while ERRFI1 and NDRG1 exhibited a decrease in expression in both cell lines. Our study demonstrates that KO of the HERV-K env gene in ovarian cancer cell lines has a substantial impact on gene expression patterns and can be used to identify potential therapeutic targets for ovarian cancer and related diseases.

Function Analysis of a Maize Endo-1,4-β-xylanase Gene ZmHSL in Response to High-Temperature Stress.

Rising temperature is a major threat to the normal growth and development of maize, resulting in low yield production and quality. The mechanism of maize in response to heat stress remains uncertain. In this study, a maize mutant Zmhsl-1 (heat sensitive leaves) with wilting and curling leaves under high temperatures was identified from maize Zheng 58 (Z58) mutant lines generated by ethyl methanesulfonate (EMS) mutagenesis. The Zmhsl-1 plants were more sensitive to increased temperature than Z58 in the field during growth season. The Zmhsl-1 plants had lower plant height, lower yield, and lower content of photosynthetic pigments. A bulked segregant analysis coupled with whole-genome sequencing (BSA-seq) enabled the identification of the corresponding gene, named ZmHSL, which encodes an endo-β-1,4-xylanase from the GH10 family. The loss-of-function of ZmHSL resulted in reduced lignin content in Zmhsl-1 plants, leading to defects in water transport and more severe leaf wilting with the increase in temperature. RNA-seq analysis revealed that the differentially expressed genes identified between Z58 and Zmhsl-1 plants are mainly related to heat stress-responsive genes and unfolded protein response genes. All these data indicated that ZmHSL plays a key role in lignin synthesis, and its defective mutation causes changes in the cell wall structure and gene expression patterns, which impedes water transport and confers higher sensitivity to high-temperature stress.

Common lizard primary oviduct cell culture: A model system for the genetic and cellular basis of oviparity and viviparity

Reproduction by egg-laying (oviparity) or live-bearing (viviparity) is a genetically determined trait fundamental to the biology of amniotes. Squamates are an emerging model for the genetics of reproductive mode yet lack cell culture models valuable for exploring molecular mechanisms. Here, we report a novel primary culture model for reproductive biology: cell cultures derived from the oviduct tissues (infundibulum, uterus and vagina) of oviparous and viviparous common lizards (Lacertidae: Zootoca vivipara). We maintained and expanded these cultures for over 100 days, including repeated subculturing and successful revival of cryopreserved cells. Immunocytochemical investigation suggested expression of both epithelial and fibroblast-like proteins, and RNA sequencing of cultured cells as compared to in vivo oviduct tissue showed changes in gene expression in response to the cell culture environment. Despite this, we confirmed the maintenance of distinct gene expression patterns in viviparous and oviparous cells after 60+ days of cell culture, finding 354 differentially expressed genes between viviparous and oviparous cells. Furthermore, we confirmed the expression of 15 viviparity-associated candidate genes in cells maintained for 60+ days in culture. Our study demonstrates the feasibility and utility of oviduct cell culture for molecular analysis of reproductive mode and provides a tool for future genetic experiments.

Comparative transcriptome reveals EphA2 and c-Fos as key factors driving enhanced replication in high-passage porcine deltacoronavirus strain

Porcine deltacoronavirus (PDCoV), a cross-species transmissible enterovirus, frequently induces severe diarrhea and vomiting symptoms in piglets, which not only pose a significant menace to the global pig industry but also a potential public safety risk. In a previous study, we isolated a vaccine candidate, PDCoV CZ2020-P100, by passaging a parental PDCoV strain in vitro, exhibiting attenuated virulence and enhanced replication. However, the factors underlying these differences between primary and passaged strains remain unknown. In this study, we present the transcriptional landscapes of porcine kidney epithelial cells (LLC-PK1) cells infected with PDCoV CZ2020-P1 strain and P100 strain using the RNA-sequencing. We identified 105 differentially expressed genes (DEGs) in P1-infected cells and 295 DEGs in P100-infected cells. Enrichment analyses indicated that many DEGs showed enrichment in immune and inflammatory responses, with a more and higher upregulation of DEGs enriched in the P100-infected group. Notably, the DEGs were concentrated in the MAPK pathway within the P100-infected group, with significant upregulation in EphA2 and c-Fos. Knockdown of EphA2 and c-Fos reduced PDCoV infection and significantly impaired P100 replication compared to P1, suggesting a novel mechanism in which EphA2 and c-Fos are highly involved in passaged virus replication. Our findings illuminate the resemblances and distinctions in the gene expression patterns of host cells infected with P1 and P100, confirming that EphA2 and c-Fos play key roles in high-passage PDCoV replication. These results enhance our understanding of the changes in virulence and replication capacity during the process of passaging.

H3K27-Altered Diffuse Midline Glioma of the Brainstem: From Molecular Mechanisms to Targeted Interventions.

Pediatric high-grade gliomas are a devastating subset of brain tumors, characterized by their aggressive pathophysiology and limited treatment options. Among them, H3 K27-altered diffuse midline gliomas (DMG) of the brainstem stand out due to their distinct molecular features and dismal prognosis. Recent advances in molecular profiling techniques have unveiled the critical role of H3 K27 alterations, particularly a lysine-to-methionine mutation on position 27 (K27M) of the histone H3 tail, in the pathogenesis of DMG. These mutations result in epigenetic dysregulation, which leads to altered chromatin structure and gene expression patterns in DMG tumor cells, ultimately contributing to the aggressive phenotype of DMG. The exploration of targeted therapeutic avenues for DMG has gained momentum in recent years. Therapies, including epigenetic modifiers, kinase inhibitors, and immunotherapies, are under active investigation; these approaches aim to disrupt aberrant signaling cascades and overcome the various mechanisms of therapeutic resistance in DMG. Challenges, including blood-brain barrier penetration and DMG tumor heterogeneity, require innovative approaches to improve drug delivery and personalized treatment strategies. This review aims to provide a comprehensive overview of the evolving understanding of DMG, focusing on the intricate molecular mechanisms driving tumorigenesis/tumor progression and the current landscape of emerging targeted interventions.

The Advancement and Application of the Single-Cell Transcriptome in Biological and Medical Research.

Single-cell RNA sequencing technology (scRNA-seq) has been steadily developing since its inception in 2009. Unlike bulk RNA-seq, scRNA-seq identifies the heterogeneity of tissue cells and reveals gene expression changes in individual cells at the microscopic level. Here, we review the development of scRNA-seq, which has gone through iterations of reverse transcription, in vitro transcription, smart-seq, drop-seq, 10 × Genomics, and spatial single-cell transcriptome technologies. The technology of 10 × Genomics has been widely applied in medicine and biology, producing rich research results. Furthermore, this review presents a summary of the analytical process for single-cell transcriptome data and its integration with other omics analyses, including genomes, epigenomes, proteomes, and metabolomics. The single-cell transcriptome has a wide range of applications in biology and medicine. This review analyzes the applications of scRNA-seq in cancer, stem cell research, developmental biology, microbiology, and other fields. In essence, scRNA-seq provides a means of elucidating gene expression patterns in single cells, thereby offering a valuable tool for scientific research. Nevertheless, the current single-cell transcriptome technology is still imperfect, and this review identifies its shortcomings and anticipates future developments. The objective of this review is to facilitate a deeper comprehension of scRNA-seq technology and its applications in biological and medical research, as well as to identify avenues for its future development in alignment with practical needs.

Targeting bacterial biofilm-related genes with nanoparticle-based strategies.

Persistent infection caused by biofilm is an urgent in medicine that should be tackled by new alternative strategies. Low efficiency of classical treatments and antibiotic resistance are the main concerns of the persistent infection due to biofilm formation which increases the risk of morbidity and mortality. The gene expression patterns in biofilm cells differed from those in planktonic cells. One of the promising approaches against biofilms is nanoparticle (NP)-based therapy in which NPs with multiple mechanisms hinder the resistance of bacterial cells in planktonic or biofilm forms. For instance, NPs such as silver (Ag), zinc oxide (ZnO), titanium dioxide (TiO2), copper oxide (Cu), and iron oxide (Fe3O4) through the different strategies interfere with gene expression of bacteria associated with biofilm. The NPs can penetrate into the biofilm structure and affect the expression of efflux pump, quorum-sensing, and adhesion-related genes, which lead to inhibit the biofilm formation or development. Therefore, understanding and targeting of the genes and molecular basis of bacterial biofilm by NPs point to therapeutic targets that make possible control of biofilm infections. In parallel, the possible impact of NPs on the environment and their cytotoxicity should be avoided through controlled exposure and safety assessments. This study focuses on the biofilm-related genes that are potential targets for the inhibition of bacterial biofilms with highly effective NPs, especially metal or metal oxide NPs.

Particulate matter-induced gene expression patterns in human-derived cells based on 11 public gene expression datasets.

Exposure to particulate matter (PM) and house dust mite (HDM) can change the expression patterns of inflammation-, oxidative stress-, and cell death-related genes. We investigated the changes in gene expression patterns owing to PM exposure. This study examined the changes in gene expression patterns following PM exposure. We searched for differentially expressed genes (DEGs) following PM exposure using five cell line-based RNA-seq or microarray datasets and six human-derived datasets. The enrichment terms of the DEGs were assessed. DEG analysis yielded two gene sets. Thus, enrichment analysis was performed for each gene set, and the enrichment terms related to respiratory diseases were presented. The intersection of six human-derived datasets and two gene sets was obtained, and the expression patterns following PM exposure were observed. Two gene sets were obtained for cells treated with PM and their expression patterns were presented following verification in human-derived cells. Our findings suggest that exposure to PM2.5 and HDM may reveal changes in genes that are associated with diseases, such as allergies, highlighting the importance of mitigating PM2.5 and HDM exposure for disease prevention.

Claudin-10 Expression and the Gene Expression Pattern of Thick Ascending Limb Cells.

Many genomic, anatomical and functional differences exist between the medullary (MTAL) and the cortical thick ascending limb of the loop of Henle (CTAL), including a higher expression of claudin-10 (CLDN10) in the MTAL than in the CTAL. Therefore, we assessed to what extent the Cldn10 gene expression is a determinant of differential gene expression between MTAL and CTAL. RNAs extracted from CTAL and MTAL microdissected from wild type (WT) and Cldn10 knock out mice (cKO) were analyzed by RNAseq. Differential and enrichment analyses (GSEA) were performed with interactive R Shiny software. Between WT and cKO MTAL, 637 genes were differentially expressed, whereas only 76 were differentially expressed between WT and cKO CTAL. Gene expression patterns and GSEA analyses in all replicates showed that WT MTAL did not cluster with the other replicates; no hierarchical clustering could be found between WT CTAL, cKO CTAL and cKO MTAL. Compared to WT replicates, cKO replicates were enriched in Cldn16, Cldn19, Pth1r, (parathyroid hormone receptor type 1), Casr (calcium sensing receptor) and Vdr (Vitamin D Receptor) mRNA in both the cortex and medulla. Cldn10 is associated with gene expression patterns, including genes specifically involved in divalent cations reabsorption in the TAL.

Inorganic ions activate lineage-specific gene regulatory networks

Inorganic biomaterials have been shown to direct cellular responses, including cell−cell and cell−matrix interactions. Notably, ions released from these inorganic biomaterials play a vital role in defining cell identity, and promoting tissue-specific functions. However, the effect of inorganic ions on cellular functions have yet to be investigated at the transcriptomic level, representing a critical knowledge gap in the development of next-generation bioactive materials. To address this gap, we investigated the impact of various inorganic ions including silver, copper, titanium, and platinum on human mesenchymal stem cells (hMSCs). Our finding showed that silver and copper induce osteogenic and chondrogenic differentiation respectively, through enrichment of lineage-specific gene expression program. In particular, silver effectively induced Wingless/Integrated (Wnt) and mitogen-activated protein kinase (MAPK) signaling, which are vital for osteogenesis. On the other hand, copper specifically stimulated Transforming growth factor beta (TGFβ) signaling, while suppressing Janus kinase/signal transducers and activators of transcription (JAK-STAT) signaling, thereby promoting chondrogenesis. In contrast, platinum, and tantalum, ions didn't stimulate regenerative responses. Together, our findings highlight the potential of inorganic biomaterials in tissue regeneration strategies, which currently rely largely on growth factors and small molecule therapeutics. Statement of significanceThis research emphasizes the critical role of bioactive inorganic ions in controlling lineage-specific gene expression patterns in mesenchymal stem cells, effectively modulating the transcriptome landscape and directing cell fate. The study lays the foundation for a systematic database of biomaterial candidates and their effects on cellular functions, which will ultimately streamline the translation of new biomaterials into clinical applications.

A dynamic role for transcription factors in restoring transcription through mitosis.

Mitosis involves intricate steps, such as DNA condensation, nuclear membrane disassembly, and phosphorylation cascades that temporarily halt gene transcription. Despite this disruption, daughter cells remarkably retain the parent cell's gene expression pattern, allowing for efficient transcriptional memory after division. Early studies in mammalian cells suggested that transcription factors (TFs) mark genes for swift reactivation, a phenomenon termed 'mitotic bookmarking', but conflicting data emerged regarding TF presence on mitotic chromosomes. Recent advancements in live-cell imaging and fixation-free genomics challenge the conventional belief in universal formaldehyde fixation, revealing dynamic TF interactions during mitosis. Here, we review recent studies that provide examples of at least four modes of TF-DNA interaction during mitosis and the molecular mechanisms that govern these interactions. Additionally, we explore the impact of these interactions on transcription initiation post-mitosis. Taken together, these recent studies call for a paradigm shift toward a dynamic model of TF behavior during mitosis, underscoring the need for incorporating dynamics in mechanistic models for re-establishing transcription post-mitosis.

Abstract 2687: NK cell plasticity in SCLC is linked to tumor transcriptional diversity

Abstract Natural Killer (NK) cells are innate lymphoid cells that are pivotal in the body’s response to cancer. To harness their cytotoxicity in clinical interventions, it is critical to understand their biology, which is highly tissue- and tumor-specific. In Small Cell Lung Cancer (SCLC), reports have described widely divergent levels of NK activity in neuroendocrine (NE) and non-neuroendocrine (non-NE) subtypes. Diminished self-antigen presentation by MHC-I is a classic trigger of NK cell activity but, counter-intuitively, NK cells exhibit reduced cytotoxicity in MHC-I downregulated NE SCLC. To explore potential causes for depressed NK activity in NE SCLC relative to non-NE, we applied scRNA-Seq to 45 SCLC patient biopsies and rapid autopsies, and identified 4000 NK cells based on gene expression signature. There was a notable dip in NK cell infiltration in NE tumors compared to non-NE tumors. Tumor cell-surface ligands recognized by activating and, unexpectedly, inhibitory NK cell receptors were generally both more highly expressed in non-NE tumors. While inhibitory NK receptors were comparable in NE and non-NE tumors, activating NK receptors were expressed significantly, though modestly, higher in non-NE. We observed tumor site-specific gene expression patterns in NK cells from different metastatic sites. Overall, there were a remarkably high number of differentially expressed genes (>1k) between NK cells from NE and non-NE tumors, even after controlling for tumor site. Together, our work suggests that NK cell plasticity is manifest at the level of SCLC transcriptional subtypes. Citation Format: Justin Malin, Sophie Zhuang, Ajit Kumar Sharma, Joseph Clara, Anish Thomas. NK cell plasticity in SCLC is linked to tumor transcriptional diversity [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 2687.

Abstract 5323: Inflammation drives lineage plasticity and a club cell-like phenotype in prostate epithelial cells

Abstract Background: Club cells are a type of bronchiolar epithelial cell that serve a protective and antimicrobial function in the lung. An epithelial cell population in adult human prostates was recently identified as sharing a similar transcriptomic profile and morphology to lung club cells. Club-like epithelial cells were identified in the prostatic urethra, collecting ducts, and rarely in the peripheral zone of normal organ donor prostates. However, we recently reported that cells expressing club cell markers (CP, LTF, MMP7, PIGR, SCGB1A1) are present in the peripheral zone of radical prostatectomy specimens. Here, expression of club cell markers was restricted to luminal epithelial cells in an inflammation-associated lesion termed proliferative inflammatory atrophy (PIA). We also demonstrated that these club-like cells within PIA express the same markers as intermediate cells, a proposed prostate cancer progenitor cell. We now aim to identify exogenous factors that drive the club cell phenotype in vitro. Our ultimate goal is to understand how club cell gene expression affects the oncogenic and metastatic potential of prostate epithelial cells. Methods: We characterized the expression of club cell genes in prostate cell lines (LNCaP, PREC, and PREC-AR-Myc) following exposure to inflammatory stimuli (TNFα, hydrogen peroxide, uropathogenic Escherichia coli) in vitro. We performed quantitative reverse transcription PCR (RT-qPCR) to measure expression of club cell markers (LTF, MMP7, PIGR) and intermediate cell markers (AR, NKX3.1, SMOX). Additionally, we used chromogenic RNA in situ hybridization (RISH) to visualize LTF, MMP7, and PIGR expression following exposure to inflammatory stimuli. Results: We report that TNFα exposure in vitro increased expression of club cell genes (LTF, MMP7, PIGR). Surprisingly, TNFα exposure induced LTF expression in LNCaP cells, despite prior findings that LTF is silenced by CpG island hypermethylation in prostate cancer and in LNCaP cells. This finding was confirmed by both RT-qPCR and RISH. Additionally, we found that TNFα exposure altered intermediate cell markers (AR, NKX3.1, SMOX) expression consistent with the previously reported gene expression pattern of luminal epithelial cells in PIA. Conclusions: Overall, our results suggest that TNFα-mediated signaling drives the expression of club cell markers associated with lineage plasticity. Our next step will be to analyze inflammation-driven alterations to methylation and chromatin accessibility around loci associated with club cell genes, proton-oncogenes, and tumor suppressor genes. Citation Format: Megan Hess, Hanbing Song, Jessica Hicks, Luke Mummert, Angelo De Marzo, Franklin W. Huang, Karen Sfanos. Inflammation drives lineage plasticity and a club cell-like phenotype in prostate epithelial 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 5323.

Abstract 6622: Identification of early oncogenic lesions following concomitant Vhl and Pbrm1 loss in the murine kidney

Abstract Characterizing the early events preceding tumorigenesis is a major challenge in understanding cancer. To address this, we are investigating the paradigm of clear cell renal cell carcinoma (ccRCC) that is driven by the biallelic loss of the von Hippel Lindau (VHL) tumor suppressor gene. Though VHL loss is required, additional mutations have been found in the evolution of the cancer, most commonly in Polybromo-1 (PBRM1), a subunit of the PBAF SWI/SNF chromatin remodeling complex. Concomitant Vhl and Pbrm1 deletion, but not Vhl deletion alone, in the renal tubular epithelium (RTE) has been reported to be sufficient to drive the formation of renal tumors in mice. However, the early in vivo consequences of Vhl loss, and the exact requirement for Pbrm1 loss to tumorigenesis remain poorly studied. We have previously reported the development of a novel lineage-tracing model of Vhl deletion that directly couples loss of a conditional Vhl allele to the expression of a tdTomato reporter and allows accurate identification and retrieval of marked Vhl-null cells. In the work reported here, we have combined this system with conditional Pbrm1 deletion using an RTE-restricted Pax8-CreERT2 to precisely identify and assay Vhl/Pbrm1-null cells. We describe the histological abnormalities specifically comprising Vhl/Pbrm1-null cells observed at early (1-3 weeks), intermediate (10 months), and late (17 months) intervals post recombination. We found that Vhl/Pbrm1-null cells formed extensive ‘tumorlets’, disorganized, multilayered tubules, and cystic dilated tubules specifically in the renal cortex at 17 months post recombination. Immunohistochemistry confirmed that all epithelial cells in the lesions were Vhl/Pbrm1 null and expressed HIF1 and HIF2. These lesions contained cycling cells as identified by dual tdTomato/Ki67 staining, foci of immune cells and were strikingly surrounded by cells expressing high levels of p-ERK. We then traced these lesions to earlier timepoints, and observed Vhl/Pbrm1-null lesions resembling ‘tumorlet’ precursors and disorganized tubules at 10 months post recombination specifically in the renal cortex. Interestingly, these precursors were encompassed by macrophages and could be discerned in situ by a ring of p-ERK-expressing cells surrounding these lesions. No similar lesions were detected at 3 weeks after recombination. Interestingly, the presence of cortical lesions was associated with a region-specific expansion of the total Vhl/Pbrm1-null population over time. Our data suggest that morphological abnormalities arise after a general proliferation of Vhl/Pbrm1-null cells in the renal cortex and from a subset of cells expressing specific markers. We are now comparing the differential gene expression patterns of Vhl-null cells and Vhl/Pbrm1-null normal and transformed cells at single cell resolution to expand on these observations. Citation Format: Joanna Lima, Samvid Kurlekar, Norma Masson, Ayslan B. Barros, Maria Fernanda Soares, Christopher W. Pugh, Julie Adam, Peter J. Ratcliffe. Identification of early oncogenic lesions following concomitant Vhl and Pbrm1 loss in the murine kidney [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 6622.

Gene Expression Patterns of Colorectal Cancer Stem Cells Following Ibuprofen and Hyperthermia Treatment.

Cancer stem cells (CSCs) substantially influence the development of colorectal cancer (CRC), metastasis, relapse, and resistance to therapy. Ibuprofen and hyperthermia can be effective in the treatment of cancer. Herein, we evaluated the effects of hyperthermia and ibuprofen on the isolated-CSCs of CRC. This experimental study was conducted between Sep 2020 and Jan 2022 at the Department of Pathology, School of Medicine, Shiraz University of Medical Sciences, Iran. A non-adhesive culture system was used to isolate CSCs from HT-29 cells. To confirm the stemness nature of isolated-CSCs, the expression of stemness genes and protein markers was evaluated by quantitative Real-time PCR (qRT-PCR) and flow cytometry assay. The isolated-CSCs were treated with hyperthermia and ibuprofen. The cell viability was determined by MTT assay and trypan blue staining. The expression of stemness, proliferation, Wnt signaling pathway and apoptosis genes was assessed by qRT-PCR. CSCs were isolated within 14 days. The expression of CD-133 marker and OCT3/4, C-MYC, KLF4, and NANOG genes in isolated-CSCs was higher than HT-29 cells (P<0.05). Cell viability of treated-CSCs were considerably reduced (P<0.05). Hperthermia reduced the expression of OCT3/4, NANOG, PCNA, WNT1 and CTNNB1 genes and increased the expression of P53, BAX, and KLF4 genes (P<0.05). Ibuprofen decreased the expression of OCT3/4, BCL2, NANOG, PCNA, WNT1, and CTNNB1 genes and increased the expression of P53, BAX, and KLF4 genes in treated-CSCs (P<0.05). Hyperthermia and ibuprofen treatment demonstrate an inhibitory effect on colorectal CSCs. However, using combination therapy is remaining to be tested.

Deciphering Glioblastoma: Fundamental and Novel Insights into the Biology and Therapeutic Strategies of Gliomas.

Gliomas constitute a diverse and complex array of tumors within the central nervous system (CNS), characterized by a wide range of prognostic outcomes and responses to therapeutic interventions. This literature review endeavors to conduct a thorough investigation of gliomas, with a particular emphasis on glioblastoma (GBM), beginning with their classification and epidemiological characteristics, evaluating their relative importance within the CNS tumor spectrum. We examine the immunological context of gliomas, unveiling the intricate immune environment and its ramifications for disease progression and therapeutic strategies. Moreover, we accentuate critical developments in understanding tumor behavior, focusing on recent research breakthroughs in treatment responses and the elucidation of cellular signaling pathways. Analyzing the most novel transcriptomic studies, we investigate the variations in gene expression patterns in glioma cells, assessing the prognostic and therapeutic implications of these genetic alterations. Furthermore, the role of epigenetic modifications in the pathogenesis of gliomas is underscored, suggesting that such changes are fundamental to tumor evolution and possible therapeutic advancements. In the end, this comparative oncological analysis situates GBM within the wider context of neoplasms, delineating both distinct and shared characteristics with other types of tumors.

SCancerRNA: Expression at the Single-cell Level and Interaction Resource of Non-coding RNA Biomarkers for Cancers.

Non-coding RNAs (ncRNAs) participate in multiple biological processes associated with cancers as tumor suppressors or oncogenic drivers. Due to their high stability in plasma, urine, and many other fluids, ncRNAs have the potential to serve as key biomarkers for early diagnosis and screening of cancers. During cancer progression, tumor heterogeneity plays a crucial role, and it is particularly important to understand the gene expression patterns of individual cells. With the development of single-cell RNA sequencing (scRNA-seq) technologies, uncovering gene expression in different cell types for human cancers has become feasible by profiling transcriptomes at the cellular level. However, a well-organized and comprehensive online resource that provides access to the expression of genes corresponding to ncRNA biomarkers in different cell types at the single-cell level is not available yet. Therefore, we developed the SCancerRNA database to summarize experimentally supported data on long ncRNA, microRNA, PIWI-interacting RNA, small nucleolar RNA, and circular RNA biomarkers, as well as data on their differential expression at the cellular level. Furthermore, we collected biological functions and clinical applications of biomarkers to facilitate the application of ncRNA biomarkers to cancer diagnosis, as well as the monitoring of progression and targeted therapies. SCancerRNA also allows users to explore interaction networks of different types of ncRNAs, and build computational models in the future. SCancerRNA is freely accessible at http://www.scancerrna.com/BioMarker.