Transcription Factor Research Articles

Mitochondrial calcium uptake orchestrates vertebrate pigmentation via transcriptional regulation of keratin filaments.

Mitochondria regulate several physiological functions through mitochondrial Ca2+ dynamics. However, role of mitochondrial Ca2+ signaling in melanosome biology remains unknown. Here, we show that pigmentation requires mitochondrial Ca2+ uptake. In vitro gain and loss of function studies demonstrate that mitochondrial Ca2+ uniporter (MCU) is crucial for melanogenesis while MCU rheostat, MCUb negatively control melanogenesis. Zebrafish, MCU+/- and MCUb-/- mice models show that MCU complex drives pigmentation in vivo. Mechanistically, MCU silencing activates transcription factor NFAT2 to induce expression of keratin (5, 7, and 8) filaments. Interestingly, keratin5 in turn augments mitochondrial Ca2+ uptake and potentiates melanogenesis by regulating melanosome biogenesis and maturation. Hence this signaling module acts as a negative feedback loop that fine-tunes both mitochondrial Ca2+ signaling and pigmentation. Notably, mitoxantrone, an FDA approved drug that inhibits MCU, reduces pigmentation thereby highlighting therapeutic potential of targeting mitochondrial Ca2+ uptake for clinical management of pigmentary disorders. Taken together, we reveal an MCU-NFAT2-Keratin5 driven signaling axis that acts as a critical determinant of mitochondrial Ca2+ uptake and pigmentation. Given the vital role of mitochondrial Ca2+ signaling and keratin filaments in cellular physiology, this feedback loop could be operational in a variety of other patho-physiological processes.

The master regulator OxyR orchestrates bacterial oxidative stress response genes in space and time.

Bacteria employ diverse gene regulatory networks to survive stress, but deciphering the underlying logic of these complex networks has proved challenging. Here, we use time-resolved single-cell imaging to explore the functioning of the E.coli regulatory response to oxidative stress. We observe diverse gene expression dynamics within the network. However, by controlling for stress-induced growth-rate changes, we show that these patterns involve just three classes of regulation: downregulated genes, upregulated pulsatile genes, and gradually upregulated genes. The two upregulated classes are distinguished by differences in the binding of the transcription factor, OxyR, and appear to play distinct roles during stress protection. Pulsatile genes activate transiently in a few cells for initial protection of a group of cells, whereas gradually upregulated genes induce evenly, generating a lasting protection involving many cells. Our study shows how bacterial populations use simple regulatory principles to coordinate stress responses in space and time. A record of this paper's transparent peer review process is included in the supplemental information.

CNSC-46. GLIOBLASTOMA INITIATION, MIGRATION, AND CELL TYPES ARE REGULATED BY CORE BHLH TRANSCRIPTION FACTORS ASCL1 AND OLIG2

Abstract Glioblastomas (GBMs) are highly aggressive, infiltrative, and heterogeneous brain tumors driven by complex genetic alterations. The neurodevelopmental transcription factors ASCL1 and OLIG2 are highly co-expressed in GBMs. However, their combinatorial roles in regulating the hierarchy and heterogeneity of GBM cells are unknown. Here, we show that induction of somatic mutations in neural progenitor cells lead to the dysregulation of ASCL1 and OLIG2, which then function redundantly and are required for brain tumor formation in a mouse model of GBM. Subsequently, the binding of ASCL1 and OLIG2 to each other’s loci and to downstream target genes then determine the cell types and degree of migration of tumor cells. Notably, single-cell RNA sequencing (scRNA-seq) reveals that a high level of ASCL1 is key in promoting neural stem cell (NSC)/astrocyte-like tumor cell types, which are highly proliferative, migratory, and are marked by upregulation of ribosomal protein, oxidative phosphorylation, cancer metastasis, and therapeutic resistance genes.

The histone deacetylase RhHDA15 represses petal senescence by epigenetically regulating reactive oxygen species homeostasis in rose.

Epigenetic modifications play vital roles in many biological processes. Flower senescence involves epigenetic factors that influence the chromatin state and gene expression. However, the molecular mechanism underlying the role of histone deacetylation in regulating flower senescence has not been elucidated. Here, we demonstrate that histone deacetylation is involved in flower senescence by fine-tuning reactive oxygen species (ROS) homeostasis in rose (Rosa hybrida). Our data reveal that the histone lysine deacetyltransferase RhHDA15 inhibits ROS accumulation and petal senescence by downregulating the expression of NADPH OXIDASE/RESPIRATORY BURST OXIDASE HOMOLOG (RhRboh) genes. Furthermore, the transcription factor RELATED TO ABI3/VP1 2 (RhRAV2) recruits RhHDA15 and the co-repressor TOPLESS (RhTPL) to suppress flower senescence by reducing H3 lysine 9 acetylation (H3K9ac) at the RhRbohA1/2 promoter and thus directly inhibiting precocious RhRbohA1/2 expression. Our work sheds light on an epigenetic mechanism in which histone deacetylation plays a crucial role in controlling petal senescence by precisely fine-tuning ROS homeostasis, providing insights into the regulatory network of organ senescence.

SURG-56. TRANSCRIPTION FACTOR CELL LINEAGE DETERMINES AGGRESSIVE BEHAVIOR AMONG NONFUNCTIONAL PITUITARY ADENOMAS

Abstract INTRODUCTION Pituitary neuroendocrine tumors (PitNETs) are the second most common intracranial neoplasm. Silent or non-functional PitNETs represent a common neuro-oncological challenge due to their size and unpredictable invasion of parasellar structures. This makes their complete surgical resection difficult and puts patients at higher risk for recurrence. OBJECTIVE To characterize differences in tumor volume, cavernous sinus invasion, and recurrence in nonfunctional PitNETs. METHODS Out of 615 transsphenoidal surgeries from 2012 to 2024, there were 309 overall PitNETs and 181 nonfunctional PitNETs. Based on new WHO criteria for transcription factor profiling (PIT-1, TPIT, and SF-1), there were 98 silent gonadotroph adenomas (SGA), 31 silent corticotroph adenomas (SCA), 10 silent pleurihormonal tumors (PHT), and 2 null cell adenomas. Tumor volume was calculated using (AxBxC/2). Knosp grade was calculated. Cavernous sinus invasion was defined as Knosp ≥ 3. Residual disease at 3 months postoperative was recorded and surgical intervention for growing residual disease was captured. RESULTS Average tumor volume of PHT (18.08 cm3) was significantly higher than SGA (8.44 cm3, p=0.007) and similar to SCA (10.33 cm3, p=0.11). Percentage of cavernous sinus invasion was significantly different between SCA (31.82%) and SGA (10.20%, p=0.003), but not PHT (33.33%; p>0.05). There was a significantly higher residual rate in SCA compared to SGA (31.71% vs. 11.22%, p=0.006). There were significantly more reoperations in the SCA group compared to SGA and PHT. CONCLUSIONS Despite similar tumor volume between SCA and SGA, there was a significantly higher rate of cavernous sinus invasion and residual disease among SCA. This has important neuro-oncological implications regarding surgical technique, postoperative surveillance, and need for adjuvant therapy to mitigate aggressive recurrent disease.

Shared genetic architecture of psychiatric disorders and hemorrhoidal disease: a large-scale genome-wide cross-trait analysis

BackgroundThe genetic association between psychiatric disorders and hemorrhoidal disease (HEM) is still not well known. The work aims to investigate their comorbidity at a genetic level.MethodsUtilizing recent large-scale genome-wide association studies (GWAS), we investigated the genetic overlap at the single nucleotide polymorphism (SNP), gene, and molecular level between depression and HEM, bipolar disorder (BD) and HEM, neuroticism and HEM, as well as schizophrenia (SCZ) and HEM. The cross-trait genes were validated through the utilization of transcriptome and proteome methodologies. The causal link was assessed using bidirectional two-sample Mendelian randomization analysis (MR) analysis. MRlap corrects for the potential bias in estimation caused by sample overlap.ResultsWe discovered significant positive genetic associations between these four types of psychiatric disorders and HEM. Cross-phenotypic association analyses identified shared SNPs along with 17 specific loci between psychiatric disorders and HEM. MAGMA identified a total of 2304 pleiotropic genes, several of which showed significant expression in the results of transcriptome and proteome analyses. We observed that these genes are mostly associated with the regulation of transcription factors and particular DNA binding activities. Lastly, MR analysis provided evidence supporting a correlation between these conditions.ConclusionThis study revealed a genetic correlation between four psychiatric disorders and HEM, identified pleiotropic loci, found multiple candidate genes, and confirmed causal relationships. This has enhanced our comprehension of the common genetic mechanisms of psychiatric disorders and HEM.

Profile of Cytokines and T Cell Subsets Transcription Factors in Cerebrospinal Fluid of Patients with Viral Encephalitis.

This study investigates the demographic, clinical characteristics, virological profiles, and immunological responses of patients with viral encephalitis (VE) compared with a control group. The VE group displayed a wide range of neurological symptoms. Virological analysis revealed the predominance of Herpesviridae family viruses. Immune responses in cerebrospinal fluid (CSF) from patients with VE were examined, highlighting an immunological shift toward T helper 1 (Th1) cells dominance, altered T helper 17 cells/regulatory T cells (Th17/Tregs) balance, and high interleukin-6 expression. These findings provide insights into the complex immunological landscape of VE, highlighting the role of specific cytokines and T cell subsets in its pathogenesis and potentially guiding targeted therapeutic strategies.

TMOD-06. ESTABLISHMENT OF PITUITARY NEUROENDOCRINE TUMOR (PITNET) ORGANOID MODELS

Abstract Pituitary neuroendocrine tumors (PitNETs) are tumors that originate from the anterior pituitary gland. While they are mostly benign, they can cause various clinical symptoms due to hormonal secretion abnormalities. PitNET can be broadly categorized into three lineages, each of which has its differentiation controlled by specific transcription factors. Treatment options for PitNET include surgical resection and pharmacotherapy, but available drugs are limited, and their efficacy markers remain unclear. To develop novel drugs for PitNET and identify their efficacy markers, it is necessary to develop new research models. In recent years, organoid models have garnered attention as novel research models for various cancer types. In this study, we established novel organoid models derived from PitNET patients and analyzed their histological and endocrinological characteristics. Surgical specimens from 4 cases of somatotroph adenoma, 1 case of double adenoma, 1 case of gonadotroph adenoma, and 1 case of corticotroph adenoma were minced in dissection medium and embedded in Matrigel basement membrane matrix respectively. The organoids were cultured under 5% CO2 at 37°C using media supplemented with various growth factors. After 28 days of culture, formalin-fixed paraffin-embedded sections were prepared to examine histological homology with patient tumors. Hematoxylin and eosin staining results showed that all organoids maintained cell morphology and tissue structure similar to the original tumors. Immunohistochemical staining for markers specific to each PitNET revealed similar staining patterns between the organoids and the original tumors. Furthermore, analysis of hormone levels in the culture medium showed that hormone production capabilities, such as growth hormone and follicle stimulating hormone, were maintained even after long-term culture. In this study, we successfully established PitNET organoid models that retain histological and endocrinological characteristics. These models are expected to serve as useful research tools for elucidating the molecular mechanisms of PitNET and developing novel therapies targeting critical molecular abnormalities.

Perception and processing of stress signals by plant mitochondria.

In the course of their life, plants continuously experience a wide range of unfavourable environmental conditions in the form of biotic and abiotic stress factors. The perception of stress via various organelles and rapid, tailored cellular responses are essential for the establishment of plant stress resilience. Mitochondria as the biosynthetic sites of energy equivalents in the form of ATP-provided in order to enable a multitude of biological processes in the cell-are often directly impacted by external stress factors. At the same time, mitochondrial function may fluctuate to a tolerable extent without the need to activate downstream retrograde signalling cascades for stress adaptation. In this Focus Review, we summarise the current state of knowledge on the perception and processing of stress signals by mitochondria and show which layers of retrograde signalling, that is, those involving transcription factors, metabolites, but also enzymes with moonlighting functions, enable communication with the nucleus. Also, light is shed on signal integration between mitochondria and chloroplasts as part of retrograde signalling. With this Focus Review, we aim to show ways in which organelle-specific communication can be further researched and the collected data used in the long-term to strengthen plant resilience in the context of climate change.

TMET-39. DUAL TARGETING OF MAP KINASE SIGNALING AND METABOLISM IS A THERAPEUTIC VULNERABILITY IN PEDIATRIC DIFFUSE MIDLINE GLIOMAS

Abstract Metabolic reprogramming in pediatric diffuse midline glioma is driven by gene expression changes induced by the hallmark histone mutation H3K27M, which results in aberrantly permissive activation of oncogenic signaling pathways. Previous studies of diffuse midline glioma with altered H3K27 (DMG-H3K27a) have shown that the RAS pathway, specifically through its downstream kinase, extracellular-signal-related kinase 5 (ERK5), is critical for tumor growth. However, there remains a knowledge gap in identifying effectors of ERK signaling and their roles in DMG-H3K27a. Leveraging several patient multi-omic datasets, we identified a unique relationship between ERK5 and brain tumor glycolysis. We establish that ERK5 is a critical regulator of cell proliferation and glycolysis in DMG-H3K27a, where ERK5 knockdown leads to decreases in glycolysis and increases in mitochondrial metabolism. We demonstrate through loss of function and overexpression studies that ERK5 mediates glycolysis through regulation of glycolytic enzyme Phosphofructo-2-Kinase/Fructose-2,6-Biphosphatase 3 (PFKFB3). PFKFB3 is a protein that catalyzes the synthesis or degradation of fructose-2,6-bisphosphate (F2,6BP). Furthermore, we establish a novel mechanism, in which ERK5 mediates PFKFB3 expression via activation of MEF2A, an oncogenic transcription factor. DMG-H3K27a cells are sensitive to the loss or inhibition of PFKFB3 both in vitro and in orthotopic mouse models. However, monotherapy is ineffective in heterogenous diseases such as gliomas, including DMG-H3K27a. To combat this, we show multi-targeted therapy against the ERK5-PFKFB3 axis is synergistic in vitro and in vivo. Multi-targeted drug therapy against the ERK5-PFKFB3 axis, such as with small-molecule inhibitors, may represent a promising therapeutic approach in patients with pediatric diffuse midline glioma. Moreover, we have identified resistance to our dual therapy using several systems biology approaches, which we are currently investigating and validating. Collectively, our study defines a novel, targetable axis in DMG-H3K27a and highlights the importance of tumor cell metabolism in defining therapeutic vulnerabilities.

TMOD-17. FOXR2-ACTIVATED CNS NEUROBLASTOMA ORIGINATES FROM THE MEDIAL GANGLIONIC EMINENCE LINEAGE

Abstract FOXR2-activated CNS neuroblastoma is a rare subtype of pediatric brain tumor characterized by the elevated expression of the transcription factor FOXR2 due to genomic rearrangement. However, the precise pathogenic mechanisms, including the cell type of origin and the function of FOXR2, are not fully understood. An expression profile analysis of patient tumors showed increased expression of the marker genes associated with the Medial Ganglionic Eminence (MGE) in the ventral forebrain such as NKX2.1 and SOX6. Based on this result, we hypothesized that FOXR2-activated CNS neuroblastoma originates from the MGE lineage. To test this hypothesis, we overexpressed FOXR2 in human embryonic stem cell-derived MGE progenitors and, as a control cell type, cortical progenitors. Our data showed that FOXR2 overexpression resulted in a significantly increased proliferation in the MGE progenitors but not in the cortical progenitors, indicating a cell-type-specific effect of FOXR2. When injected into the brain of immunodeficient mice, FOXR2-expressing MGE progenitors formed tumors that molecularly resemble patient tumors, while FOXR2-expressing cortical progenitors did not. To further dissect the underlying molecular mechanisms, we conducted an RNA-seq experiment and found that FOXR2 overexpression activated the MEK/ERK signaling pathway through a suppression of the endogenous RAS inhibitor DIRAS3. Moreover, the FDA-approved MEK inhibitor trametinib significantly suppressed the proliferation and induced cell death in FOXR2-expressing MGE progenitors in vitro with the IC50 value of 30 nM. Collectively, our study has revealed the cell type of origin of FOXR2-activated CNS neuroblastoma and identified the MEK/ERK pathway as a critical downstream of FOXR2 that can be targeted therapeutically.

A full genome assembly reveals drought stress effects on gene expression and metabolite profiles in blackcurrant (Ribes nigrum L.)

Abstract Blackcurrant (Ribes nigrum L., family Grossulariaceae) is a perennial shrub that is widely cultivated for its edible berries. These are rich in antioxidants, vitamin C and anthocyanins, making them a valuable ingredient in the food and beverage industry. However, prolonged periods of drought during the fruiting season lead to drought stress, which has serious ecological and agricultural implications, inhibiting blackcurrant growth and reducing yields. To facilitate the analysis of underlying molecular processes, we present the first high-quality chromosome-scale and partially haplotype-resolved assembly of the blackcurrant genome (cv. Rosenthals Langtraubige), also the first in the family Grossulariaceae. We used this genomic reference to analyze the transcriptomic response of blackcurrant leaves and roots to drought stress, revealing differentially expressed genes with diverse functions, including those encoding the transcription factors bZIP, bHLH, MYB and WRKY, and tyrosine kinase-like kinases such as PERK and DUF26. Gene expression was correlated with the abundance of primary metabolites, revealing 14 with significant differences between stressed leaves and controls indicating a metabolic response to drought stress. Amino acids such as proline were more abundant under stress conditions, whereas organic acids were depleted. The genomic and transcriptomic data from this study can be used to develop more robust blackcurrant cultivars that thrive under drought stress conditions.

New targets and biomarkers for doxorubicin-induced cardiotoxicity in humans: implications drawn from toxicogenomic data and molecular modelling

The doxorubicin-induced cardiotoxicity continues to be a life-threatening adverse effect in the clinic. Doxorubicin-induced acute cardiotoxicity is reversible, whereas chronic cardiotoxicity is irreversible, leading to dilated cardiomyopathy and heart failure. The aim of this study was to identify the molecular mechanisms associated with doxorubicin metabolites in doxorubicin-induced chronic cardiotoxicity. For this purpose, literature searches and in silico toxicogenomic analyses were conducted using various tools, including the Comparative Toxicogenomic Database, GeneMANIA, Metascape, MIENTURNET, ChEA3, and AutoDock. Additionally, molecular dynamics simulations were performed for 500 ns using Schrödinger software to assess the stability and dynamics of the representative docked complexes. We observed that doxorubicin biotransformed into five metabolites in the human heart and identified 11 common genes related to doxorubicin, its metabolites, dilated cardiomyopathy, and heart failure. Our findings revealed that doxorubicin and its metabolites primarily exhibited binding affinity to the beta-1 adrenergic receptor and fatty acid synthase. Furthermore, we identified several key transcription factors, especially the Homeobox protein Nkx-2.6, and hsa-miR-183-3p associated with this cardiotoxicity. Finally, we observed that, in addition to doxorubicinol, 7-deoxidoxorubicinone, another metabolite of doxorubicin, may also contribute to this cardiotoxicity. These findings contribute to our understanding of the processes underlying doxorubicin-induced chronic cardiotoxicity.

The homeostasis of AtMYB4 is maintained by ARA4, HY5, and CAM7 during Arabidopsis seedling development.

Calmodulin7 (CAM7) is a key transcription factor of Arabidopsis seedling development. CAM7 works together with HY5 bZIP protein to promote photomorphogenesis at various wavelengths of light. In this study, we show that AtMYB4, identified from a yeast two-hybrid screen, physically interacts with CAM7 and works as a positive regulator of photomorphogenesis at various wavelengths of light. CAM7 and HY5 directly bind to the promoter of AtMYB4 to promote its expression for photomorphogenic growth. On the other hand, ARA4, identified from the same yeast two-hybrid screen, works as a negative regulator of photomorphogenic growth specifically in white light. The double mutant analysis reveals that the altered hypocotyl elongation of atmyb4 and ara4 is either partly or completely suppressed by additional loss of function of CAM7. Furthermore, ARA4 genetically interacts with AtMYB4 in an antagonistic manner to suppress the elongated hypocotyl phenotype of atmyb4. The transactivation studies reveal that while CAM7 activates the promoter of AtMYB4 in association with HY5, ARA4 negatively regulates AtMYB4 expression. Taken together, these results demonstrate that working as a negative regulator of photomorphogenesis, ARA4 plays a balancing act on CAM7 and HY5-mediated regulation of AtMYB4.

CSIG-12. YAP1-TEAD SIGNALING CONTRIBUTES TO LUNG CANCER COLONIZATION OF THE BRAIN

Abstract BACKGROUND Lung cancer is the most common primary tumor to metastasize to the brain. Poor understanding of the brain metastasis process limits therapeutic options. Despite emerging data on the role of YAP signaling pathway in the development of brain tumors, a gap in knowledge remains regarding its role in brain colonization. We hypothesize that YAP1 and the associated TEAD transcriptional factors are responsible for lung cancer progression and that intervention with YAP/TEAD-targeted therapy will prevent brain dissemination. METHODS Lung cancer cells at different stages of cancer progression (VMRC-LCD and NCI-H1975, lung cancer cell lines; NCI-H1915 and LN001, brain metastatic cancer cells) were genetically engineered to overexpress or silence/knockout YAP1 using lentiviral transduction of constructs expressing shRNA/sgRNA or YAP1. We tested several tumor cell capabilities using well-established in vitro assays, including invasion and migration assays, and an in vivo brain metastasis (BM) model to determine the biological effects of YAP1/TEAD suppression at different stages of lung cancer progression. Dose-response assays were performed to determine IC50 of the TEAD inhibitor, verteporfin. RESULTS YAP overexpression enhances the proliferative phenotype in metastatic and BM cell lines but not in non-metastatic cells. However, YAP overexpression only increases the migration ability of BM cell lines such as LN001 and NCI-H1915 compared to the metastatic NCI-H1975 and non-metastatic lung cancer cells. Moreover, we showed the efficacy of the YAP1/TEAD inhibitor in patient-derived lung BM cells. Finally, our in vivo preliminary data showed YAP1 involvement in lung cancer cell colonization of the brain but not in sustaining brain metastasis growth. CONCLUSIONS Our findings showed YAP1 involvement in lung cancer cell colonization of the brain, suggesting YAP1 inhibition is a clinically actionable target for preventing lung cancer dissemination and providing a strong rationale to further evaluate the benefit of a YAP1/TEAD-targeted therapy in in vivo models of BM.

EXTH-83. CHARACTERIZATION OF SPLICING ABERRATIONS INDUCED BY PRMT5 INHIBITION IN GLIOBLASTOMA

Abstract Glioblastoma (GBM) are lethal tumors with limited treatment options, with a high unmet need for novel therapies. PRMT5, an arginine methyltransferase that regulates several histone and non-histone targets, is overexpressed in cancers including GBM and is a promising therapeutic target given its role in promoting oncogenic processes. Here, we characterized the effects of pharmacological inhibition of PRMT5 by the brain-penetrant, orally bioavailable inhibitors, PRT811 and PRT808 on treatment-induced splicing aberrations using a panel of patient-derived glioma stem cells (GSC) and organotypic human glioma slice cultures. Changes in protein expression upon PRMT5 inhibition were assessed using reverse phase protein array (RPPA) analysis. Additionally, splicing aberrations due to PRMT5 inhibition in were tested in several GSCs using RNA-seq. We observed a potent reduction of symmetrical dimethylation of arginine (SDMA protein marks by both PRMT5 inhibitors across various GSC lines and ex-vivo brain tumor tissue slices. Notably, GSCs with both mutant and wild-type p53 displayed similar sensitivity to PRMT5 inhibition, independent of MTAP status. Moreover, PRMT5 inhibition elicited reduced cell proliferation, altered cell survival pathways, and apoptosis. RNA-seq analysis revealed that splicing alterations affecting numerous oncogenes and regulatory factors, encompassing 42 cancer driver genes, 80 splicing factors, and 42 transcriptional factors across these GSC lines, with specific splicing events observed in nuclear and cytoplasmic subcellular compartments. Additional analysis is ongoing to determine which of the aberrant transcripts are translated to yield tumor specific neoantigens that can be potential targets for tumor specific immune strategies. These results underscore the efficacy of pharmacological PRMT5 inhibition in genetically and epigenetically diverse GSCs, and highlight its potential for developing novel immune and non-immune therapeutic strategies against GBM.

Brca1 haploinsufficiency promotes early tumor onset and epigenetic alterations in a mouse model of hereditary breast cancer.

Germline BRCA1 mutation carriers face a high breast cancer risk; however, the underlying mechanisms for this risk are not completely understood. Using a new genetically engineered mouse model of germline Brca1 heterozygosity, we demonstrate that early tumor onset in a Brca1 heterozygous background cannot be fully explained by the conventional 'two-hit' hypothesis, suggesting the existence of inherent tumor-promoting alterations in the Brca1 heterozygous state. Single-cell RNA sequencing and assay for transposase-accessible chromatin with sequencing analyses uncover a unique set of differentially accessible chromatin regions in ostensibly normal Brca1 heterozygous mammary epithelial cells, distinct from wild-type cells and partially mimicking the chromatin and RNA-level changes in tumor cells. Transcription factor analyses identify loss of ELF5 and gain of AP-1 sites in these epigenetically primed regions; in vivo experiments further implicate AP-1 and Wnt10a as strong promoters of Brca1-related breast cancer. These findings reveal a previously unappreciated epigenetic effect of Brca1 haploinsufficiency in accelerating tumorigenesis, advancing our mechanistic understanding and informing potential therapeutic strategies.

TMOD-27. RECURRENT GLIOBLASTOMA AFTER LASER ABLATION OF THE PRIMARY TUMOR IN A PRECLINICAL ORTHOTOPIC MODEL EXHIBIT INCREASED GENETIC SIGNATURES OF CELL CYCLE AND CELL MOTILITY

Abstract Image-guided laser interstitial thermal therapy (LITT) is a minimally invasive tumor cytoreductive treatment for recurrent gliomas and tumors in eloquent loci. We have adapted this technique to develop an image-guided glioblastoma (GBM) ablation model, its recurrence and have tested the efficacy of imaging biomarkers in evaluating tumor ablation and recurrence. The cytopathology and molecular signatures of the primary and recurrent tumors were compared. Immune-compromised female rats were implanted with U251N tumor cells in one brain hemisphere (n=20). Tumor growth was monitored using magnetic resonance imaging (MRI). When tumors reached about 4 mm in diameter, they were ablated using a clinical LITT system (Visualase®), under MRI guidance. Five other rats implanted with U251N tumors were used as unablated controls. MRI data were acquired at 24 h post-LITT, and at 2 and 4 weeks. Rats were sacrificed for histopathology at 2 and 4 weeks, and brain sections stained for hematoxylin and eosin, human major histocompatibility complex, Ki67, a cell proliferation marker and sex determining region Y)-box 2 (SOX2), a stem cell transcription factor. An additional cohort of rats with primary (n=4) and post-LITT recurrent (n=4) U251N tumors were used for analysis of their molecular composition using RNA-Seq approach. In the treated groups, MRI showed little tumor tissue at 24 h, evidence of recurrence at 2 weeks and significant tumor tissue at 4 weeks. Tumor DCE-MRI parameters showed elevated intra-tumoral vascular permeability (i.e., Ktrans) values at pre-LITT imaging, that shifted to peri-ablation periphery at 24 h. A trend of progressive decrease in Ktrans was seen until 1-week post-ablation. Increasing Ktrans values at 2 weeks and after 4 weeks coincided with tumor recurrence. RNA-Seq data showed that cell cycle, cellular movement and inflammatory disease genes were the most differentially expressed genes in the recurrent tumors suggesting increased infiltration may primarily underlie treatment resistance.

TMET-10. TARGETING PGM3 TO INHIBIT HEXOSAMINE PRODUCTION IS AN EFFECTIVE APPROACH TO TREAT GLIOBLASTOMA

Abstract Hexosamine biosynthesis pathway (HBP) mediated by a series of enzymes is highly upregulated in many cancers to promote the glycosylation of proteins and lipids, while which enzyme in this pathway is better to serve as anti-tumor target is unclear. Here, we found targeting GFAT1, the initial and rate-limiting enzyme in HBP, fails to inhibit the growth of glioblastoma (GBM), the most lethal brain tumor, due to this cancer dramatically elevating NAGK-mediated hexosamine salvage pathway. In contrast, targeting PGM3, the latter enzyme in HBP, blocks both de novo hexosamine synthesis and salvage pathways, effectively inhibiting GBM growth. We further identified HBP enzymes are upregulated by SREBP-1, a key lipogenic transcriptional factor. In turn, this upregulation promotes SREBP-1 activation to stimulate lipogenesis via stabilization of its key transporter SCAP by N-glycosylation modification. Together, our study uncovered a previously unrecognized HBP-SCAP-SREBP-1 feedforward loop and demonstrated targeting PGM3 is an effective approach to treat GBM.

The Expression of HPV-16 E5 Oncoprotein Impacts the Transcript Profiles of FGFR2 and EMT-Related Genes in Preneoplastic Anal Epithelium Lesions

Anal Squamous Cell Carcinoma (SCCA) is a rare Human Papillomavirus type 16 (HPV16)-associated carcinoma whose pathogenesis is still poorly understood. Recent studies based on biopsy and Next Generation Sequencing (NGS) approaches have linked the viral episomal status to aggressive SCCA phenotypes, suggesting a potential role of the 16E5 oncoprotein in tumor development. Our previous findings indicated that 16E5 induces Fibroblast Growth Factor Receptor 2 (FGFR2) isoform switching, aberrant mesenchymal FGFR2c expression, Epithelial Mesenchymal Transition (EMT), and cell invasion in various in vitro human keratinocyte models, as well as in the in vivo context of cervical Low-grade Squamous Intraepithelial Lesions (LSILs). To further explore the role of 16E5 in epithelial carcinogenesis, this study aims to investigate the molecular profile in HPV-related anal lesions. The results showed a significant positive correlation between 16E5 and FGFR2c, as well as 16E5 or FGFR2c and key EMT-related transcription factors, particularly in the group of HPV16 positive anal samples not containing without high grade lesions. Additionally, by coupling the molecular analysis with an interactome investigation, we hypothesized a potential functional interplay between the Ca2+ channel Transient Receptor Potential Ankyrin 1 (TRPA1) and FGFR2c, mediated by 16E5 during the establishment of the oncogenic signaling. These findings will help to elucidate the actual relevance of 16E5 in the early progression of anal lesions and contribute to determine its potential as target for future preventive approaches for HPV16-positive SCCA.