Critical role of G3BP1 in bovine parainfluenza virus type 3 (BPIV3)-inhibition of stress granules formation and viral replication.

Background: It remains unclear whether BPIV3 infection leads to stress granules formation and whether G3BP1 plays a role in this process and in viral replication. This study aims to clarify the association between BPIV3 and stress granules, explore the effect of G3BP1 on BPIV3 replication, and provide significant insights into the mechanisms by which BPIV3 evades the host's antiviral immunity to support its own survival. Methods: Here, we use Immunofluorescence staining to observe the effect of BPIV3 infection on the assembly of stress granules. Meanwhile, the expression changes of eIF2α and G3BP1 were determined. Overexpression or siRNA silencing of intracellular G3BP1 levels was examined for its regulatory control of BPIV3 replication. Results: We identify that the BPIV3 infection elicited phosphorylation of the eIF2α protein. However, it did not induce the assembly of stress granules; rather, it inhibited the formation of stress granules and downregulated the expression of G3BP1. G3BP1 overexpression facilitated the formation of stress granules within cells and hindered viral replication, while G3BP1 knockdown enhanced BPIV3 expression. Conclusion: This study suggest that G3BP1 plays a crucial role in BPIV3 suppressing stress granule formation and viral replication.

The correlation study between TOP2A gene expression in circulating tumor cells and chemotherapeutic drug resistance of patients with breast cancer.

BACKGROUND: Patients with breast cancer (BC) at advanced stages have poor outcomes because of high rate of recurrence and metastasis. Biomarkers for predicting prognosis remain to be explored. This study aimed to evaluate the relationships between circulating tumor cells (CTCs) and outcomes of BC patients. PATIENTS AND METHODS: A total of 50 female were enrolled in this study. Their diagnoses were determined by clinical characteristics, image data, and clinical pathology. CTC subtypes and TOP2A gene expression on CTCs were detected by CanPatrol™ technology and triple color in situ RNA hybridization (RNA-ISH), which divided into epithelial CTCs (eCTCs), mesenchymal CTCs (MCTCs), and hybrid CTCs (HCTCs) based on their surface markers. Hormone receptor, including estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER-2) expression, was measured by immunohistochemistry (IHC) method before treatment. The risk factors for predicting recurrence and metastasis were calculated by COX risk regression model. The progression-free survival (PFS) of patients was determined using Kaplan-Meier survival curve. RESULTS: The patients with a large tumor size (≥ 3 cm) and advanced tumor node metastasis (TNM) stages had high total CTCs (TCTCs) (P < 0.05). These patients also had high TOP2A expression level. COX risk regression analysis indicated that TOP2A expression levels in TCTCs, ER + , HER-2 + , and TNM stages were critical risk factors for recurrence and metastasis of patients (P < 0.05). The PFS of patients with ≥ 5 TCTCs, ≥ 3 HCTCs, and positive TOP2A expression in ≥ 3 TCTCs was significantly longer than that in patient with < 5 TCTCs, < 3 HCTCs, and TOP2A expression in < 3 TCTCs (P < 0.05). In contrast, the PFS of patients with positive hormone receptors (ER + , PR + , HER-2 +) also was dramatically lived longer than that in patients with negative hormone receptor expression. CONCLUSIONS: High TCTC, HCTCs, and positive TOP2A gene expression on CTCs were critical biomarkers for predicting outcomes of BC patients. Positive hormone receptor expression in BC patients has significant favor PFS.

Cockayne Syndrome Patient iPSC-Derived Brain Organoids and Neurospheres Show Early Transcriptional Dysregulation of Biological Processes Associated with Brain Development and Metabolism.

Cockayne syndrome (CS) is a rare hereditary autosomal recessive disorder primarily caused by mutations in Cockayne syndrome protein A (CSA) or B (CSB). While many of the functions of CSB have been at least partially elucidated, little is known about the actual developmental dysregulation in this devasting disorder. Of particular interest is the regulation of cerebral development as the most debilitating symptoms are of neurological nature. We generated neurospheres and cerebral organoids utilizing Cockayne syndrome B protein (CSB)-deficient induced pluripotent stem cells derived from two patients with distinct severity levels of CS and healthy controls. The transcriptome of both developmental timepoints was explored using RNA-Seq and bioinformatic analysis to identify dysregulated biological processes common to both patients with CS in comparison to the control. CSB-deficient neurospheres displayed upregulation of the VEGFA-VEGFR2 signalling pathway, vesicle-mediated transport and head development. CSB-deficient cerebral organoids exhibited downregulation of brain development, neuron projection development and synaptic signalling. We further identified the upregulation of steroid biosynthesis as common to both timepoints, in particular the upregulation of the cholesterol biosynthesis branch. Our results provide insights into the neurodevelopmental dysregulation in patients with CS and strengthen the theory that CS is not only a neurodegenerative but also a neurodevelopmental disorder.

Spectrum of ERCC6-Related Cockayne Syndrome (Type B): From Mild to Severe Forms.

(1) Background: Cockayne syndrome (CS) is an ultra-rare multisystem disorder, classically subdivided into three forms and characterized by a clinical spectrum without a clear genotype-phenotype correlation for both the two causative genes ERCC6 (CS type B) and ERCC8 (CS type A). We assessed this, presenting a series of patients with genetically confirmed CSB. (2) Materials and Methods: We retrospectively collected demographic, clinical, genetic, neuroimaging, and serum neurofilament light-chain (sNFL) data about CSB patients; diagnostic and severity scores were also determined. (3) Results: Data of eight ERCC6/CSB patients are presented. Four patients had CS I, three patients CS II, and one patient CS III. Various degrees of ataxia and spasticity were cardinal neurologic features, with variably combined systemic characteristics. Mean age at diagnosis was lower in the type II form, in which classic CS signs were more evident. Interestingly, sNFL determination appeared to reflect clinical classification. Two novel premature stop codon and one novel missense variants were identified. All CS I subjects harbored the p.Arg735Ter variant; the milder CS III subject carried the p.Leu764Ser missense change. (4) Conclusion: Our work confirms clinical variability also in the ERCC6/CSB type, where manifestations may range from severe involvement with prenatal or neonatal onset to normal psychomotor development followed by progressive ataxia. We propose, for the first time in CS, sNFL as a useful peripheral biomarker, with increased levels compared to currently available reference values and with the potential ability to reflect disease severity.

Exosomal miRNA-92a derived from cancer-associated fibroblasts promote invasion and metastasis in breast cancer by regulating G3BP2.

Cancer-associated Fibroblasts (CAFs) exert a tumor-promoting effect in various cancers, including breast cancer. CAFs secrete exosomes containing miRNA and proteins, influencing the tumor microenvironment. In this study, we identified CAF-derived exosomes that transport functional miR-92a from CAFs to tumor cells, thereby intensifying the aggressiveness of breast cancer. CAFs downregulate the expression of G3BP2 in breast cancer cells, and a significant elevation in miR-92a levels in CAF-derived exosomes was observed. Both in vitro and in vivo experiments demonstrate that miR-92a enhances breast cancer cell migration and invasion by directly targeting G3BP2, functioning as a tumor-promoting miRNA. We validated that the RNA-binding proteins SNRPA facilitate the transfer of CAF-derived exosomal miR-92a to breast cancer cells. The reduction of G3BP2 protein by CAF-derived exosomes releases TWIST1 into the nucleus, promoting epithelial-mesenchymal transition (EMT) and further exacerbating breast cancer progression. Moreover, CAF-derived exosomal miR-92a induces tumor invasion and metastasis in mice. Overall, our study reveals that CAF-derived exosomal miR-92a serves as a promoter in the migration and invasion of breast cancer cells by reducing G3BP2 and may represent a potential novel tumor marker for breast cancer.

Phosphorylation of G3BP1 is involved in the regulation of PDCoV-induced inflammatory response.

Stress granules (SGs), the main component is GTPase-activating protein-binding protein 1 (G3BP1), which are assembled during viral infection and function to sequester host and viral mRNAs and proteins, are part of the antiviral responses. In this study, we found that porcine deltacoronavirus (PDCoV) infection induced stable formation of robust SGs in cells through a PERK (protein kinase R-like endoplasmic reticulum kinase)-dependent mechanism. Overexpression of SGs marker proteins G3BP1 significantly reduced PDCoV replication in vitro, while inhibition of endogenous G3BP1 enhanced PDCoV replication. Moreover, PDCoV infected LLC-PK1 cells raise the phosphorylation level of G3BP1. By overexpression of the G3BP1 phosphorylated protein or the G3BP1 dephosphorylated protein, we found that phosphorylation of G3BP1 is involved in the regulation of PDCoV-induced inflammatory response. Taken together, our study presents a vital aspect of the host innate response to invading pathogens and reveals attractive host targets for antiviral target.

DNA topoisomerase II inhibition potentiates osimertinib's therapeutic efficacy in EGFR-mutant non-small cell lung cancer models.

Development of effective strategies to manage the inevitable acquired resistance to osimertinib, a third-generation EGFR inhibitor for the treatment of EGFR-mutant (EGFRm) non-small cell lung cancer (NSCLC), is urgently needed. This study reports that DNA topoisomerase II (Topo II) inhibitors, doxorubicin and etoposide, synergistically decreased cell survival, with enhanced induction of DNA damage and apoptosis in osimertinib-resistant cells; suppressed the growth of osimertinib-resistant tumors; and delayed the emergence of osimertinib-acquired resistance. Mechanistically, osimertinib decreased Topo IIα levels in EGFRm NSCLC cells by facilitating FBXW7-mediated proteasomal degradation, resulting in induction of DNA damage; these effects were lost in osimertinib-resistant cell lines that possess elevated levels of Topo IIα. Increased Topo IIα levels were also detected in the majority of tissue samples from patients with NSCLC after relapse from EGFR tyrosine kinase inhibitor treatment. Enforced expression of an ectopic TOP2A gene in sensitive EGFRm NSCLC cells conferred resistance to osimertinib, whereas knockdown of TOP2A in osimertinib-resistant cell lines restored their susceptibility to osimertinib-induced DNA damage and apoptosis. Together, these results reveal an essential role of Topo IIα inhibition in mediating the therapeutic efficacy of osimertinib against EGFRm NSCLC, providing scientific rationale for targeting Topo II to manage acquired resistance to osimertinib.

DNA fragility at topologically associated domain boundaries is promoted by alternative DNA secondary structure and topoisomerase II activity.

CCCTC-binding factor (CTCF) binding sites are hotspots of genome instability. Although many factors have been associated with CTCF binding site fragility, no study has integrated all fragility-related factors to understand the mechanism(s) of how they work together. Using an unbiased, genome-wide approach, we found that DNA double-strand breaks (DSBs) are enriched at strong, but not weak, CTCF binding sites in five human cell types. Energetically favorable alternative DNA secondary structures underlie strong CTCF binding sites. These structures coincided with the location of topoisomerase II (TOP2) cleavage complex, suggesting that DNA secondary structure acts as a recognition sequence for TOP2 binding and cleavage at CTCF binding sites. Furthermore, CTCF knockdown significantly increased DSBs at strong CTCF binding sites and at CTCF sites that are located at topologically associated domain (TAD) boundaries. TAD boundary-associated CTCF sites that lost CTCF upon knockdown displayed increased DSBs when compared to the gained sites, and those lost sites are overrepresented with G-quadruplexes, suggesting that the structures act as boundary insulators in the absence of CTCF, and contribute to increased DSBs. These results model how alternative DNA secondary structures facilitate recruitment of TOP2 to CTCF binding sites, providing mechanistic insight into DNA fragility at CTCF binding sites.

PRRX1-TOP2A interaction is a malignancy-promoting factor in human malignant peripheral nerve sheath tumours.

BACKGROUND: Paired related-homeobox 1 (PRRX1) is a transcription factor in the regulation of developmental morphogenetic processes. There is growing evidence that PRRX1 is highly expressed in certain cancers and is critically involved in human survival prognosis. However, the molecular mechanism of PRRX1 in cancer malignancy remains to be elucidated. METHODS: PRRX1 expression in human Malignant peripheral nerve sheath tumours (MPNSTs) samples was detected immunohistochemically to evaluate survival prognosis. MPNST models with PRRX1 gene knockdown or overexpression were constructed in vitro and the phenotype of MPNST cells was evaluated. Bioinformatics analysis combined with co-immunoprecipitation, mass spectrometry, RNA-seq and structural prediction were used to identify proteins interacting with PRRX1. RESULTS: High expression of PRRX1 was associated with a poor prognosis for MPNST. PRRX1 knockdown suppressed the tumorigenic potential. PRRX1 overexpressed in MPNSTs directly interacts with topoisomerase 2 A (TOP2A) to cooperatively promote epithelial-mesenchymal transition and increase expression of tumour malignancy-related gene sets including mTORC1, KRAS and SRC signalling pathways. Etoposide, a TOP2A inhibitor used in the treatment of MPNST, may exhibit one of its anticancer effects by inhibiting the PRRX1-TOP2A interaction. CONCLUSION: Targeting the PRRX1-TOP2A interaction in malignant tumours with high PRRX1 expression might provide a novel tumour-selective therapeutic strategy.

Escherichia coli-Induced cGLIS3-Mediated Stress Granules Activate the NF-κB Pathway to Promote Intrahepatic Cholangiocarcinoma Progression.

Patients with concurrent intrahepatic cholangiocarcinoma (ICC) and hepatolithiasis generally have poor prognoses. Hepatolithiasis is once considered the primary cause of ICC, although recent insights indicate that bacteria in the occurrence of hepatolithiasis can promote the progression of ICC. By constructing in vitro and in vivo ICC models and patient-derived organoids (PDOs), it is shown that Escherichia coli induces the production of a novel RNA, circGLIS3 (cGLIS3), which promotes tumor growth. cGLIS3 binds to hnRNPA1 and G3BP1, resulting in the assembly of stress granules (SGs) and suppression of hnRNPA1 and G3BP1 ubiquitination. Consequently, the IKKα mRNA is blocked in SGs, decreasing the production of IKKα and activating the NF-κB pathway, which finally results in chemoresistance and produces metastatic phenotypes of ICC. This study shows that a combination of Icaritin (ICA) and gemcitabine plus cisplatin (GP) chemotherapy can be a promising treatment strategy for ICC.

Chromatin damage generated by DNA intercalators leads to degradation of RNA Polymerase II.

In cancer therapy, DNA intercalators are mainly known for their capacity to kill cells by inducing DNA damage. Recently, several DNA intercalators have attracted much interest given their ability to inhibit RNA Polymerase I transcription (BMH-21), evict histones (Aclarubicin) or induce chromatin trapping of FACT (Curaxin CBL0137). Interestingly, these DNA intercalators lack the capacity to induce DNA damage while still retaining cytotoxic effects and stabilize p53. Herein, we report that these DNA intercalators impact chromatin biology by interfering with the chromatin stability of RNA polymerases I, II and III. These three compounds have the capacity to induce degradation of RNA polymerase II and they simultaneously enable the trapping of Topoisomerases TOP2A and TOP2B on the chromatin. In addition, BMH-21 also acts as a catalytic inhibitor of Topoisomerase II, resembling Aclarubicin. Moreover, BMH-21 induces chromatin trapping of the histone chaperone FACT and propels accumulation of Z-DNA and histone eviction, similarly to Aclarubicin and CBL0137. These DNA intercalators have a cumulative impact on general transcription machinery by inducing accumulation of topological defects and impacting nuclear chromatin. Therefore, their cytotoxic capabilities may be the result of compounding deleterious effects on chromatin homeostasis.

Prognosis of polymerase epsilon (POLE) mutation in high-grade endometrioid endometrial cancer: Systematic review and meta-analysis.

BACKGROUND: POLE mutated endometrial carcinomas may represent a subspecific type of tumors harboring a more favorable prognosis. Grade 3 (G3 or high-grade) endometrioid endometrial carcinomas remain a clinical dilemma, with some tumors behaving as the low-grade counterparts and others presenting a more aggressive behavior. OBJECTIVES: To determine the association between POLE mutational status and the overall-survival (OS) and progression-free-survival (PFS) of patients with G3 endometrioid endometrial cancer (EC). We also aimed to determine the prevalence of POLE mutations in G3 endometrioid EC. METHODS: We conducted a systematic review in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines (PROSPERO No: CRD4202340008). We searched the following electronic databases: PubMed/Medline, EMBASE, Cochrane Library, Scopus, and Web of Science. For time-to-event data, the effect of POLE mutation in G3 EC was described using hazard ratios (HRs) and corresponding 95% confidence intervals (CIs). Individual patient data for each study was investigated if available from the study authors. If individual patient data were not available, information regarding time-to-event outcomes was extracted using an appropriate methodology. OS and PFS were analyzed using both one-stage and two-stage approaches, the Kaplan-Meier method, and Cox-proportional hazards models. RESULTS: This systematic review and meta-analysis included 19 studies with 3092 patients who had high-grade endometrioid EC. Patients with POLE mutations had lower risks of death (HR = 0.36, 95% CI 0.26 to 0.50, I2 = 0%, 10 trials) and disease progression (HR = 0.31, 95% CI 0.17 to 0.57, I2 = 33%, 10 trials). The pooled prevalence of POLE mutation was 11% (95% CI 9 to 13, I2 = 68%, 18 studies). CONCLUSION: POLE mutations in high-grade endometrioid EC are associated with a more favorable prognosis with increased OS and PFS.

DNA polymerase ε and δ variants drive mutagenesis in polypurine tracts in human tumors.

Alterations in the exonuclease domain of DNA polymerase ε cause ultramutated cancers. These cancers accumulate AGA>ATA transversions; however, their genomic features beyond the trinucleotide motifs are obscure. We analyze the extended DNA context of ultramutation using whole-exome sequencing data from 524 endometrial and 395 colorectal tumors. We find that G>T transversions in POLE-mutant tumors predominantly affect sequences containing at least six consecutive purines, with a striking preference for certain positions within polypurine tracts. Using this signature, we develop a machine-learning classifier to identify tumors with hitherto unknown POLE drivers and validate two drivers, POLE-E978G and POLE-S461L, by functional assays in yeast. Unlike other pathogenic variants, the E978G substitution affects the polymerase domain of Pol ε. We further show that tumors with POLD1 drivers share the extended signature of POLE ultramutation. These findings expand the understanding of ultramutation mechanisms and highlight peculiar mutagenic properties of polypurine tracts in the human genome.

Properties of biomolecular condensates defined by Activator of G-protein Signaling 3.

Activator of G-protein signaling 3 (AGS3; also known as GPSM1), a receptor-independent activator of G-protein signaling, oscillates among defined subcellular compartments and biomolecular condensates (BMCs) in a regulated manner that is likely related to the functional diversity of the protein. We determined the influence of cell stress on the cellular distribution of AGS3 and core material properties of AGS3 BMCs. Cellular stress (oxidative, pHi and thermal) induced the formation of AGS3 BMCs in HeLa and COS-7 cells, as determined by fluorescent microscopy. Oxidative stress-induced AGS3 BMCs were distinct from G3BP1 stress granules and from RNA processing BMCs defined by the P-body protein Dcp1a. Immunoblots indicated that cellular stress shifted AGS3, but not the stress granule protein G3BP1 to a membrane pellet fraction following cell lysis. The stress-induced generation of AGS3 BMCs was reduced by co-expression of the signaling protein Gαi3, but not the AGS3-binding partner DVL2. Fluorescent recovery following photobleaching of individual AGS3 BMCs indicated that there are distinct diffusion kinetics and restricted fluidity for AGS3 BMCs. These data suggest that AGS3 BMCs represent a distinct class of stress granules that serve as a previously unrecognized signal processing node.

Identification of small molecule inhibitors of G3BP-driven stress granule formation.

Stress granule formation is triggered by the release of mRNAs from polysomes and is promoted by the action of the RNA-binding proteins G3BP1/2. Stress granules have been implicated in several disease states, including cancer and neurodegeneration. Consequently, compounds that limit stress granule formation or promote their dissolution have potential as both experimental tools and novel therapeutics. Herein, we describe two small molecules, G3BP inhibitor a and b (G3Ia and G3Ib), designed to bind to a specific pocket in G3BP1/2 that is targeted by viral inhibitors of G3BP1/2 function. In addition to disrupting the co-condensation of RNA, G3BP1, and caprin 1 in vitro, these compounds inhibit stress granule formation in cells treated prior to or concurrent with stress and dissolve pre-existing stress granules. These effects are consistent across multiple cell types and a variety of initiating stressors. Thus, these compounds represent powerful tools to probe the biology of stress granules and hold promise for therapeutic interventions designed to modulate stress granule formation.

Validation of a one-step genomics-based molecular classifier for endometrial carcinoma in a large Chinese population.

OBJECTIVE: The aim of this study is to delineate the molecular classification features within Chinese endometrial cancer (EC) patients and to evaluate the concurrence between two widely employed methods for diagnosing EC molecular subtypes. METHODS: This retrospective observational cohort study encompassed 479 cases of EC for analysis. Utilizing next-generation sequencing (NGS) panels targeting POLE, TP53, and microsatellite instability (MSI) status, four subtypes [POLE ultramutated (POLE mut), MMR-deficient (MMRd), p53 abnormal (p53abn), and no specific molecular profile (NSMP)] were classified. Immunohistochemistry (IHC) was employed to ascertain the expression of p53 and MMR proteins. RESULTS: Among the 479 patients, the distribution of EC subtypes was as follows: 28 (5.85%) POLE mut, 67 (13.99%) MMRd, 60 (12.53%) p53abn, and 324 (67.64%) NSMP. When compared to published findings on EC subtypes in the Caucasian population, our real-world data on Chinese ECs revealed a notably higher proportion of NSMP/CNL (copy number low). The evaluation of MSI/MMR status through NGS-based and IHC-based methods displayed substantial concordance (Kappa = 0.91). Slight discordance between the two techniques in identifying p53 abnormalities (Kappa = 0.83) might stem from TP53 truncating mutations, cytoplasmic p53 expression, null TP53 mutants, and well-documented challenges in interpreting p53 IHC. CONCLUSIONS: Chinese ECs exhibit distinctive molecular attributes. For accurate molecular subtyping of Chinese ECs, additional molecular markers that align with the Chinese population's characteristics should be incorporated into existing classifiers. The study's outcomes underscore a strong agreement between NGS and IHC in TP53/p53 detection and MSI assessment.

Exploring Co-occurring POLE Exonuclease and Non-exonuclease Domain Mutations and Their Impact on Tumor Mutagenicity.

POLE driver mutations in the exonuclease domain (ExoD driver) are prevalent in several cancers, including colorectal cancer and endometrial cancer, leading to dramatically ultra-high tumor mutation burden (TMB). To understand whether POLE mutations that are not classified as drivers (POLE Variant) contribute to mutagenesis, we assessed TMB in 447 POLE-mutated colorectal cancers, endometrial cancers, and ovarian cancers classified as TMB-high ≥10 mutations/Mb (mut/Mb) or TMB-low <10 mut/Mb. TMB was significantly highest in tumors with "POLE ExoD driver plus POLE Variant" (colorectal cancer and endometrial cancer, P < 0.001; ovarian cancer, P < 0.05). TMB increased with additional POLE variants (P < 0.001), but plateaued at 2, suggesting an association between the presence of these variants and TMB. Integrated analysis of AlphaFold2 POLE models and quantitative stability estimates predicted the impact of multiple POLE variants on POLE functionality. The prevalence of immunogenic neoepitopes was notably higher in the "POLE ExoD driver plus POLE Variant" tumors. Overall, this study reveals a novel correlation between POLE variants in POLE ExoD-driven tumors, and ultra-high TMB. Currently, only select pathogenic ExoD mutations with a reliable association with ultra-high TMB inform clinical practice. Thus, these findings are hypothesis-generating, require functional validation, and could potentially inform tumor classification, treatment responses, and clinical outcomes. SIGNIFICANCE: Somatic POLE ExoD driver mutations cause proofreading deficiency that induces high TMB. This study suggests a novel modifier role for POLE variants in POLE ExoD-driven tumors, associated with ultra-high TMB. These data, in addition to future functional studies, may inform tumor classification, therapeutic response, and patient outcomes.

Profiling of RNA-binding protein binding sites by in situ reverse transcription-based sequencing.

RNA-binding proteins (RBPs) regulate diverse cellular processes by dynamically interacting with RNA targets. However, effective methods to capture both stable and transient interactions between RBPs and their RNA targets are still lacking, especially when the interaction is dynamic or samples are limited. Here we present an assay of reverse transcription-based RBP binding site sequencing (ARTR-seq), which relies on in situ reverse transcription of RBP-bound RNAs guided by antibodies to identify RBP binding sites. ARTR-seq avoids ultraviolet crosslinking and immunoprecipitation, allowing for efficient and specific identification of RBP binding sites from as few as 20 cells or a tissue section. Taking advantage of rapid formaldehyde fixation, ARTR-seq enables capturing the dynamic RNA binding by RBPs over a short period of time, as demonstrated by the profiling of dynamic RNA binding of G3BP1 during stress granule assembly on a timescale as short as 10 minutes.

Race, Prevalence of POLE and POLD1 Alterations, and Survival Among Patients With Endometrial Cancer.

Importance: Black patients with endometrial cancer (EC) in the United States have higher mortality than patients of other races with EC. The prevalence of POLE and POLD1 pathogenic alterations in patients of different races with EC are not well studied. Objective: To explore the prevalence of and outcomes associated with POLE and POLD1 alterations in differential racial groups. Design, Setting, and Participants: This retrospective cohort study incorporated the largest available data set of patients with EC, including American Association for Cancer Research Project GENIE (Genomics Evidence Neoplasia Information Exchange; 5087 participants), Memorial Sloan Kettering-Metastatic Events and Tropisms (1315 participants), and the Cancer Genome Atlas Uterine Corpus Endometrial Carcinoma (517 participants), collected from 2015 to 2023, 2013 to 2021, and 2006 to 2012, respectively. The prevalence of and outcomes associated with POLE or POLD1 alterations in EC were evaluated across self-reported racial groups. Exposure: Patients of different racial groups with EC and with or without POLE or POLD1 alterations. Main Outcomes and Measures: The main outcome was overall survival. Data on demographic characteristics, POLE and POLD1 alteration status, histologic subtype, tumor mutation burden, fraction of genome altered, and microsatellite instability score were collected. Results: A total of 6919 EC cases were studied, of whom 444 (6.4%), 694 (10.0%), and 4869 (70.4%) patients were self-described as Asian, Black, and White, respectively. Within these large data sets, Black patients with EC exhibited a lower weighted average prevalence of pathogenic POLE alterations (0.5% [3 of 590 cases]) compared with Asian (6.1% [26 of 424]) or White (4.6% [204 of 4520]) patients. By contrast, the prevalence of POLD1 pathogenic alterations was 5.0% (21 cases), 3.2% (19 cases), and 5.6% (255 cases) in Asian, Black, and White patients with EC, respectively. Patients with POLD1 alterations had better outcomes regardless of race, histology, and TP53 alteration status. For a total of 241 clinically annotated Black patients with EC, a composite biomarker panel of either POLD1 or POLE alterations identified 7.1% (17 patients) with positive outcomes (1 event at 70 months follow up) in the small sample of available patients. Conclusions and Relevance: In this retrospective clinicopathological study of patients of different racial groups with EC, a composite biomarker panel of either POLD1 or POLE alteration could potentially guide treatment de-escalation, which is especially relevant for Black patients.