Preexisting maternal immunity to AAV but not Cas9 impairs in utero gene editing in mice.

In utero gene editing (IUGE) is a potential treatment for inherited diseases that cause pathology before or soon after birth. Preexisting immunity to adeno-associated virus (AAV) vectors and Cas9 endonuclease may limit postnatal gene editing. The tolerogenic fetal immune system minimizes a fetal immune barrier to IUGE. However, the ability of maternal immunity to limit fetal gene editing remains a question. We investigated whether preexisting maternal immunity to AAV or Cas9 impairs IUGE. Using a combination of fluorescent reporter mice and a murine model of a metabolic liver disease, we demonstrated that maternal anti-AAV IgG antibodies were efficiently transferred from dam to fetus and impaired IUGE in a maternal titer-dependent fashion. By contrast, maternal cellular immunity was inefficiently transferred to the fetus, and neither maternal cellular nor humoral immunity to Cas9 impaired IUGE. Using human umbilical cord and maternal blood samples collected from mid- to late-gestation pregnancies, we demonstrated that maternal-fetal transmission of anti-AAV IgG was inefficient in midgestation compared with term, suggesting that the maternal immune barrier to clinical IUGE would be less relevant at midgestation. These findings support immunologic advantages for IUGE and inform maternal preprocedural testing protocols and exclusion criteria for future clinical trials.

Microtubule forces drive nuclear damage in LMNA cardiomyopathy.

Nuclear homeostasis requires a balance of forces between the cytoskeleton and nucleus. Variants in LMNA disrupt this balance by weakening the nuclear lamina, resulting in nuclear damage in contractile tissues and ultimately muscle disease. Intriguingly, disrupting the LINC complex that connects the cytoskeleton to the nucleus has emerged as a promising strategy to ameliorate LMNA cardiomyopathy. Yet how LINC disruption protects the cardiomyocyte nucleus remains unclear. To address this, we developed an assay to quantify the coupling of cardiomyocyte contraction to nuclear deformation and interrogated its dependence on the lamina and LINC complex. We found that the LINC complex was surprisingly dispensable for transferring the majority of contractile strain into the nucleus, and that increased nuclear strain in Lmna-deficient myocytes was not rescued by LINC disruption. However, LINC disruption eliminated the cage of microtubules encircling the nucleus, and disrupting microtubules was sufficient to prevent nuclear damage induced by LMNA deficiency. Through computational modeling we simulated the mechanical stress fields surrounding cardiomyocyte nuclei and show how microtubule compression exploits local vulnerabilities to damage LMNA-deficient nuclei. Our work pinpoints localized, microtubule-dependent force transmission through the LINC complex as a pathological driver and therapeutic target for LMNA cardiomyopathy.

Epigenetics.

Epigenetics is the study of heritable changes to the genome and gene expression patterns that are not caused by direct changes to the DNA sequence. Examples of these changes include posttranslational modifications to DNA-bound histone proteins, DNA methylation, and remodeling of nuclear architecture. Collectively, epigenetic changes provide a layer of regulation that affects transcriptional activity of genes while leaving DNA sequences unaltered. Sequence variants or mutations affecting enzymes responsible for modifying or sensing epigenetic marks have been identified in patients with congenital heart disease (CHD), and small-molecule inhibitors of epigenetic complexes have shown promise as therapies for adult heart diseases. Additionally, transgenic mice harboring mutations or deletions of genes encoding epigenetic enzymes recapitulate aspects of human cardiac disease. Taken together, these findings suggest that the evolving field of epigenetics will inform our understanding of congenital and adult cardiac disease and offer new therapeutic opportunities.

T2-FLAIR mismatch sign predicts DNA methylation subclass and CDKN2A/B status in IDH-mutant astrocytomas.

PURPOSE: DNA methylation profiling stratifies isocitrate dehydrogenase (IDH)-mutant astrocytomas into methylation low-grade and high-grade groups. We investigated the utility of the T2-FLAIR mismatch sign for predicting DNA methylation grade and cyclin-dependent kinase inhibitor 2A/B (CDKN2A/B) homozygous deletion, a molecular biomarker for grade 4 IDH-mutant astrocytomas, according to the 2021 World Health Organization (WHO) classification. EXPERIMENTAL DESIGN: Preoperative MRI scans of IDH-mutant astrocytomas subclassified by DNA methylation profiling (n=71) were independently evaluated by two radiologists for the T2-FLAIR mismatch sign. The diagnostic utility of T2-FLAIR mismatch in predicting methylation grade, CDKN2A/B status, copy number variation, and survival was analyzed. RESULTS: The T2-FLAIR mismatch sign was present in 21 of 45 (46.7%) methylation low-grade and 1 of 26 (3.9%) methylation high-grade cases (p<0.001), resulting in 96.2% specificity, 95.5% positive predictive value, and 51.0% negative predictive value for predicting low methylation grade. The T2-FLAIR mismatch sign was also significantly associated with intact CDKN2A/B status (p=0.028) with 87.5% specificity, 86.4% positive predictive value, and 42.9% negative predictive value. Overall multivariable Cox analysis showed that retained CDKN2A/B status remained significant for PFS (p=0.01). Multivariable Cox analysis of the histologic grade 3 subset, which was nearly evenly divided by CDKN2A/B status, CNV, and methylation grade, showed trends toward significance for DNA methylation grade with OS (p=0.045) and CDKN2A/B status with PFS (p=0.052). CONCLUSIONS: The T2-FLAIR mismatch sign is highly specific for low methylation grade and intact CDKN2A/B in IDH-mutant astrocytomas.

Leptomeningeal Metastases in IDH-wildtype Glioblastomas Revisited: Comprehensive Analysis of Incidence, Risk Factors, and Prognosis Based on Post-contrast FLAIR.

BACKGROUND: The incidence of leptomeningeal metastases (LM) has been reported diversely. This study aimed to investigate the incidence, risk factors, and prognosis of LM in patients with IDH-wildtype glioblastoma. METHODS: A total of 828 patients with IDH-wildtype glioblastoma were enrolled between 2005 and 2022. Baseline preoperative MRI including post-contrast fluid-attenuated inversion recovery (FLAIR) was used for LM diagnosis. Qualitative and quantitative features, including distance between tumor and subventricular zone (SVZ) and tumor volume by automatic segmentation of the lateral ventricles and tumor, were assessed. Logistic analysis of LM development was performed using clinical, molecular, and imaging data. Survival analysis was performed. RESULTS: The incidence of LM was 11.4%. MGMTp unmethylation (odds ratio [OR] = 1.92, P = 0.014), shorter distance between tumor and SVZ (OR = 0.94, P = 0.010), and larger contrast-enhancing tumor volume (OR = 1.02, P < 0.001) were significantly associated with LM. The overall survival (OS) was significantly shorter in patients with LM than in those without (log-rank test; P < 0.001), with median OS of 12.2 and 18.5 months, respectively. Presence of LM remained an independent prognostic factor for OS in IDH-wildtype glioblastoma (hazard ratio = 1.42, P = 0.011), along with other clinical, molecular, imaging, and surgical prognostic factors. CONCLUSION: The incidence of LM is high in patients with IDH-wildtype glioblastoma, and aggressive molecular and imaging factors are correlated with LM development. The prognostic significance of LM based on post-contrast FLAIR imaging suggests acknowledgement of post-contrast FLAIR as a reliable diagnostic tool for clinicians.

Two-Stage Training Framework Using Multicontrast MRI Radiomics for IDH Mutation Status Prediction in Glioma.

Purpose To develop a radiomics framework for preoperative MRI-based prediction of isocitrate dehydrogenase (IDH) mutation status, a crucial glioma prognostic indicator. Materials and Methods Radiomics features (shape, first-order statistics, and texture) were extracted from the whole tumor or the combination of nonenhancing, necrosis, and edema regions. Segmentation masks were obtained via the federated tumor segmentation tool or the original data source. Boruta, a wrapper-based feature selection algorithm, identified relevant features. Addressing the imbalance between mutated and wild-type cases, multiple prediction models were trained on balanced data subsets using random forest or XGBoost and assembled to build the final classifier. The framework was evaluated using retrospective MRI scans from three public datasets (The Cancer Imaging Archive [TCIA, 227 patients], the University of California San Francisco Preoperative Diffuse Glioma MRI dataset [UCSF, 495 patients], and the Erasmus Glioma Database [EGD, 456 patients]) and internal datasets collected from the University of Texas Southwestern Medical Center (UTSW, 356 patients), New York University (NYU, 136 patients), and University of Wisconsin-Madison (UWM, 174 patients). TCIA and UTSW served as separate training sets, while the remaining data constituted the test set (1617 or 1488 testing cases, respectively). Results The best performing models trained on the TCIA dataset achieved area under the receiver operating characteristic curve (AUC) values of 0.89 for UTSW, 0.86 for NYU, 0.93 for UWM, 0.94 for UCSF, and 0.88 for EGD test sets. The best performing models trained on the UTSW dataset achieved slightly higher AUCs: 0.92 for TCIA, 0.88 for NYU, 0.96 for UWM, 0.93 for UCSF, and 0.90 for EGD. Conclusion This MRI radiomics-based framework shows promise for accurate preoperative prediction of IDH mutation status in patients with glioma. Keywords: Glioma, Isocitrate Dehydrogenase Mutation, IDH Mutation, Radiomics, MRI Supplemental material is available for this article. Published under a CC BY 4.0 license. See also commentary by Moassefi and Erickson in this issue.

Synthesizing 3D Multi-Contrast Brain Tumor MRIs Using Tumor Mask Conditioning.

Data scarcity and data imbalance are two major challenges in training deep learning models on medical images, such as brain tumor MRI data. The recent advancements in generative artificial intelligence have opened new possibilities for synthetically generating MRI data, including brain tumor MRI scans. This approach can be a potential solution to mitigate the data scarcity problem and enhance training data availability. This work focused on adapting the 2D latent diffusion models to generate 3D multi-contrast brain tumor MRI data with a tumor mask as the condition. The framework comprises two components: a 3D autoencoder model for perceptual compression and a conditional 3D Diffusion Probabilistic Model (DPM) for generating high-quality and diverse multi-contrast brain tumor MRI samples, guided by a conditional tumor mask. Unlike existing works that focused on generating either 2D multi-contrast or 3D single-contrast MRI samples, our models generate multi-contrast 3D MRI samples. We also integrated a conditional module within the UNet backbone of the DPM to capture the semantic class-dependent data distribution driven by the provided tumor mask to generate MRI brain tumor samples based on a specific brain tumor mask. We trained our models using two brain tumor datasets: The Cancer Genome Atlas (TCGA) public dataset and an internal dataset from the University of Texas Southwestern Medical Center (UTSW). The models were able to generate high-quality 3D multi-contrast brain tumor MRI samples with the tumor location aligned by the input condition mask. The quality of the generated images was evaluated using the Fréchet Inception Distance (FID) score. This work has the potential to mitigate the scarcity of brain tumor data and improve the performance of deep learning models involving brain tumor MRI data.

Sex-specific Differences in IDH1-Wildtype Glioblastoma patients in the ReSPOND Consortium.

BACKGROUND: Understanding sex-based differences in glioblastoma patients is necessary for accurate personalized treatment planning to improve patient outcomes. PURPOSE: To investigate sex-specific differences in molecular, clinical and radiological tumor parameters, as well as survival outcomes in glioblastoma, isocitrate dehydrogenase-1 wildtype (IDH1-WT), grade 4 patients. METHODS: Retrospective data of 1832 glioblastoma, IDH1-WT patients with comprehensive information on tumor parameters was acquired from the Radiomics Signatures for Precision Oncology in Glioblastoma (ReSPOND) consortium. Data imputation was performed for missing values. Sex-based differences in tumor parameters, such as, age, molecular parameters, pre-operative KPS score, tumor volumes, epicenter and laterality were assessed through non-parametric tests. Spatial atlases were generated using pre-operative MRI maps to visualize tumor characteristics. Survival time analysis was performed through log-rank tests and Cox proportional hazard analyses. RESULTS: GBM was diagnosed at a median age of 64 years in females compared to 61.9 years in males (FDR = 0.003). Males had a higher Karnofsky Performance Score (above 80) as compared to females (60.4% females Vs 69.7% males, FDR = 0.044). Females had lower tumor volumes in enhancing (16.7 cm3 Vs. 20.6 cm3 in males, FDR = 0.001), necrotic core (6.18 cm3 Vs. 7.76 cm3 in males, FDR = 0.001) and edema regions (46.9 cm3 Vs. 59.2 cm3 in males, FDR = 0.0001). Right temporal region was the most common tumor epicenter in the overall population. Right as well as left temporal lobes were more frequently involved in males. There were no significant differences in survival outcomes and mortality ratios. Higher age, unmethylated O6-methylguanine-DNAmethyltransferase (MGMT) promoter and undergoing subtotal resection increased the mortality risk in both males and females. CONCLUSIONS: Our study demonstrates significant sex-based differences in clinical and radiological tumor parameters of glioblastoma, IDH1-WT, grade 4 patients. Sex is not an independent prognostic factor for survival outcomes and the tumor parameters influencing patient outcomes are identical for males and females. ABBREVIATIONS: IDH1-WT = isocitrate dehydrogenase-1 wildtype; MGMTp = O6-methylguanine-DNA-methyltransferase promoter; KPS = Karnofsky performance score; EOR = extent of resection; WHO = world health organization; FDR = false discovery rate.

Listeria monocytogenes brain abscesses presenting as contiguous, tubular rim-enhancing lesions on Magnetic Resonance Imaging: Case series and literature review.

Listeriosis has more than a 50% mortality when the central nervous system is involved, necessitating rapid diagnosis and treatment. We present four patients with brain abscesses in the setting of diagnosed neurolisteriosis, all of which demonstrated an odd presentation of multiple small, contiguous tubular lesions with rim enhancement on magnetic resonance imaging. Our review of published cases of neurolisteriosis suggests that this may be a useful pattern to identify neurolisteriosis abscesses, allowing earlier detection and therapy.

Characterization of human extreme heat exposure using an outdoor thermal manikin.

Extreme heat is a current and growing global health concern. Current heat exposure models include meteorological and human factors that dictate heat stress, comfort, and risk of illness. However, radiation models simplify the human body to a cylinder, while convection ones provide conflicting predictions. To address these issues, we introduce a new method to characterize human exposure to extreme heat with unprecedented detail. We measure heat loads on 35 body surface zones using an outdoor thermal manikin ("ANDI") alongside an ultrasonic anemometer array and integral radiation measurements (IRM). We show that regardless of body orientation, IRM and ANDI agree even under high solar conditions. Further, body parts can be treated as cylinders, even in highly turbulent flow. This geometry-rooted insight yields a whole-body convection correlation that resolves prior conflicts and is valid for diverse indoor and outdoor wind flows. Results will inform decision-making around heat protection, adaptation, and mitigation.

Multinodular and Vacuolating Neuronal Tumor-like Lesion of the Spinal Cord: Two Case Reports.

We describe 2 cases of a spinal cord lesion with imaging features closely resembling those described in supratentorial multinodular and vacuolating neuronal tumor (MVNT) or infratentorial multinodular and vacuolating posterior fossa lesions of unknown significance. Multiple well-delineated nonenhancing T2-hyperintense intramedullary cystic ovoid nodules were visualized within the white matter of the spinal cord, including some immediately abutting the gray matter. No alterations in signal intensity or morphology were detected in a follow-up. Moreover, no relevant clinical symptoms attributable to the lesions were present. We describe these lesions as presumed MVNT, and we therefore use the term MVNT-like spinal cord lesions.

Prognostic value of DNA methylation subclassification, aneuploidy, and CDKN2A/B homozygous deletion in predicting clinical outcome of IDH mutant astrocytomas.

BACKGROUND: Isocitrate dehydrogenase (IDH) mutant astrocytoma grading, until recently, has been entirely based on morphology. The 5th edition of the Central Nervous System World Health Organization (WHO) introduces CDKN2A/B homozygous deletion as a biomarker of grade 4. We sought to investigate the prognostic impact of DNA methylation-derived molecular biomarkers for IDH mutant astrocytoma. METHODS: We analyzed 98 IDH mutant astrocytomas diagnosed at NYU Langone Health between 2014 and 2022. We reviewed DNA methylation subclass, CDKN2A/B homozygous deletion, and ploidy and correlated molecular biomarkers with histological grade, progression free (PFS), and overall (OS) survival. Findings were confirmed using 2 independent validation cohorts. RESULTS: There was no significant difference in OS or PFS when stratified by histologic WHO grade alone, copy number complexity, or extent of resection. OS was significantly different when patients were stratified either by CDKN2A/B homozygous deletion or by DNA methylation subclass (P value = .0286 and .0016, respectively). None of the molecular biomarkers were associated with significantly better PFS, although DNA methylation classification showed a trend (P value = .0534). CONCLUSIONS: The current WHO recognized grading criteria for IDH mutant astrocytomas show limited prognostic value. Stratification based on DNA methylation shows superior prognostic value for OS.

MRI-Based Deep Learning Method for Classification of IDH Mutation Status.

Isocitrate dehydrogenase (IDH) mutation status has emerged as an important prognostic marker in gliomas. This study sought to develop deep learning networks for non-invasive IDH classification using T2w MR images while comparing their performance to a multi-contrast network. Methods: Multi-contrast brain tumor MRI and genomic data were obtained from The Cancer Imaging Archive (TCIA) and The Erasmus Glioma Database (EGD). Two separate 2D networks were developed using nnU-Net, a T2w-image-only network (T2-net) and a multi-contrast network (MC-net). Each network was separately trained using TCIA (227 subjects) or TCIA + EGD data (683 subjects combined). The networks were trained to classify IDH mutation status and implement single-label tumor segmentation simultaneously. The trained networks were tested on over 1100 held-out datasets including 360 cases from UT Southwestern Medical Center, 136 cases from New York University, 175 cases from the University of Wisconsin-Madison, 456 cases from EGD (for the TCIA-trained network), and 495 cases from the University of California, San Francisco public database. A receiver operating characteristic curve (ROC) was drawn to calculate the AUC value to determine classifier performance. Results: T2-net trained on TCIA and TCIA + EGD datasets achieved an overall accuracy of 85.4% and 87.6% with AUCs of 0.86 and 0.89, respectively. MC-net trained on TCIA and TCIA + EGD datasets achieved an overall accuracy of 91.0% and 92.8% with AUCs of 0.94 and 0.96, respectively. We developed reliable, high-performing deep learning algorithms for IDH classification using both a T2-image-only and a multi-contrast approach. The networks were tested on more than 1100 subjects from diverse databases, making this the largest study on image-based IDH classification to date.

Clinical Efficacy of ONC201 in H3K27M-Mutant Diffuse Midline Gliomas Is Driven by Disruption of Integrated Metabolic and Epigenetic Pathways.

Patients with H3K27M-mutant diffuse midline glioma (DMG) have no proven effective therapies. ONC201 has recently demonstrated efficacy in these patients, but the mechanism behind this finding remains unknown. We assessed clinical outcomes, tumor sequencing, and tissue/cerebrospinal fluid (CSF) correlate samples from patients treated in two completed multisite clinical studies. Patients treated with ONC201 following initial radiation but prior to recurrence demonstrated a median overall survival of 21.7 months, whereas those treated after recurrence had a median overall survival of 9.3 months. Radiographic response was associated with increased expression of key tricarboxylic acid cycle-related genes in baseline tumor sequencing. ONC201 treatment increased 2-hydroxyglutarate levels in cultured H3K27M-DMG cells and patient CSF samples. This corresponded with increases in repressive H3K27me3 in vitro and in human tumors accompanied by epigenetic downregulation of cell cycle regulation and neuroglial differentiation genes. Overall, ONC201 demonstrates efficacy in H3K27M-DMG by disrupting integrated metabolic and epigenetic pathways and reversing pathognomonic H3K27me3 reduction. SIGNIFICANCE: The clinical, radiographic, and molecular analyses included in this study demonstrate the efficacy of ONC201 in H3K27M-mutant DMG and support ONC201 as the first monotherapy to improve outcomes in H3K27M-mutant DMG beyond radiation. Mechanistically, ONC201 disrupts integrated metabolic and epigenetic pathways and reverses pathognomonic H3K27me3 reduction. This article is featured in Selected Articles from This Issue, p. 2293.

Association of partial T2-FLAIR mismatch sign and isocitrate dehydrogenase mutation in WHO grade 4 gliomas: results from the ReSPOND consortium.

PURPOSE: While the T2-FLAIR mismatch sign is highly specific for isocitrate dehydrogenase (IDH)-mutant, 1p/19q-noncodeleted astrocytomas among lower-grade gliomas, its utility in WHO grade 4 gliomas is not well-studied. We derived the partial T2-FLAIR mismatch sign as an imaging biomarker for IDH mutation in WHO grade 4 gliomas. METHODS: Preoperative MRI scans of adult WHO grade 4 glioma patients (n = 2165) from the multi-institutional ReSPOND (Radiomics Signatures for PrecisiON Diagnostics) consortium were analyzed. Diagnostic performance of the partial T2-FLAIR mismatch sign was evaluated. Subset analyses were performed to assess associations of imaging markers with overall survival (OS). RESULTS: One hundred twenty-one (5.6%) of 2165 grade 4 gliomas were IDH-mutant. Partial T2-FLAIR mismatch was present in 40 (1.8%) cases, 32 of which were IDH-mutant, yielding 26.4% sensitivity, 99.6% specificity, 80.0% positive predictive value, and 95.8% negative predictive value. Multivariate logistic regression demonstrated IDH mutation was significantly associated with partial T2-FLAIR mismatch (odds ratio [OR] 5.715, 95% CI [1.896, 17.221], p = 0.002), younger age (OR 0.911 [0.895, 0.927], p < 0.001), tumor centered in frontal lobe (OR 3.842, [2.361, 6.251], p < 0.001), absence of multicentricity (OR 0.173, [0.049, 0.612], p = 0.007), and presence of cystic (OR 6.596, [3.023, 14.391], p < 0.001) or non-enhancing solid components (OR 6.069, [3.371, 10.928], p < 0.001). Multivariate Cox analysis demonstrated cystic components (p = 0.024) and non-enhancing solid components (p = 0.003) were associated with longer OS, while older age (p < 0.001), frontal lobe center (p = 0.008), multifocality (p < 0.001), and multicentricity (p < 0.001) were associated with shorter OS. CONCLUSION: Partial T2-FLAIR mismatch sign is highly specific for IDH mutation in WHO grade 4 gliomas.

Occipital Nocardia Abscess Presenting With Positive Visual Phenomenon and Quadrantanopsia.

ABSTRACT: A 74-year-old man with chronic obstructive pulmonary disease, glaucoma, and Stage IIIB squamous cell lung cancer experienced several minutes of flashing lights in his right visual hemifield, followed by onset of a right visual field defect. On examination, the patient had a right homonymous hemianopsia that was most dense inferiorly by confrontation testing. Emergent CT scan of the head revealed a 2.5 × 3 cm hypodensity in the left occipital lobe, which was interpreted as an acute stroke. Continuous EEG monitoring captured left posterior quadrant seizures that were temporally correlated to the positive visual phenomena. Subsequent MRI of the brain with and without contrast revealed a conglomerate of centrally necrotic and peripherally enhancing mass lesions. On biopsy, a thick purulent material was drained and Gram stain of the sample revealed gram-positive beaded rods, which speciated to Nocardia farcinica . The patient was treated with a six-week course of intravenous meropenem and a one-year course of oral trimethroprim-sulfamethoxazole. On follow-up, the patient experienced resolution of the right visual field deficit.

Ionizable Lipid Nanoparticles for Therapeutic Base Editing of Congenital Brain Disease.

Delivery of mRNA-based therapeutics to the perinatal brain holds great potential in treating congenital brain diseases. However, nonviral delivery platforms that facilitate nucleic acid delivery in this environment have yet to be rigorously studied. Here, we screen a diverse library of ionizable lipid nanoparticles (LNPs) via intracerebroventricular (ICV) injection in both fetal and neonatal mice and identify an LNP formulation with greater functional mRNA delivery in the perinatal brain than an FDA-approved industry standard LNP. Following in vitro optimization of the top-performing LNP (C3 LNP) for codelivery of an adenine base editing platform, we improve the biochemical phenotype of a lysosomal storage disease in the neonatal mouse brain, exhibit proof-of-principle mRNA brain transfection in vivo in a fetal nonhuman primate model, and demonstrate the translational potential of C3 LNPs ex vivo in human patient-derived brain tissues. These LNPs may provide a clinically translatable platform for in utero and postnatal mRNA therapies including gene editing in the brain.

High-throughput Oligopaint screen identifies druggable 3D genome regulators.

The human genome functions as a three-dimensional chromatin polymer, driven by a complex collection of chromosome interactions1-3. Although the molecular rules governing these interactions are being quickly elucidated, relatively few proteins regulating this process have been identified. Here, to address this gap, we developed high-throughput DNA or RNA labelling with optimized Oligopaints (HiDRO)-an automated imaging pipeline that enables the quantitative measurement of chromatin interactions in single cells across thousands of samples. By screening the human druggable genome, we identified more than 300 factors that influence genome folding during interphase. Among these, 43 genes were validated as either increasing or decreasing interactions between topologically associating domains. Our findings show that genetic or chemical inhibition of the ubiquitous kinase GSK3A leads to increased long-range chromatin looping interactions in a genome-wide and cohesin-dependent manner. These results demonstrate the importance of GSK3A signalling in nuclear architecture and the use of HiDRO for identifying mechanisms of spatial genome organization.