WDR5 represents a therapeutically exploitable target for cancer stem cells in glioblastoma.

Glioblastomas (GBMs) are heterogeneous, treatment-resistant tumors driven by populations of cancer stem cells (CSCs). However, few molecular mechanisms critical for CSC population maintenance have been exploited for therapeutic development. We developed a spatially resolved loss-of-function screen in GBM patient-derived organoids to identify essential epigenetic regulators in the SOX2-enriched, therapy-resistant niche and identified WDR5 as indispensable for this population. WDR5 is a component of the WRAD complex, which promotes SET1 family-mediated Lys4 methylation of histone H3 (H3K4me), associated with positive regulation of transcription. In GBM CSCs, WDR5 inhibitors blocked WRAD complex assembly and reduced H3K4 trimethylation and expression of genes involved in CSC-relevant oncogenic pathways. H3K4me3 peaks lost with WDR5 inhibitor treatment occurred disproportionally on POU transcription factor motifs, including the POU5F1(OCT4)::SOX2 motif. Use of a SOX2/OCT4 reporter demonstrated that WDR5 inhibitor treatment diminished cells with high reporter activity. Furthermore, WDR5 inhibitor treatment and WDR5 knockdown altered the stem cell state, disrupting CSC in vitro growth and self-renewal, as well as in vivo tumor growth. These findings highlight the role of WDR5 and the WRAD complex in maintaining the CSC state and provide a rationale for therapeutic development of WDR5 inhibitors for GBM and other advanced cancers.

Near-infrared imaging in orthodontic intraoral scanners for early interproximal caries detection.

INTRODUCTION: Intraoral scanners commonly used in orthodontic offices now offer near-infrared imaging (NIRI) technology, advertised as a screening tool to identify interproximal caries. This study aimed to evaluate the reliability and validity of NIRI detection of interproximal carious lesions in a common intraoral scanner (iTero Element 5D; Align Technology, San Jose, Calif) with and without bitewing radiograph complement, compared with a microcomputed tomography (micro-CT) reference standard. METHODS: Extracted human posterior teeth (premolars and molars) were selected for early (noncavitated) interproximal carious lesions (n = 39) and sound control surfaces (n = 47). The teeth were scanned via micro-CT for evaluation by 2 blinded evaluators using consensus scoring. The teeth were mounted to simulate anatomic interproximal contacts and underwent a NIRI scan using iTero Element 5D and bitewing radiographs. Two trained, calibrated examiners independently evaluated (1) near-infrared images alone with clinical photograph, (2) bitewing radiograph alone with clinical photograph, and (3) near-infrared images with bitewing radiograph and clinical photograph in combination, after at least a 10-day washout period between each evaluation. RESULTS: Interrater reliability was highest for NIRI alone (k = 0.533) compared with bitewing radiograph alone (k = 0.176) or in combination (k = 0.256). NIRI alone showed high specificity (0.83-0.96) and moderate sensitivity (0.42-0.63) compared with a micro-CT reference standard. Dentin lesions were significantly more reliably detected than enamel lesions. CONCLUSIONS: After rigorous training and calibration, NIRI can be used with moderate reliability, high specificity, and moderate sensitivity to detect noncavitated interproximal carious lesions.

Influence of BMI percentile on craniofacial morphology and development in adolescents,Part II: elevated BMI is associated with larger final facial dimensions.

BACKGROUND: Prevalence of adolescent obesity has markedly increased from 5.2% in 1974 to 19.7% in 2021. Understanding the impacts of obesity is important to orthodontists, as growth acceleration and greater pre-pubertal facial dimensions are seen in children with elevated body mass index (BMI). METHODS: To identify whether adolescent obesity shifts the timing and rate of craniofacial growth resulting in larger post-treatment dimensions, we evaluated cephalometric outcomes in overweight/obese (BMI > 85%, n = 168) and normal weight (n = 158) adolescents (N = 326 total). Cephalometric measurements were obtained from pre- and post-treatment records to measure growth rates and final dimensions and were statistically evaluated with repeated measures analysis of variance and linear regression models. RESULTS: Overweight and obese adolescents began and finished treatment with significantly larger, bimaxillary prognathic craniofacial dimensions, with elevated mandibular length [articulare-gnathion (Ar-Gn)], maxillary length [condylion-anterior nasal spine (Co-ANS), posterior nasal spine-ANS (PNS-ANS)], and anterior lower face height (ANS-Me), suggesting overweight children grow more overall. However, there was no difference between weight cohorts in the amount of cephalometric change during treatment, and regression analyses demonstrated no correlation between change in growth during treatment and BMI. BMI percentile was a significant linear predictor (P < 0.05) for cephalometric post-treatment outcomes, including Ar-Gn, Co-ANS, ANS-Me, upper face height percentage (UFH:total FH, inverse relationship), lower face height percentage (LFH:total FH), sella-nasion-A-point (SNA), and SN-B-point (SNB). LIMITATIONS: The study is retrospective. CONCLUSIONS: Growth begins earlier in overweight and obese adolescents and continues at a rate similar to normal-weight children during orthodontic treatment, resulting in larger final skeletal dimensions. Orthodontics could begin earlier in overweight patients to time care with growth, and clinicians can anticipate that overweight/obese patients will finish treatment with proportionally larger, bimaxillary-prognathic craniofacial dimensions.

Roadmap on plasticity and epigenetics in cancer.

The role of plasticity and epigenetics in shaping cancer evolution and response to therapy has taken center stage with recent technological advances including single cell sequencing. This roadmap article is focused on state-of-the-art mathematical and experimental approaches to interrogate plasticity in cancer, and addresses the following themes and questions: is there a formal overarching framework that encompasses both non-genetic plasticity and mutation-driven somatic evolution? How do we measure and model the role of the microenvironment in influencing/controlling non-genetic plasticity? How can we experimentally study non-genetic plasticity? Which mathematical techniques are required or best suited? What are the clinical and practical applications and implications of these concepts?

New IDH1 mutant inhibitors for treatment of acute myeloid leukemia.

Neomorphic mutations in isocitrate dehydrogenase 1 (IDH1) are driver mutations in acute myeloid leukemia (AML) and other cancers. We report the development of new allosteric inhibitors of mutant IDH1. Crystallographic and biochemical results demonstrated that compounds of this chemical series bind to an allosteric site and lock the enzyme in a catalytically inactive conformation, thereby enabling inhibition of different clinically relevant IDH1 mutants. Treatment of IDH1 mutant primary AML cells uniformly led to a decrease in intracellular 2-HG, abrogation of the myeloid differentiation block and induction of granulocytic differentiation at the level of leukemic blasts and more immature stem-like cells, in vitro and in vivo. Molecularly, treatment with the inhibitors led to a reversal of the DNA cytosine hypermethylation patterns caused by mutant IDH1 in the cells of individuals with AML. Our study provides proof of concept for the molecular and biological activity of novel allosteric inhibitors for targeting different mutant forms of IDH1 in leukemia.

Identifying factors influencing orthodontic residency program selection.

PURPOSE: The desirability of Orthodontic Residency remains high today relative to the past 30 years. This investigation seeks to re-assess factors influencing Orthodontic Residency program selection amidst contemporary challenges and changes, including increasing student debt, reduced residency stipends, and shifting practice models. METHODS: This mixed-methods study consisted of interviews (qualitative) and surveys (quantitative). Fifteen final-year dental students and first-year orthodontic residents were interviewed one-on-one following a topic guide and then transcripts were analyzed using MAXQDA2022 to identify values, factors, and influences related to program selection and ranking. Qualitative findings and previous studies provided the basis for a survey distributed to residency applicants in 2020-22. Data were analyzed with bivariate and descriptive statistics with stratification by debt group. RESULTS: Interviews (N = 15) elevated the importance of location, interview experience, program cost, and clinical education. Surveys (N = 239) identified the most desirable factors for program selection: satisfied residents, strong clinical training with multiple techniques, good interview impressions, low cost, high patient numbers, a strong reputation, and good clinical facilities with new technology. Less desirable factors include programs that are hospital-based, certificate-only, research-intensive, and require considerable after-hours work. CONCLUSIONS: Clinical education and interview experiences are key for residency selection, consistent with prior studies, though program cost has grown in importance. Findings can help guide orthodontic programs in recruiting applicants and suggest a need to limit educational costs.

FBXO42 activity is required to prevent mitotic arrest, spindle assembly checkpoint activation and lethality in glioblastoma and other cancers.

Glioblastoma (GBM) is the most common and aggressive brain tumor in adults. To identify genes differentially required for the viability of GBM stem-like cells (GSCs), we performed functional genomic lethality screens comparing GSCs and control human neural stem cells. Among top-scoring hits in a subset of GBM cells was the F-box-containing gene FBXO42, which was also predicted to be essential in ∼15% of cell lines derived from a broad range of cancers. Mechanistic studies revealed that, in sensitive cells, FBXO42 activity prevents chromosome alignment defects, mitotic cell cycle arrest and cell death. The cell cycle arrest, but not the cell death, triggered by FBXO42 inactivation could be suppressed by brief exposure to a chemical inhibitor of Mps1, a key spindle assembly checkpoint (SAC) kinase. FBXO42's cancer-essential function requires its F-box and Kelch domains, which are necessary for FBXO42's substrate recognition and targeting by SCF (SKP1-CUL1-F-box protein) ubiquitin ligase complex. However, none of FBXO42's previously proposed targets, including ING4, p53 and RBPJ, were responsible for the observed phenotypes. Instead, our results suggest that FBOX42 alters the activity of one or more proteins that perturb chromosome-microtubule dynamics in cancer cells, which in turn leads to induction of the SAC and cell death.

Benzimidazole F167Y polymorphism in the canine hookworm, Ancylostoma caninum: Widespread geographic, seasonal, age, and breed distribution in United States and Canada dogs.

Surveillance data for Ancylostoma spp. and the A. caninum benzimidazole treatment resistance associated F167Y polymorphism using molecular diagnostics was obtained in a large population of dogs from the United States and Canada. Real-time PCR (qPCR) for Ancylostoma spp. and allele-specific qPCR detecting a single nucleotide polymorphism (SNP) F167Y was used in 262,872 canine stool samples collected between March and December of 2022. Ancylostoma spp. was found at an overall prevalence of 2.5% (6538/262,872), with the highest prevalence in the Southern US, 4.4% (4490/103,095), and the lowest prevalence in Canada 0.6% (101/15,829). The A. caninum F167Y polymorphism was found with the highest prevalence (13.4%, n = 46/343) in the Western US and the lowest in Canada at 4.1% (4/97). The F167Y polymorphism was detected every month over the 10-month collection period. Seasonal distribution showed a peak in June for both Ancylostoma spp. (3.08%, 547/17,775) and A. caninum F167Y (12.25%, 67/547). However, the A. caninum F167Y polymorphism prevalence was highest in September (13.9%, 119/856). Age analysis indicates a higher prevalence of both hookworm infections and occurrence of resistant isolates in puppies. The breeds with the highest F167Y polymorphism prevalence in Ancylostoma spp. detected samples were poodles (28.9%), followed by Bernese Mountain dogs (25%), Cocker spaniels (23.1%), and greyhounds (22.4%). Our data set describes widespread geographic distribution of the A. caninum benzimidazole resistance associated F167Y polymorphism in the United States and Canada, with no clear seasonality compared to the Ancylostoma spp. prevalence patterns. The F167 polymorphism was present in all geographic areas with detected hookworms, including Canada. Our study highlights that the F167Y polymorphism is represented in many dog breeds, including greyhounds.

The in vitro efficacy of betadine antiseptic solution and colloidal silver gel combination in inhibiting the growth of bacterial biofilms.

INTRODUCTION: Betadine (Povidone-Iodine) solution is a topically applied antiseptic, which has been used routinely used in wound care and general surgery to prevent skin and wound infections. However, several studies have documented the ineffectiveness of betadine. Other topical antimicrobial dressings, including those that contain silver, have been used in the management of infected wounds. The present study was undertaken to determine if the combination of 5% betadine solution and silver colloidal gel (Ag-gel) is more effective than either substance alone in inhibiting the growth gram-negative and gram-positive bacteria. METHODS: The effectiveness of 5% betadine solution and Ag-gel as anti-microbial agents were assessed using both colony forming unit (CFU) assay and confocal laser scanning microscopy (CLSM). RESULTS: Ag-gel showed complete inhibition on all the bacteria species examined except the Klebsiella pneumoniae clinical isolate (CL) strain while 5% betadine concentrations did not completely kill any of the tested bacteria. In contrast, K. pneumoniae was completely eliminated in the presence of both 5% betadine solution and Ag-gel together. The CLSM showed similar findings to the CFU results examining the 5% betadine solution and Ag-gel combination. CONCLUSIONS: This study demonstrated that while the individual treatments using either 5% betadine solution and Ag-gel alone were infective antimicrobial agents, the combination of 5% betadine solution and Ag-gel was superior at eliminating all tested bacteria, including K. pneumoniae CL.

Reducing Specimen Rejection Rates Using Concentration-Dependent Hemolysis Rejection Thresholds.

BACKGROUND: Using middleware solutions, it is possible to implement concentration-dependent analyte-specific hemolysis rejection limits. This makes day-to-day reporting of clinical specimens more efficient and potentially lowers sample rejection rates compared to a "one-size-fits-all" approach (i.e., solely based on a single cutoff provided in the package insert). METHODS: Hemolysis interference studies were performed at multiple analyte concentrations for three frequently ordered tests. For each assay, concentration-dependent hemolysis rejection limits were designed based on the total allowable error (TAE) for the analyte as well as the clinical significance of such incurred inaccuracy at the respective concentrations. In general, the following rationale was used: if the interference exceeds 10% (or package insert cutoffs), a comment is placed on the result. If the interference exceeds the TAE, the result will not be reported. Reduction in specimen rejection rates were estimated by comparing the incurred specimen rejection rates when package inserts' vs concentration-dependent hemolysis interference limits were applied to a data set in our institute during a three-month period. RESULTS: Concentration-dependent analyte-specific hemolysis rejection thresholds were designed for three commonly ordered assays that are especially susceptible to hemolysis interference. It is estimated that these novel thresholds for aspartate aminotransferase (AST), lactate dehydrogenase (LDH), and direct bilirubin (DBIL) reduced specimen rejection rates from 9.3% to 1.3%, 31.4% to 4.8%, and 19.9% to 7.1%, respectively. CONCLUSIONS: Concentration-dependent analyte-specific hemolysis rejection thresholds for three commonly ordered assays can reduce rejection rates without significantly compromising the quality of test results.

Novel molecular diagnostic (PCR) diagnosis and outcome of intestinal Echinococcus multilocularis in a dog from western Canada.

OBJECTIVE: To describe the novel PCR diagnosis and outcome of intestinal Echinococcus multilocularis in a dog. ANIMAL: A 13-month-old female intact dog with naturally occurring intestinal E multilocularis. CLINICAL PRESENTATION, PROGRESSION, AND PROCEDURES: The 13-month-old dog initially presented with a reduced appetite and weight loss and then developed hematochezia. The clinical history included a lack of endoparasite preventive care (fecal testing, deworming), exposure to coyotes, fox, sheep, and rodents and the dog had intermittently been fed a raw food diet. Physical examination revealed a thin dog, with a 2/9 body condition score, that was otherwise clinically unremarkable. A fecal sample was submitted for screening for gastrointestinal parasites as part of an infectious disease assessment. The fecal PCR test reported detection of E multilocularis. This result was sequenced as the European haplotype E3/E4. Centrifugal flotation (same sample) did not detect taeniid eggs. TREATMENT AND OUTCOME: The dog was treated with metronidazole, maropitant, and milbemycin oxime/praziquantel. Clinical improvement was noted within 48 hours. No DNA of E multilocularis was detected in a fecal sample collected approximately 10 days after treatment. The dog's owner was advised to provide monthly deworming (praziquantel) for all dogs on the property and to contact their human health-care provider due to potential zoonotic exposure risk. CLINICAL RELEVANCE: Increasing detection of E multilocularis is occurring in dogs in Canada and the US. Alveolar echinococcosis can cause severe disease in dogs and humans. Fecal PCR detection and surveillance may alert practitioners to canine intestinal cases and allow dogs to serve as sentinels for human exposure risk.

Comparative study of a broad qPCR panel and centrifugal flotation for detection of gastrointestinal parasites in fecal samples from dogs and cats in the United States.

BACKGROUND: For decades, zinc sulfate centrifugal fecal flotation microscopy (ZCF) has been the mainstay technique for gastrointestinal (GI) parasite screening at veterinary clinics and laboratories. Elsewhere, PCR has replaced microscopy because of generally increased sensitivity and detection capabilities; however, until recently it has been unavailable commercially. Therefore, the primary aim of this study was to compare the performance of real-time PCR (qPCR) and ZCF for fecal parasite screening. Secondary aims included further characterization of markers for hookworm treatment resistance and Giardia spp. assemblages with zoonotic potential and qPCR optimization. METHODS: A convenience sampling of 931 canine/feline fecal samples submitted to a veterinary reference laboratory for routine ZCF from the Northeast US (11/2022) was subsequently evaluated by a broad qPCR panel following retention release. Detection frequency and agreement (kappa statistics) were evaluated between ZCF and qPCR for seven GI parasites [hookworm/(Ancylostoma spp.), roundworm/(Toxocara spp.), whipworm/(Trichuris spp.), Giardia duodenalis, Cystoisospora spp., Toxoplasma gondii, and Tritrichomonas blagburni] and detections per sample. Total detection frequencies were compared using a paired t-test; positive sample and co-infection frequencies were compared using Pearson's chi-squared test (p ≤ 0.05 significant) and qPCR frequency for hookworm benzimidazole (BZ) resistance (F167Y) and zoonotic Giardia spp. assemblage markers calculated. Confirmatory testing, characterization, and qPCR optimization were carried out with Sanger sequencing. RESULTS: qPCR detected a significantly higher overall parasite frequency (n = 679) compared to ZCF (n = 437) [p = < 0.0001, t = 14.38, degrees-of-freedom (df) = 930] and 2.6 × the co-infections [qPCR (n = 172) vs. ZCF (n = 66)], which was also significant (p = < 0.0001, X2 = 279.49; df = 1). While overall agreement of parasite detection was substantial [kappa = 0.74; (0.69-0.78], ZCF-undetected parasites reduced agreement for individual and co-infected samples. qPCR detected markers for Ancylostoma caninum BZ resistance (n = 5, 16.1%) and Giardia with zoonotic potential (n = 22, 9.1%) as well as two parasites undetected by ZCF (T. gondii/T. blagburni). Sanger sequencing detected novel roundworm species, and qPCR optimization provided detection beyond ZCF. CONCLUSIONS: These results demonstrate the statistically significant detection frequency advantage offered by qPCR compared to routine ZCF for both single and co-infections. While overall agreement was excellent, this rapid, commercially available qPCR panel offers benefits beyond ZCF with detection of markers for Giardia assemblages with zoonotic potential and hookworm (A. caninum) BZ resistance.

Minocycline and Diacetyl Minocycline Eye Drops Reduce Ocular Neovascularization in Mice.

Purpose: To evaluate the efficacy of minocycline and a novel, modified minocycline analogue that lacks antimicrobial action, diacetyl minocycline (DAM), on choroidal neovascularization (CNV) in mice of both sexes. Methods: CNV was induced via laser injury in female and male C57BL/6J mice. Minocycline, DAM, or saline was administered via topical eye drops twice a day for 2 weeks starting the day after laser injury. CNV volume was measured using immunohistochemistry labeling and confocal microscopy. Results: Minocycline reduced lesion volume by 79% (P ≤ 0.0004) in female and male mice. DAM reduced lesion volume by 73% (P ≤ 0.001) in female and male mice. There was no significant difference in lesion volume between minocycline and DAM treatment groups or between female and male mice. Conclusions: Both minocycline and DAM eye drops significantly reduced laser-induced CNV lesion volume in female and male mice. While oral tetracyclines have been shown to mitigate pathologic neovascularization in both preclinical studies and clinical trials, the present data are the first to suggest that tetracycline derivatives may be effective to reduce pathologic CNV when administered via topical eye drops. However, the action is unrelated to antimicrobial action. Targeted delivery of these medications via eye drops may reduce the potential for systemic side effects. Translational Relevance: Topical administration of minocycline and/or DAM via eye drops may represent a novel therapeutic strategy for disorders involving pathologic CNV.

Functional genomic analysis of adult and pediatric brain tumor isolates.

Background: Adult and pediatric tumors display stark differences in their mutation spectra and chromosome alterations. Here, we attempted to identify common and unique gene dependencies and their associated biomarkers among adult and pediatric tumor isolates using functional genetic lethal screens and computational modeling. Methods: We performed CRISRP-Cas9 lethality screens in two adult glioblastoma (GBM) tumor isolates and five pediatric brain tumor isolates representing atypical teratoid rhabdoid tumors (ATRT), diffuse intrinsic pontine glioma, GBM, and medulloblastoma. We then integrated the screen results with machine learning-based gene-dependency models generated from data from >900 cancer cell lines. Results: We found that >50% of candidate dependencies of 280 identified were shared between adult GBM tumors and individual pediatric tumor isolates. 68% of screen hits were found as nodes in our network models, along with shared and tumor-specific predictors of gene dependencies. We investigated network predictors associated with ADAR, EFR3A, FGFR1 (pediatric-specific), and SMARCC2 (ATRT-specific) gene dependency among our tumor isolates. Conclusions: The results suggest that, despite harboring disparate genomic signatures, adult and pediatric tumor isolates share a preponderance of genetic dependences. Further, combining data from primary brain tumor lethality screens with large cancer cell line datasets produced valuable insights into biomarkers of gene dependency, even for rare cancers. Importance of the Study: Our results demonstrate that large cancer cell lines data sets can be computationally mined to identify known and novel gene dependency relationships in adult and pediatric human brain tumor isolates. Gene dependency networks and lethality screen results represent a key resource for neuro-oncology and cancer research communities. We also highlight some of the challenges and limitations of this approach.

An in vitro Study of Betadine's Ability to Eliminate Live Bacteria on the Eye: Should It Be Used for Protection against Endophthalmitis?

BACKGROUND: Povidone-iodide (Betadine) is an antiseptic that is applied topically and has many uses in the medical community, such as in wound care and pre- and post-operative surgical procedures. This study was done to measure the effectiveness of Betadine solutions in inhibiting the growth of Gram-negative and Gram-positive bacteria. METHODS: The ability of 2.5 and 10% Betadine solutions to inhibit bacterial growth was measured against Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, and Acinetobacter baumannii. We grew the bacteria independently and together to simulate a hospital environment. RESULTS: All the bacteria showed zones of inhibition. However, discs were also tested for live bacteria using the colony-forming unit assay. Complete killing was only seen for S. aureus with the 10% Betadine solution. All other bacteria showed growth on the disc. CONCLUSIONS: This study showed several things. First, the zone of inhibition assay does not give an accurate assessment of antimicrobial properties when used alone and should be followed by a colony-forming unit assay. Second, 2.5% and 5% Betadine do not have effective antimicrobial properties against any of the bacteria tested, and 10% Betadine is only effective against S. aureus and not effective against the other bacteria tested.

Effects of the COVID-19 pandemic on dentists' workforce confidence and workflow.

BACKGROUND: The COVID-19 pandemic has affected the US economy and workforce, including marked effects on small businesses. Researchers have evaluated workers' views of financial confidence and advancement, but there has been limited focus on the dental industry. METHODS: To extend investigations to dentistry, the authors used published scales and pretested questions to determine workforce confidence and workflow changes among dentists. Data were evaluated using descriptive and bivariate statistics. In the wake of the pandemic, surveys were distributed to the memberships of the American Dental Association and American Association of Orthodontists (n = 656). RESULTS: Dentists' top concern was increased cost of providing treatment (57.4%; 95% CI, 53.5% to 61.3%), associated with widely adopted workflow changes including reduced patient volumes (66.0%; 95% CI, 62.4% to 69.6%) and increased safety protocols and equipment (health screening: 75.5%; 95% CI, 72.2% to 78.8%; KN/N95 respirators: 76.7%; 95% CI, 73.5% to 80.0%). However, most respondents did not expect their personal or practice finances to be negatively affected after the pandemic, as only 18.5% (95% CI, 15.4% to 21.7%) predicted their practice's gross revenue would decrease. CONCLUSIONS: Dentists were optimistic in the wake of vaccinations and lifting restrictions. Most expected their finances and practice performance to remain the same or grow in the short term and expected long-term improvements postpandemic. PRACTICAL IMPLICATIONS: Results suggest that despite shutdowns and workflow changes, dentists have rebounded financially and anticipate future growth.

Endothelial OCT4 is atheroprotective by preventing metabolic and phenotypic dysfunction.

AIMS: Until recently, the pluripotency factor Octamer (ATGCAAAT)-binding transcriptional factor 4 (OCT4) was believed to be dispensable in adult somatic cells. However, our recent studies provided clear evidence that OCT4 has a critical atheroprotective role in smooth muscle cells. Here, we asked if OCT4 might play a functional role in regulating endothelial cell (EC) phenotypic modulations in atherosclerosis. METHODS AND RESULTS: Specifically, we show that EC-specific Oct4 knockout resulted in increased lipid, LGALS3+ cell accumulation, and altered plaque characteristics consistent with decreased plaque stability. A combination of single-cell RNA sequencing and EC-lineage-tracing studies revealed increased EC activation, endothelial-to-mesenchymal transitions, plaque neovascularization, and mitochondrial dysfunction in the absence of OCT4. Furthermore, we show that the adenosine triphosphate (ATP) transporter, ATP-binding cassette (ABC) transporter G2 (ABCG2), is a direct target of OCT4 in EC and establish for the first time that the OCT4/ABCG2 axis maintains EC metabolic homeostasis by regulating intracellular heme accumulation and related reactive oxygen species production, which, in turn, contributes to atherogenesis. CONCLUSIONS: These results provide the first direct evidence that OCT4 has a protective metabolic function in EC and identifies vascular OCT4 and its signalling axis as a potential target for novel therapeutics.

SerpinB3 drives cancer stem cell survival in glioblastoma.

Despite therapeutic interventions for glioblastoma (GBM), cancer stem cells (CSCs) drive recurrence. The precise mechanisms underlying CSC resistance, namely inhibition of cell death, are unclear. We built on previous observations that the high cell surface expression of junctional adhesion molecule-A drives CSC maintenance and identified downstream signaling networks, including the cysteine protease inhibitor SerpinB3. Using genetic depletion approaches, we found that SerpinB3 is necessary for CSC maintenance, survival, and tumor growth, as well as CSC pathway activation. Knockdown of SerpinB3 also increased apoptosis and susceptibility to radiation therapy. SerpinB3 was essential to buffer cathepsin L-mediated cell death, which was enhanced with radiation. Finally, we found that SerpinB3 knockdown increased the efficacy of radiation in pre-clinical models. Taken together, our findings identify a GBM CSC-specific survival mechanism involving a cysteine protease inhibitor, SerpinB3, and provide a potential target to improve the efficacy of GBM therapies against therapeutically resistant CSCs.

VRK1 Is a Synthetic-Lethal Target in VRK2-Deficient Glioblastoma.

Synthetic lethality is a genetic interaction that results in cell death when two genetic deficiencies co-occur but not when either deficiency occurs alone, which can be co-opted for cancer therapeutics. Pairs of paralog genes are among the most straightforward potential synthetic-lethal interactions by virtue of their redundant functions. Here, we demonstrate a paralog-based synthetic lethality by targeting vaccinia-related kinase 1 (VRK1) in glioblastoma (GBM) deficient of VRK2, which is silenced by promoter methylation in approximately two thirds of GBM. Genetic knockdown of VRK1 in VRK2-null or VRK2-methylated cells resulted in decreased activity of the downstream substrate barrier to autointegration factor (BAF), a regulator of post-mitotic nuclear envelope formation. Reduced BAF activity following VRK1 knockdown caused nuclear lobulation, blebbing, and micronucleation, which subsequently resulted in G2-M arrest and DNA damage. The VRK1-VRK2 synthetic-lethal interaction was dependent on VRK1 kinase activity and was rescued by ectopic expression of VRK2. In VRK2-methylated GBM cell line-derived xenograft and patient-derived xenograft models, knockdown of VRK1 led to robust tumor growth inhibition. These results indicate that inhibiting VRK1 kinase activity could be a viable therapeutic strategy in VRK2-methylated GBM. SIGNIFICANCE: A paralog synthetic-lethal interaction between VRK1 and VRK2 sensitizes VRK2-methylated glioblastoma to perturbation of VRK1 kinase activity, supporting VRK1 as a drug discovery target in this disease.