Increasing intracellular dNTP levels improves prime editing efficiency.

In primary cell types, intracellular deoxynucleotide triphosphate (dNTP) levels are tightly regulated in a cell cycle-dependent manner. We report that prime editing efficiency is increased by mutations that improve the enzymatic properties of Moloney murine leukemia virus reverse transcriptase and treatments that increase intracellular dNTP levels. In combination, these modifications produce substantial increases in precise editing rates.

Next generation APOBEC3 inhibitors: Optimally designed for potency and nuclease stability.

APOBEC3 (or A3) enzymes have emerged as potential therapeutic targets due to their role in introducing heterogeneity in viruses and cancer, often leading to drug resistance. Inhibiting these enzymes has remained elusive as initial phosphodiester (PO) linked DNA based inhibitors lack stability and potency. We have enhanced both potency and nuclease stability, of 2'-deoxy-zebularine (dZ), substrate-based oligonucleotide inhibitors for two critical A3's: A3A and A3G. While replacing the phosphate backbone with phosphorothioate (PS) linkages increased nuclease stability, fully PS-modified inhibitors lost potency (1.4-3.7 fold) due to the structural constraints of the active site. For both enzymes, mixed PO/PS backbones enhanced potency (2.3-9.2 fold), while also vastly improving nuclease resistance. We also strategically introduced 2'-fluoro sugar modifications, creating the first nanomolar inhibitor of A3G-CTD2. With hairpin-structured inhibitors containing optimized PS patterns and LNA sugar modifications, we characterize the first single-digit nanomolar inhibitor targeting A3A. These extremely potent A3A inhibitors, were highly resistant to nuclease degradation in serum stability assays. Overall, our optimally designed A3 oligonucleotide inhibitors show improved potency and stability, compared to previous attempts to inhibit these critical enzymes, opening the door to realize the therapeutic potential of A3 inhibition.

A Comparative Study of SonoVue and Sonazoid for Contrast-Enhanced Ultrasound CT/MRI Fusion Guidance During Radiofrequency Ablation of Poorly Visualized Hepatic Malignancies: A Prospective Intra-Individual Analysis.

PURPOSE: This study aimed to evaluate the effectiveness of two contrast agents, SonoVue (SV) and Sonazoid (SZ), by comparing them intra-individually in contrast-enhanced ultrasound (CEUS)-CT/MRI fusion imaging (FI) to improve the visibility of inconspicuous liver malignancies on B-mode sonography for guiding percutaneous radiofrequency ablation (RFA). Additionally, the radiologists' preference between SonoVue- CT/MRI FI (SV-FI) and Sonazoid-CT/MRI FI (SZ-FI) was determined. METHODS: This prospective study enrolled 23 patients with inconspicuous hepatic malignancies (≤ 3 cm) on B-mode US who underwent both SV-FI and SZ-FI for RFA guidance. The patients underwent real-time CEUS FI with CT/MRI on the same day, utilizing both SV and SZ with at least 15-min intervals. Tumor visibility and radiologists' preferences were assessed and graded using a 4-point scale during the dynamic phases of both SV-FI and SZ-FI and the Kupffer phase of SZ-FI. RESULTS: The tumor visibility scores obtained from CEUS-CT/MRI FI were significantly better than those obtained from US-FI. Indeed, SV-FI and SZ-FI demonstrated comparable visibility scores when corresponding phases were compared (p > 0.05). However, the Kupffer phase images of SZ-FI displayed superior visibility scores (3.70 ± 0.56 vs. 2.96 ± 0.88; p = 0.002) than the late vascular phase images of SV-FI. The radiologists favored SZ-FI in many cases, exhibiting moderate inter-observer agreement (Kappa value = 0.587; 95% CI, 0.403-0.772). CONCLUSION: Although CEUS-CT/MRI FI with either SV or SZ substantially improved the visibility of inconspicuous tumors on US-CT/MRI FI, radiologists preferred SZ to SV to guide the RFA procedure.

Conversion of acidified lignin containing sulfur discharged from a biorefinery process into neutralized biochar: Characterization and metal adsorption.

The objectives of this study were to produce biochars using sulfur-rich acidified lignin discharged from a biorefinery process and to evaluate their physicochemical properties and Pb adsorption capacity. As the pyrolysis temperature increased, the lignin acidified by the desulfurization process was converted to neutralized biochar (LBC), which exhibited high carbon content and stability. The carbon content of biochar manufactured at a pyrolysis temperature of 600 °C or higher was over 90 % and showed no significant difference, and their surface structures were found to be different, as revealed through XRD and FTIR analyses. The adsorption capacity of Pb by LBC increased with increasing pyrolysis temperature, and their adsorption capacity was well described by the pseudo-second-order model and the Langmuir isotherm adsorption model. In particular, the internal diffusion effect on the adsorption capacity of Pb was greater for LBC900 than for LBC600. In complex heavy metal solutions, LBC selectively exhibited high affinity for Pb, while the adsorption capacity of other metals was significantly reduced. The adsorption mechanism of Pb by LBC was verified through various analytical methods, and these results demonstrated that the adsorption of Pb by LBC was influenced by functional groups existing on the surface and inside of LBC and by some cation exchange.

Strongylocentrotus intermedius Extract Suppresses Adiposity by Inhibiting Adipogenesis and Promoting Adipocyte Browning via AMPK Activation in 3T3-L1 Cells.

The current study aimed to determine whether Strongylocentrotus intermedius (S. intermedius) extract (SIE) exerts anti-obesity potentials employing 3T3-L1 cells as in vitro model. Herein we reported that treatment of SIE for 6 days reduced lipid accretion and triglyceride content whereas it increased the release of free glycerol. The inhibited lipid accumulation and induced lipolysis were evidenced by the downregulation of lipogenesis proteins, such as fatty acid synthase and lipoprotein lipase, and the upregulation of hormone-sensitive lipase expression. Furthermore, the downregulation of adipogenic transcription factors, including peroxisome proliferator-activated receptor gamma, CCAAT/enhancer-binding protein α, and sterol regulatory element-binding protein 1, highlights that reduced lipid accumulation is supported by lowering adipocyte differentiation. Additionally, treatment activates brown adipocyte phenotype in 3T3-L1 cells by inducing expression of brown adipose tissue-specific proteins, such as uncoupling protein 1 and peroxisome proliferator-activated receptor-γ coactivator 1α. Moreover, SIE induced the phosphorylation of AMP-activated protein kinase (AMPK). The pharmacological approach using AMPK inhibitor revealed that the restraining effect of SIE on adipogenesis and promotion of adipocyte browning were blocked. In GC-MS analysis, SIE was mainly composed of cholest-5-en-3-ol (36.71%) along with saturated and unsaturated fatty acids which have favorable anti-obesity potentials. These results reveal that SIE has the possibility as a lipid-lowering agent for the intervention of obesity.

Induction of Cell Death by Bifidobacterium infantis DS1685 in Colorectal and Breast Cancers via SMAD4/TGF-Beta Activation.

Therapeutic advancements in treatments for cancer, a leading cause of mortality worldwide, have lagged behind the increasing incidence of this disease. There is a growing interest in multifaceted approaches for cancer treatment, such as chemotherapy, targeted therapy, and immunotherapy, but due to their low efficacy and severe side effects, there is a need for the development of new cancer therapies. Recently, the human microbiome, which is comprised of various microorganisms, has emerged as an important research field due to its potential impact on cancer treatment. Among these microorganisms, Bifidobacterium infantis has been shown to significantly improve the efficacy of various anticancer drugs. However, research on the role of B. infantis in cancer treatment remains insufficient. Thus, in this study, we explored the anticancer effect of treatment with B. infantis DS1685 supernatant (BI sup) in colorectal and breast cancer cell lines. Treatment with BI sup induced SMAD4 expression to suppress cell growth in colon and breast cancer cells. Furthermore, a decrease in tumor cohesion was observed through the disruption of the regulation of EMT-related genes by BI sup in 3D spheroid models. Based on these findings, we anticipate that BI sup could play an adjunctive role in cancer therapy, and future cotreatment of BI sup with various anticancer drugs may lead to synergistic effects in cancer treatment.

Non-invasive CT radiomic biomarkers predict microsatellite stability status in colorectal cancer: a multicenter validation study.

BACKGROUND: Microsatellite instability (MSI) status is a strong predictor of response to immunotherapy of colorectal cancer. Radiogenomic approaches promise the ability to gain insight into the underlying tumor biology using non-invasive routine clinical images. This study investigates the association between tumor morphology and the status of MSI versus microsatellite stability (MSS), validating a novel radiomic signature on an external multicenter cohort. METHODS: Preoperative computed tomography scans with matched MSI status were retrospectively collected for 243 colorectal cancer patients from three hospitals: Seoul National University Hospital (SNUH); Netherlands Cancer Institute (NKI); and Fondazione IRCCS Istituto Nazionale dei Tumori, Milan Italy (INT). Radiologists delineated primary tumors in each scan, from which radiomic features were extracted. Machine learning models trained on SNUH data to identify MSI tumors underwent external validation using NKI and INT images. Performances were compared in terms of area under the receiving operating curve (AUROC). RESULTS: We identified a radiomic signature comprising seven radiomic features that were predictive of tumors with MSS or MSI (AUROC 0.69, 95% confidence interval [CI] 0.54-0.84, p = 0.018). Integrating radiomic and clinical data into an algorithm improved predictive performance to an AUROC of 0.78 (95% CI 0.60-0.91, p = 0.002) and enhanced the reliability of the predictions. CONCLUSION: Differences in the radiomic morphological phenotype between tumors MSS or MSI could be detected using radiogenomic approaches. Future research involving large-scale multicenter prospective studies that combine various diagnostic data is necessary to refine and validate more robust, potentially tumor-agnostic MSI radiogenomic models. RELEVANCE STATEMENT: Noninvasive radiomic signatures derived from computed tomography scans can predict MSI in colorectal cancer, potentially augmenting traditional biopsy-based methods and enhancing personalized treatment strategies. KEY POINTS: Noninvasive CT-based radiomics predicted MSI in colorectal cancer, enhancing stratification. A seven-feature radiomic signature differentiated tumors with MSI from those with MSS in multicenter cohorts. Integrating radiomic and clinical data improved the algorithm's predictive performance.

Incremental high power radiofrequency ablation with multi-electrodes for small hepatocellular carcinoma: a prospective study.

Radiofrequency ablation (RFA) offers a minimally invasive treatment for small hepatocellular carcinoma (HCC), but it faces challenges such as high local recurrence rates. This prospective study, conducted from January 2020 to July 2022, evaluated a novel approach using a three-channel, dual radiofrequency (RF) generator with separable clustered electrodes to improve RFA's efficacy and safety. The study employed a high-power, gradual, stepwise RFA method on HCCs (≤ 4 cm), utilizing real-time ultrasound-computed tomography (CT)/magnetic resonance imaging (MRI) fusion imaging. Involving 110 participants with 116 HCCs, the study reported no major complications. Local tumor progression (LTP) and intrahepatic remote recurrence (IRR) rates were low, with promising cumulative incidences at 1, 2, and 3 years for LTP (0.9%, 3.6%, 7.0%) and IRR (13.9%, 20.5%, 31.4%). Recurrence-free survival (RFS) rates were similarly encouraging: LTP (99.1%, 96.4%, 93.0%) and IRR (86.1%, 79.5%, 68.6%). This innovative gradual, incremental high-power RFA technique, featuring a dual switching monopolar mode and three electrodes, represents an effective and safer management option for small HCCs. TRIAL REGISTRATION: clinicaltrial.gov identifier: NCT05397860, first registered on 26/05/2022.

Deep learning-based 3D quantitative total tumor burden predicts early recurrence of BCLC A and B HCC after resection.

OBJECTIVES: This study aimed to evaluate the potential of deep learning (DL)-assisted automated three-dimensional quantitative tumor burden at MRI to predict postoperative early recurrence (ER) of hepatocellular carcinoma (HCC). MATERIALS AND METHODS: This was a single-center retrospective study enrolling patients who underwent resection for BCLC A and B HCC and preoperative contrast-enhanced MRI. Quantitative total tumor volume (cm3) and total tumor burden (TTB, %) were obtained using a DL automated segmentation tool. Radiologists' visual assessment was used to ensure the quality control of automated segmentation. The prognostic value of clinicopathological variables and tumor burden-related parameters for ER was determined by Cox regression analyses. RESULTS: A total of 592 patients were included, with 525 and 67 patients assigned to BCLC A and B, respectively (2-year ER rate: 30.0% vs. 45.3%; hazard ratio (HR) = 1.8; p = 0.007). TTB was the most important predictor of ER (HR = 2.2; p < 0.001). Using 6.84% as the threshold of TTB, two ER risk strata were obtained in overall (p < 0.001), BCLC A (p < 0.001), and BCLC B (p = 0.027) patients, respectively. The BCLC B low-TTB patients had a similar risk for ER to BCLC A patients and thus were reassigned to a BCLC An stage; whilst the BCLC B high-TTB patients remained in a BCLC Bn stage. The 2-year ER rate was 30.5% for BCLC An patients vs. 58.1% for BCLC Bn patients (HR = 2.8; p < 0.001). CONCLUSIONS: TTB determined by DL-based automated segmentation at MRI was a predictive biomarker for postoperative ER and facilitated refined subcategorization of patients within BCLC stages A and B. CLINICAL RELEVANCE STATEMENT: Total tumor burden derived by deep learning-based automated segmentation at MRI may serve as an imaging biomarker for predicting early recurrence, thereby improving subclassification of Barcelona Clinic Liver Cancer A and B hepatocellular carcinoma patients after hepatectomy. KEY POINTS: Total tumor burden (TTB) is important for Barcelona Clinic Liver Cancer (BCLC) staging, but is heterogenous. TTB derived by deep learning-based automated segmentation was predictive of postoperative early recurrence. Incorporating TTB into the BCLC algorithm resulted in successful subcategorization of BCLC A and B patients.

Enhanced Deposition Selectivity of High-k Dielectrics by Vapor Dosing and Selective Removal of Phosphonic Acid Inhibitors.

Area-selective atomic layer deposition (AS-ALD), which provides a bottom-up nanofabrication method with atomic-scale precision, has attracted a great deal of attention as a means to alleviate the problems associated with conventional top-down patterning. In this study, we report a methodology for achieving selective deposition of high-k dielectrics by surface modification through vapor-phase functionalization of octadecylphosphonic acid (ODPA) inhibitor molecules accompanied by post-surface treatment. A comparative evaluation of deposition selectivity of ZrO2 thin films deposited with the O2 and O3 reactants was performed on SiO2, TiN, and W substrates, and we confirmed that high enough deposition selectivity over 10 nm can be achieved even after 200 cycles of ALD with the O2 reactant. Subsequently, the electrical properties of ZrO2 films deposited with O2 and O3 reactants were investigated with and without post-deposition treatment. We successfully demonstrated that high-quality ZrO2 thin films with high dielectric constants and stable antiferroelectric properties can be produced by subjecting the films to ozone, which can eliminate carbon impurities within the films. We believe that this work provides a new strategy to achieve highly selective deposition for AS-ALD of dielectric on dielectric (DoD) applications toward upcoming bottom-up nanofabrication.