Expanding Roles of the E2F-RB-p53 Pathway in Tumor Suppression.

The transcription factor E2F links the RB pathway to the p53 pathway upon loss of function of pRB, thereby playing a pivotal role in the suppression of tumorigenesis. E2F fulfills a major role in cell proliferation by controlling a variety of growth-associated genes. The activity of E2F is controlled by the tumor suppressor pRB, which binds to E2F and actively suppresses target gene expression, thereby restraining cell proliferation. Signaling pathways originating from growth stimulative and growth suppressive signals converge on pRB (the RB pathway) to regulate E2F activity. In most cancers, the function of pRB is compromised by oncogenic mutations, and E2F activity is enhanced, thereby facilitating cell proliferation to promote tumorigenesis. Upon such events, E2F activates the Arf tumor suppressor gene, leading to activation of the tumor suppressor p53 to protect cells from tumorigenesis. ARF inactivates MDM2, which facilitates degradation of p53 through proteasome by ubiquitination (the p53 pathway). P53 suppresses tumorigenesis by inducing cellular senescence or apoptosis. Hence, in almost all cancers, the p53 pathway is also disabled. Here we will introduce the canonical functions of the RB-E2F-p53 pathway first and then the non-classical functions of each component, which may be relevant to cancer biology.

HMG-CoA reductase degrader, SR-12813, counteracts statin-induced upregulation of HMG-CoA reductase and augments the anticancer effect of atorvastatin.

Statins are cholesterol-lowering drugs that have exhibited potential as cancer therapeutic agents. However, as some cancer cells are resistant to statins, broadening an anticancer spectrum of statins is desirable. The upregulated expression of the statin target enzyme, 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase (HMGCR), in statin-treated cancer cells is a well-known mechanism of statin resistance, which can be counteracted by the downregulation of HMGCR gene expression, or degradation of the HMGCR protein. However, the mechanism by which HMGCR degradation influences the anticancer effects of statins remain unreported. We tested the effect of the HMGCR degrader compound SR-12813 at a concentration that did not affect the growth of eight diverse tumor cell lines. Combined treatment with atorvastatin and a low concentration of SR-12813 led to lowering of increased HMGCR expression, and augmented the cytostatic effect of atorvastatin in both statin-resistant and -sensitive cancer cells compared with that of atorvastatin treatment alone. Dual-targeting of HMGCR using statins and SR-12813 (or similar compounds) could provide an improved anticancer therapeutic approach.

Expression of housekeeping genes varies depending on mevalonate pathway inhibition in cancer cells.

Statins have anticancer effects and may be used as anticancer agents via drug repositioning. In reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays, the internal reference gene must not be affected by any experimental conditions. As statins exert a wide range of effects on cells by inhibiting the mevalonate pathway, it is possible that statin treatment might alter the expression of housekeeping genes used as internal reference genes, thereby misleading the assessment of obtained gene expression data. Here, we evaluated the expression stability of internal reference genes in atorvastatin-treated cancer cell lines. We treated both statin-sensitive and statin-resistant cancer cell lines with atorvastatin at seven different concentrations and performed RT-qPCR on 15 housekeeping genes whose expression stability was then assessed using five different algorithms. In both statin-sensitive and statin-resistant cancer cell lines, atorvastatin affected the expression of certain internal reference genes in a dose-dependent and cancer cell line-dependent manner; therefore, caution should be exercised when comparing target gene expression between cells. Our findings emphasize the importance of the validation of internal reference genes in gene expression analyses in drug treatment-based cancer research.

Simultaneous qualitative and quantitative analyses of volatile components in Chinese honey of six botanical origins using headspace solid-phase microextraction and gas chromatography-mass spectrometry.

BACKGROUND: Honey aroma is one of its most important properties and it depends on the qualitative and quantitative composition of the volatile compounds. The volatile profile of honey could reveal its botanical origin to avoid a false characterization. Thus, it is of great significance to honey authentication. This study developed and validated a headspace solid-phase microextraction and gas chromatography-mass spectrometry (HS-SPME-GC-MS) method for simultaneous qualitative and quantitative analyses of 34 volatile components in honey. The developed method was applied to 86 honey samples from six different botanical origins, including linden honey, rape honey, jujube honey, vitex honey, lavender honey and acacia honey. RESULTS: The volatile fingerprints and quantitative results were simultaneously obtained by using the full scan and selected ion monitoring (SCAN+SIM) MS scanning mode. The limits of quantification (LOQs) and limits of detection (LODs) of 34 volatile compounds were in the ranges of 1-10 ng/g and 0.3-3 ng/g, respectively. And the spiked recoveries ranged between 70.6% and 126.2%, with the relative standard deviations (RSDs) not higher than 45.4%. A total of 98 volatile compounds were found with relative contents determined, and the 34 volatile compounds were determined with absolute concentrations. Based on the volatile fingerprints and the contents of volatile compounds, honey samples from six botanical origins were well classified by principal component analysis and orthogonal partial least-squares discrimination analysis. CONCLUSIONS: The HS-SPME-GC-MS method was successfully applied to achieve the volatile fingerprints of six types of honey and to quantitatively analyze 34 volatile compounds with satisfying sensitivity and accuracy. Chemometrics analysis showed significant correlations between honey types and volatiles. These results reveal the characteristics of volatile compounds in six types of unifloral honey and provide some supports for honey authentication. © 2023 Society of Chemical Industry.

Solvent-switchable regioselective 1,2- or 1,6-addition of quinones with boronic acids.

An efficient copper-catalyzed solvent-switchable regioselective 1,2- or 1,6-addition of quinones with boronic acids has been developed. This novel catalytic protocol for the synthesis of various quinols and 4-phenoxyphenols was enabled by a simple solvent swap between H2O and MeOH. It features mild reaction conditions, simple and easy operation, broad substrate scope and excellent regioselectivity. The gram-scale reactions as well as the further transformations of both addition products were also successfully investigated.

Lung microbiota and potential treatment of respiratory diseases.

The unique microbiome found in the lungs has been studied and shown to be associated with both pulmonary homeostasis and lung diseases. The lung microbiome has the potential to produce metabolites that modulate host-microbe interactions. Specifically, short-chain fatty acids (SCFAs) produced by certain strains of the lung microbiota have been shown to regulate immune function and maintain gut mucosal health. In response, this review described the distribution and composition of the microbiota in lung diseases and discussed the impact of the lung microbiota on health and lung disease. In addition, the review further elaborated on the mechanism of microbial metabolites in microbial-host interaction and their application in the treatment of lung diseases. A better understanding of the interaction between the microbiota, metabolites, and host will provide potential strategies for the development of novel methods for the treatment of pulmonary microbial induced lung diseases.

The TFDP1 gene coding for DP1, the heterodimeric partner of the transcription factor E2F, is a target of deregulated E2F.

The TFDP1 gene codes for the heterodimeric partner DP1 of the transcription factor E2F. E2F, principal target of the tumor suppressor pRB, plays central roles in cell proliferation by activating a group of growth-related genes. E2F also mediates tumor suppression by activating tumor suppressor genes such as ARF, an upstream activator of the tumor suppressor p53, when deregulated from pRB upon oncogenic changes. Among 8 E2F family members (E2F1∼E2F8), expression of activator E2Fs (E2F1∼E2F3a) is induced at the G1/S boundary of the cell cycle after growth stimulation by E2F itself. However, mechanisms regulating DP1 expression are not known. We show here that over-expression of E2F1 and forced inactivation of pRB, by adenovirus E1a, induced TFDP1 gene expression in human normal fibroblast HFFs, suggesting that the TFDP1 gene is a target of E2F. Serum stimulation of HFFs also induced TFDP1 gene expression, but with different kinetics from that of the CDC6 gene, a typical growth-related E2F target. Both over-expression of E2F1 and serum stimulation activated the TFDP1 promoter. We searched for E2F1-responsive regions by 5' and 3' deletion of the TFDP1 promoter and by introducing point mutations in putative E2F1-responsive elements. Promoter analysis identified several GC-rich elements, mutation of which reduced E2F1-responsiveness but not serum-responsiveness. ChIP assays showed that the GC-rich elements bound deregulated E2F1 but not physiological E2F1 induced by serum stimulation. These results suggest that the TFDP1 gene is a target of deregulated E2F. In addition, knockdown of DP1 expression by shRNA enhanced ARF gene expression, which is specifically induced by deregulated E2F activity, suggesting that activation of the TFDP1 gene by deregulated E2F may function as a failsafe feedback mechanism to suppress deregulated E2F and maintain normal cell growth in the event that DP1 expression is insufficient relative to that of its partner activator E2Fs. a maximum of 6 keywords: E2F, DP1, TFDP1 gene, pRB, gene expression.

Flexible fiber cholangioscope for detection of near-infrared fluorescence.

Video 1Flexible fiber cholangioscope for detection of near-infrared fluorescence.

Deregulated E2F Activity as a Cancer-Cell Specific Therapeutic Tool.

The transcription factor E2F, the principal target of the tumor suppressor pRB, plays crucial roles in cell proliferation and tumor suppression. In almost all cancers, pRB function is disabled, and E2F activity is enhanced. To specifically target cancer cells, trials have been undertaken to suppress enhanced E2F activity to restrain cell proliferation or selectively kill cancer cells, utilizing enhanced E2F activity. However, these approaches may also impact normal growing cells, since growth stimulation also inactivates pRB and enhances E2F activity. E2F activated upon the loss of pRB control (deregulated E2F) activates tumor suppressor genes, which are not activated by E2F induced by growth stimulation, inducing cellular senescence or apoptosis to protect cells from tumorigenesis. Deregulated E2F activity is tolerated in cancer cells due to inactivation of the ARF-p53 pathway, thus representing a feature unique to cancer cells. Deregulated E2F activity, which activates tumor suppressor genes, is distinct from enhanced E2F activity, which activates growth-related genes, in that deregulated E2F activity does not depend on the heterodimeric partner DP. Indeed, the ARF promoter, which is specifically activated by deregulated E2F, showed higher cancer-cell specific activity, compared to the E2F1 promoter, which is also activated by E2F induced by growth stimulation. Thus, deregulated E2F activity is an attractive potential therapeutic tool to specifically target cancer cells.

Alterations in the omics profiles in mevalonate pathway-inhibited cancer cells.

AIMS: Statins, cholesterol-lowering drugs, are potential therapeutic agents for inhibiting cancer proliferation. However, the mechanisms that mediate the effects of statins, the homeostatic responses of tumor cells to statin therapy, and the modes underlying the antitumor effects of statins remain unclear. MAIN METHODS: To uncover the effects of statins on cancer cells in vitro, we performed transcriptome and metabolome analyses on atorvastatin-treated statin-resistant and statin-sensitive lung cancer cells. KEY FINDINGS: The results of Gene Ontology terms and pathway enrichment analyses showed that after 24 h of atorvastatin treatment, the expression of cell cycle- and DNA replication-related genes was significantly decreased in the statin-sensitive cancer cells. The results of metabolome analysis showed that the components of polyamine metabolism and purine metabolism, glycolysis, and pentose phosphate pathway were decreased in the statin-sensitive cancer cells. SIGNIFICANCE: Differences in cellular properties between statin-sensitive and statin-resistant cancer cells revealed additional candidates for therapeutic targets in statin-treated cancer cells and suggested that inhibiting these metabolic pathways could improve efficacy. In conclusion, combining statins with inhibitors of polyamine metabolism (cell proliferation and protein translation), purine metabolism (DNA synthesis), glycolytic system (energy production), and pentose phosphate pathway (antioxidant stress) might enhance the anticancer effects of statins.

Cost-Efficient Video Synthesis and Evaluation for Development of Virtual 3D Endoscopy.

OBJECTIVE: 3D reconstruction of the shape and texture of hollow organs captured by endoscopy is important for the diagnosis and surveillance of early and recurrent cancers. Better evaluation of 3D reconstruction pipelines developed for such applications requires easy access to extensive datasets and associated ground truths, cost-efficient and scalable simulations of a range of possible clinical scenarios, and more reliable and insightful metrics to assess performance. METHODS: We present a computer-aided simulation platform for cost-effective synthesis of monocular endoscope videos and corresponding ground truths that mimic a range of potential settings and situations one might encounter during acquisition of clinical endoscopy videos. Using cystoscopy of the bladder as model case, we generated an extensive dataset comprising several synthesized videos of a bladder phantom. We then introduce a novel evaluation procedure to reliably assess an individual 3D reconstruction pipeline or to compare different pipelines. RESULTS: To illustrate the use of the proposed platform and evaluation procedure, we use the aforementioned dataset and ground truths to evaluate a proprietary 3D reconstruction pipeline (CYSTO3D) for bladder cystoscopy videos and compared it with a general-purpose 3D reconstruction pipeline (COLMAP). The evaluation results provide insight into the suggested clinical acquisition protocol and several potential areas for refinement of the pipeline to improve future performance. CONCLUSION: Our work proposes an endoscope video synthesis and reconstruction evaluation toolset and presents experimental results that illustrate usage of the toolset to efficiently assess performance and reveal possible problems of any given 3D reconstruction pipeline, to compare different pipelines, and to provide technically or clinically actionable insights.

Type B thymoma in a patient with HIV infection: A case report with a review of HIV and thymoma coexistence.

HIV infection predisposes people to cancer, including AIDS-defining cancers, such as Kaposi sarcoma, and a broad range of non-AIDS-defining cancers. Here we report a case with rare coexistence of HIV and thymoma, and summarize all the comorbid cases that currently exist. We found that in all the cases reported, thymoma occurred when CD4+ counts were within a normal range, but the immune response in peripheral T-cell repertoire remains unknown. In our case, an overview of the immune system under this complicated situation is given for the first time by showing the lymphocyte subpopulations in the blood and the immune cell distribution of the thymoma. This case expands the scope of non-AIDS-defining cancers, and provides insight into the influence of the immune system under two immunocompromising conditions, HIV infection and thymoma.

Heterologous prime-boost vaccination targeting MAGE-type antigens promotes tumor T-cell infiltration and improves checkpoint blockade therapy.

BACKGROUND: The clinical benefit of immune checkpoint blockade (ICB) therapy is often limited by the lack of pre-existing CD8+ T cells infiltrating the tumor. In principle, CD8+ T-cell infiltration could be promoted by therapeutic vaccination. However, this remains challenging given the paucity of vaccine platforms able to induce the strong cytotoxic CD8+ T-cell response required to reject tumors. A therapeutic cancer vaccine that induces a robust cytotoxic CD8+ T-cell response against shared tumor antigens and can be combined with ICB could improve the outcome of cancer immunotherapy. METHODS: Here, we developed a heterologous prime-boost vaccine based on a chimpanzee adenovirus (ChAdOx1) and a modified vaccinia Ankara (MVA) encoding MAGE-type antigens, which are tumor-specific shared antigens expressed in different tumor types. The mouse MAGE-type antigen P1A was used as a surrogate to study the efficacy of the vaccine in combination with ICB in murine tumor models expressing the P1A antigen. To characterize the vaccine-induced immune response, we performed flow cytometry and transcriptomic analyses. RESULTS: The ChAdOx1/MVA vaccine displayed strong immunogenicity with potent induction of CD8+ T cells. When combined with anti-Programmed Cell Death Protein 1 (PD-1), the vaccine induced superior tumor clearance and survival in murine tumor models expressing P1A compared with anti-PD-1 alone. Remarkably, ChAdOx1/MVA P1A vaccination promoted CD8+ T-cell infiltration in the tumors, and drove inflammation in the tumor microenvironment, turning 'cold' tumors into 'hot' tumors. Single-cell transcriptomic analysis of the P1A-specific CD8+ T cells revealed an expanded population of stem-like T cells in the spleen after the combination treatment as compared with vaccine alone, and a reduced PD-1 expression in the tumor CD8+ T cells. CONCLUSIONS: These findings highlight the synergistic potency of ChAdOx1/MVA MAGE vaccines combined with anti-PD-1 for cancer therapy, and establish the foundation for clinical translation of this approach. A clinical trial of ChadOx1/MVA MAGE-A3/NY-ESO-1 combined with anti-PD-1 will commence shortly.

Copper-Catalyzed Three-Component Reactions of α-Ketoaldehyde, 1,3-Dicarbonyl Compound, and Organic Boronic Acid in Water: A Route to 1,4-Diketones.

A novel three-component reaction of α-ketoaldehydes, 1,3-dicarbonyl compounds, and organic boronic acids catalyzed by CuO in water has been developed to give a wide range of products containing 1,3/1,4-diketones. The method has some advantages such as the use of readily available starting materials, wide substrate scopes, excellent yields, gram-scale synthesis, and mild reaction conditions.

Optimizing genome editing strategy by primer-extension-mediated sequencing.

Efficient and precise genome editing is essential for clinical applications and generating animal models, which requires engineered nucleases with high editing ability while low off-target activity. Here we present a high-throughput sequencing method, primer-extension-mediated sequencing (PEM-seq), to comprehensively assess both editing ability and specificity of engineered nucleases. We showed CRISPR/Cas9-generated breaks could lead to chromosomal translocations and large deletions by PEM-seq. We also found that Cas9 nickase possessed lower off-target activity while with some loss of target cleavage ability. However, high-fidelity Cas9 variants, including both eCas9 and the new FeCas9, could significantly reduce the Cas9 off-target activity with no obvious editing retardation. Moreover, we found AcrIIA4 inhibitor could greatly reduce the activities of Cas9, but off-target loci were not so effectively suppressed as the on-target sites. Therefore, PEM-seq fully evaluating engineered nucleases could help choose better genome editing strategy at given loci than other methods detecting only off-target activity.

Near-Infrared Imaging of Artificial Enamel Caries Lesions with a Scanning Fiber Endoscope.

Several studies have shown that near-infrared imaging has great potential for the detection of dental caries lesions. A miniature scanning fiber endoscope (SFE) operating at near-infrared (NIR) wavelengths was developed and used in this study to test whether the device could be used to discriminate demineralized enamel from sound enamel. Varying depths of artificial enamel caries lesions were prepared on 20 bovine blocks with smooth enamel surfaces. Samples were imaged with a SFE operating in the reflectance mode at 1310-nm and 1460-nm in both wet and dry conditions. The measurements acquired by the SFE operating at 1460-nm show significant difference between the sound and the demineralized enamel. There was a moderate positive correlation between the SFE measurements and micro-CT measurements, and the NIR SFE was able to detect the presence of demineralization with high sensitivity (0.96) and specificity (0.85). This study demonstrates that the NIR SFE can be used to detect early demineralization from sound enamel. In addition, the NIR SFE can differentiate varying severities of demineralization. With its very small form factor and maneuverability, the NIR SFE should allow clinicians to easily image teeth from multiple viewing angles in real-time.

Near-infrared multispectral endoscopic imaging of deep artificial interproximal lesions in extracted teeth.

BACKGROUND AND OBJECTIVE: A safer alternative method to radiographic imaging is needed. We present a multispectral near-infrared scanning fiber endoscope (nirSFE) for dental imaging which is designed to be the smallest imaging probe with near-infrared (NIR) imaging (1200-2000 nm). MATERIALS AND METHODS: The prototype nirSFE is designed for wide-field forward viewing of scanned laser illumination at 1310, 1460, or 1550 nm. Artificial lesions with varying sizes and locations were prepared on proximal surfaces of extracted human teeth to examine capability and limitation of this new dental imaging modality. Nineteen artificial interproximal lesions and several natural occlusal lesions on extracted teeth were imaged with nirSFE, OCT, and microCT. RESULTS: Our nirSFE system has a flexible shaft as well as a probe tip with diameter of 1.6 mm and a rigid length of 9 mm. The small form factor and multispectral NIR imaging capability enables multiple viewing angles and reliable detection of lesions that can extend into the dentin. Among nineteen artificial interproximal lesions, the nirSFE reflectance imaging operating at 1460-nm and OCT operating at 1310-nm scanned illumination exhibited high sensitivity for interproximal lesions that were closer to occlusal surface. Diagnosis from a non-blinded trained user by looking at real-time occlusal-side nirSFE videos indicate true positive rate of 78.9%. There were no false positives. CONCLUSIONS: This study demonstrates that nirSFE may be used for detecting occlusal lesions and interproximal lesions located less than 4 mm under the occlusal surface. Major advantages of this imaging system include multiple viewing angles due to flexibility and small form factor, as well as the ability to capture real-time video. The multispectral nirSFE has the potential to be employed as a low-cost dental camera for detecting dental lesions without exposure to ionizing radiation. Lasers Surg. Med. 51:459-465, 2019. © 2019 Wiley Periodicals, Inc.

Towards AR-assisted visualisation and guidance for imaging of dental decay.

Untreated dental decay is the most prevalent dental problem in the world, affecting up to 2.4 billion people and leading to a significant economic and social burden. Early detection can greatly mitigate irreversible effects of dental decay, avoiding the need for expensive restorative treatment that forever disrupts the enamel protective layer of teeth. However, two key challenges exist that make early decay management difficult: unreliable detection and lack of quantitative monitoring during treatment. New optically based imaging through the enamel provides the dentist a safe means to detect, locate, and monitor the healing process. This work explores the use of an augmented reality (AR) headset to improve the workflow of early decay therapy and monitoring. The proposed workflow includes two novel AR-enabled features: (i) in situ visualisation of pre-operative optically based dental images and (ii) augmented guidance for repetitive imaging during therapy monitoring. The workflow is designed to minimise distraction, mitigate hand-eye coordination problems, and help guide monitoring of early decay during therapy in both clinical and mobile environments. The results from quantitative evaluations as well as a formative qualitative user study uncover the potentials of the proposed system and indicate that AR can serve as a promising tool in tooth decay management.