Factors In Cancer Cells Research Articles

SITP: A single cell bioinformatics analysis flow captures proteasome markers in the development of breast cancer.

Single cell sequencing and related databases have been widely used in the exploration of cancer occurrence and development, but there is still no in-depth explanation of specific and complicated cellular protein modification processes. Ubiquitin-Proteasome System (UPS), as a specific and precise protein modification and degradation process, plays an important role in the biological functions of cancer cell proliferation and apoptosis. Proteasomes, vital multi-catalytic proteinases in eukaryotic cells, play a crucial role in protein degradation and contribute to tumor regulation. The 26S proteasome, part of the ubiquitin-proteasome system. In this study, we have enrolled a common SITP process including analysis of single cell sequencing to elucidate a flow that can capture typical proteasome markers in the oncogenesis and progression of breast cancer. PSMD11, a key component of the 26S proteasome regulatory particle, has been identified as a critical survival factor in cancer cells. Results suggest that PSMD11's rapid degradation is linked to acute apoptosis in cancer cells, making it a potential target for cancer treatment. Our study explored the potential mechanisms of PSMD11 in breast cancer development. The findings revealed the feasibility of disclosing ubiquitinating biomarkers from public database, as well as presented new evidence supporting PSMD11 as a potential therapeutic biomarker for breast cancer.

Evolving Tumor Characteristics and Smart Nanodrugs for Tumor Immunotherapy.

Typical physiological characteristics of tumors, such as weak acidity, low oxygen content, and upregulation of certain enzymes in the tumor microenvironment (TME), provide survival advantages when exposed to targeted attacks by drugs and responsive nanomedicines. Consequently, cancer treatment has significantly progressed in recent years. However, the evolution and adaptation of tumor characteristics still pose many challenges for current treatment methods. Therefore, efficient and precise cancer treatments require an understanding of the heterogeneity degree of various factors in cancer cells during tumor evolution to exploit the typical TME characteristics and manage the mutation process. The highly heterogeneous tumor and infiltrating stromal cells, immune cells, and extracellular components collectively form a unique TME, which plays a crucial role in tumor malignancy, including proliferation, invasion, metastasis, and immune escape. Therefore, the development of new treatment methods that can adapt to the evolutionary characteristics of tumors has become an intense focus in current cancer treatment research. This paper explores the latest understanding of cancer evolution, focusing on how tumors use new antigens to shape their "new faces"; how immune system cells, such as cytotoxic T cells, regulatory T cells, macrophages, and natural killer cells, help tumors become "invisible", that is, immune escape; whether the diverse cancer-associated fibroblasts provide support and coordination for tumors; and whether it is possible to attack tumors in reverse. This paper discusses the limitations of targeted therapy driven by tumor evolution factors and explores future strategies and the potential of intelligent nanomedicines, including the systematic coordination of tumor evolution factors and adaptive methods, to meet this therapeutic challenge.

Abstract 511: Cell cycle inhibitors exhibit anti-tumor immunomodulatory roles through the HIF-1-DDX41 cytosolic DNA sensing pathway in HCC

Abstract Cell cycle inhibitors have a long history as anti-cancer therapeutics in clinical applications. Uncontrolled and infinite cell cycles are hallmarks of cancer, which enable them to proliferate rapidly and survive. To target these key features of cancer cells, cell cycle inhibitors are used to arrest their active cell cycle and induce cell death. In this study, beyond the known primary effect on cell cycle control, we identified that an expected and a secondary effect in immune modulation by the cell cycle inhibitors. Anti-mitotic agents including Paclitaxel (microtubule stabilizer), Palbociclib (CDK4/6 inhibitor) and AZD1152 and GSK1070916 (Aurora Kinase B inhibitors) can also eliminate cancer cells via an alternative mechanism - the activation of STING signaling. Our results demonstrated that these anti-mitotic agents caused DNA damage and cytosolic DNA accumulation. The cytosolic DNA was captured by a cytosolic DNA sensor DDX41,which triggered STING-TBK1-IRF3/7 pathway to upregulate secretion of pro-inflammatory senescence-associated secretory phenotype (SASP) factors in cancer cells. In addition, we revealed that transcription of DDX41 was mediated by the transcription factor known ashypoxia-inducible factor (HIF). In HCC, hypoxia induced DDX41 expression through HIF-1,thus hypoxic HCC cells were more prone to STING activation and SASP production under mitotic stress induced by cell cycle inhibitors. In the tumor microenvironment, the SASP enhanced infiltration of immune cells into the tumor core to eliminate cancer cells. We also observed the additive effects of cell cycles inhibitors (Paclitaxel, Palbociclib, and AZD1152)with anti-PD-1 mAb to arrest growth of HCC in mouse models. In summary, this study exhibited the novel immune-mediated aspect of cell cycle inhibitors in suppressing tumor growth. Our data suggested the potential combination regimen of cell cycle inhibitors and currently available immunotherapy with promising result. Citation Format: Cerise Yuen Ki Chan, Po Yee Wong, Helen Do Gai Xue, Chi Ching Goh, Jacinth Wing Sum Cheu, Aki Pui Wah Tse, Misty Shuo Zhang, Carmen Chak Lui Wong. Cell cycle inhibitors exhibit anti-tumor immunomodulatory roles through the HIF-1-DDX41 cytosolic DNA sensing pathway in HCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 511.

Trafficking of carbonic anhydrase 12 and bicarbonate transporters by histamine stimulation mediates intracellular acidic scenario in lung cancer cells

Carbonic anhydrase 12 is considered an oncogenic and acidic microenvironmental factor in cancer cells. To verify the role of histamine signalling as an anti-cancer signal, we determined the roles of CA12 and its associated bicarbonate transporters. In this study, histamine stimulation mediated mislocalization of CA12 in lung cancer cells. Histamine receptor activation-mediated CA12 endocytosis and pH were restored by CaMKII inhibition. CA12-associated AE2 expression was enhanced, whereas NBCn1 expression and its activity were reduced by histamine stimulation. Histamine receptor activation-mediated acidification was induced by internalised CA12 and NBCn1 and, at the same time by increased bicarbonate efflux through enhanced AE2 expression. Inhibition of protein trafficking by bafilomycin restored CA12 and AE2 localisation and diminished cellular acidosis. Thus, we verified that histamine stimulation induced an acidic scenario, which revealed trafficking of CA12 and its associated bicarbonate transporters in lung cancer cells and its dysregulated pH modulation may be involved in the histamine signalling-mediated anti-cancer process.

RXR agonist, Bexarotene, effectively reduces drug resistance via regulation of RFX1 in embryonic carcinoma cells

Aberrant expression of multidrug resistance (MDR) proteins is one of the features of cancer stem cells (CSCs) that make them escape chemotherapy. A well-orchestrated regulation of multiple MDRs by different transcription factors in cancer cells confers this drug resistance. An in silico analysis of the major MDR genes revealed a possible regulation by RFX1 and Nrf2. Previous reports also noted that Nrf2 is a positive regulator of MDR genes in NT2 cells. But we, for the first time, report that Regulatory factor X1 (RFX1), a pleiotropic transcription factor, negatively regulates the major MDR genes, Abcg2, Abcb1, Abcc1, and Abcc2, in NT2 cells. The levels of RFX1 in undifferentiated NT2 cells were found to be very low, which significantly increased upon RA-induced differentiation. Ectopic expression of RFX1 reduced the levels of transcripts corresponding to MDRs and stemness-associated genes. Interestingly, Bexarotene, an RXR agonist that acts as an inhibitor of Nrf2-ARE signaling, could increase the transcription of RFX1. Further analysis revealed that the RFX1 promoter has binding sites for RXRα, and upon Bexarotene exposure RXRα could bind and activate the RFX1 promoter. Bexarotene, alone or in combination with Cisplatin, could inhibit many cancer/CSC-associated properties in NT2 cells. Also, it significantly reduced the expression of drug resistance proteins and made the cells sensitive towards Cisplatin. Our study proves that RFX1 could be a potent molecule to target MDRs, and Bexarotene can induce RXRα-mediated RFX1 expression, therefore, would be a better chemo-assisting drug during therapy.

Senescent Secretome of Blind Mole Rat Spalax Inhibits Malignant Behavior of Human Breast Cancer Cells Triggering Bystander Senescence and Targeting Inflammatory Response

Subterranean blind mole rat, Spalax, has developed strategies to withstand cancer by maintaining genome stability and suppressing the inflammatory response. Spalax cells undergo senescence without the acquisition of senescence-associated secretory phenotype (SASP) in its canonical form, namely, it lacks the main inflammatory mediators. Since senescence can propagate through paracrine factors, we hypothesize that conditioned medium (CM) from senescent Spalax fibroblasts can transmit the senescent phenotype to cancer cells without inducing an inflammatory response, thereby suppressing malignant behavior. To address this issue, we investigated the effect of CMs of Spalax senescent fibroblasts on the proliferation, migration, and secretory profile in MDA-MB-231 and MCF-7 human breast cancer cells. The results suggest that Spalax CM induced senescence in cancer cells, as evidenced by increased senescence-associated beta-galactosidase (SA-β-Gal) activity, growth suppression and overexpression of senescence-related p53/p21 genes. Contemporaneously, Spalax CM suppressed the secretion of the main inflammatory factors in cancer cells and decreased their migration. In contrast, human CM, while causing a slight increase in SA-β-Gal activity in MDA-MB-231 cells, did not decrease proliferation, inflammatory response, and cancer cell migration. Dysregulation of IL-1α under the influence of Spalax CM, especially the decrease in the level of membrane-bound IL1-α, plays an important role in suppressing inflammatory secretion in cancer cells, which in turn leads to inhibition of cancer cell migration. Overcoming of SASP in tumor cells in response to paracrine factors of senescent microenvironment or anti-cancer drugs represents a promising senotherapeutic strategy in cancer treatment.

Targeting Chromatin-Remodeling Factors in Cancer Cells: Promising Molecules in Cancer Therapy.

ATP-dependent chromatin-remodeling complexes can reorganize and remodel chromatin and thereby act as important regulator in various cellular processes. Based on considerable studies over the past two decades, it has been confirmed that the abnormal function of chromatin remodeling plays a pivotal role in genome reprogramming for oncogenesis in cancer development and/or resistance to cancer therapy. Recently, exciting progress has been made in the identification of genetic alteration in the genes encoding the chromatin-remodeling complexes associated with tumorigenesis, as well as in our understanding of chromatin-remodeling mechanisms in cancer biology. Here, we present preclinical evidence explaining the signaling mechanisms involving the chromatin-remodeling misregulation-induced cancer cellular processes, including DNA damage signaling, metastasis, angiogenesis, immune signaling, etc. However, even though the cumulative evidence in this field provides promising emerging molecules for therapeutic explorations in cancer, more research is needed to assess the clinical roles of these genetic cancer targets.

ZAP70 Activation Compensates for Loss of Class IA PI3K Isoforms Through Activation of the JAK–STAT3 Pathway

Tyrosine kinases have crucial functions in cell signaling and proliferation. The phosphatidylinositol 3-kinase (PI3K) pathway is frequently deregulated in human cancer and is an essential regulator of cellular proliferation. We aimed to determine which tyrosine kinases contribute to resistance elicited by PI3K silencing and inhibition. To mimic catalytic inactivation of p110α/β, specific p110α (BYL719) and p110β (KIN193) inhibitors were used in addition to genetic knock-out in in vitro assays. Cell viability was assessed using crystal violet staining, whereas cellular transformation ability was analyzed by soft-agar growth assays. Activated zeta chain of T-cell receptor-associated protein kinase 70 (ZAP70) generated resistance to PI3K inhibition. This resistance was via activation of the Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) axis. We demonstrated that activated ZAP70 has a high transforming capability associated with the formation of malignant phenotype in untransformed cells and has the potential to be a tumor-initiating factor in cancer cells. ZAP70 may be a potent driver of proliferation and transformation in untransformed cells and is implicated in resistance to PI3K inhibitors in cancer cells.

The Fight against Cancer by Microgravity: The Multicellular Spheroid as a Metastasis Model.

Cancer is a disease exhibiting uncontrollable cell growth and spreading to other parts of the organism. It is a heavy, worldwide burden for mankind with high morbidity and mortality. Therefore, groundbreaking research and innovations are necessary. Research in space under microgravity (µg) conditions is a novel approach with the potential to fight cancer and develop future cancer therapies. Space travel is accompanied by adverse effects on our health, and there is a need to counteract these health problems. On the cellular level, studies have shown that real (r-) and simulated (s-) µg impact survival, apoptosis, proliferation, migration, and adhesion as well as the cytoskeleton, the extracellular matrix, focal adhesion, and growth factors in cancer cells. Moreover, the µg-environment induces in vitro 3D tumor models (multicellular spheroids and organoids) with a high potential for preclinical drug targeting, cancer drug development, and studying the processes of cancer progression and metastasis on a molecular level. This review focuses on the effects of r- and s-µg on different types of cells deriving from thyroid, breast, lung, skin, and prostate cancer, as well as tumors of the gastrointestinal tract. In addition, we summarize the current knowledge of the impact of µg on cancerous stem cells. The information demonstrates that µg has become an important new technology for increasing current knowledge of cancer biology.

Calcium/calmodulin-dependent protein kinase kinase 2 mediates pleiotropic effects of epidermal growth factor in cancer cells

AimsEngagement of epidermal growth factor (EGF) with its receptor (EGFR) produces a broad range of cancer phenotypes. The overriding aim of this study was to understand EGFR signaling and its regulation by the Ca2+/calmodulin (CaM) dependent protein kinase kinase 2 (CaMKK2) in cancer cells. ResultsIn ovarian cancer cells and other cancer cell types, EGF-induced activation of oncogenic Akt is mediated by both the canonical PI3K-PDK1 pathway and by CaMKK2. Akt activation induced by EGF occurs by both calcium-dependent and calcium-independent mechanisms. In contrast to the canonical pathway, CaMKK2 neither binds to, nor is regulated by phosphoinositides but is activated by Ca2+/CaM. Akt activation at its primary activation site, T308 occurs by direct phosphorylation by CaMKK2, but activation at its secondary site (S473), is through an indirect mechanism requiring mTORC2. In cells in which another CaMKK2 target, 5′AMP-dependent protein kinase (AMPK) was deleted, Akt activation and calcium-dependency of activation were still observed. CaMKK2 accumulates in the nucleus in response to EGF and regulates transcription of phosphofructokinase platelet (PFKP) a glycolytic regulator. CaMKK2 is required for optimal PFK activity. CaMKK2 regulates transcription of plasminogen activator, urokinase (PLAU) a metastasis regulator. The EGFR inhibitor gefitinib synergizes with CaMKK2 inhibition in the regulation of cell survival and increases the dose-reduction index. CRISPR/Cas9 knockout of CaMKK2 leads to compensatory PTEN downregulation and upregulation of Akt activation. ConclusionsCaMKK2-mediation of EGFR action may enable cancer cells to use intracellular calcium elevation as a signal for growth and survival.

Role of Base Excision Repair in Innate Immune Cells and Its Relevance for Cancer Therapy

Innate immunity is critical for immediate recognition and elimination of invading pathogens or defense against cancer cell growth. Dysregulation of innate immune systems is associated with the pathogenesis of different types of inflammatory diseases, including cancer. In addition, the maintenance of innate immune cells’ genomic integrity is crucial for the survival of all organisms. Oxidative stress generated from innate immune cells may cause self-inflicted DNA base lesions as well as DNA damage on others neighboring cells, including cancer cells. Oxidative DNA base damage is predominantly repaired by base excision repair (BER). BER process different types of DNA base lesions that are presented in cancer and innate immune cells to maintain genomic integrity. However, mutations in BER genes lead to impaired DNA repair function and cause insufficient genomic integrity. Moreover, several studies have implicated that accumulation of DNA damage leads to chromosomal instability that likely activates the innate immune signaling. Furthermore, dysregulation of BER factors in cancer cells modulate the infiltration of innate immune cells to the tumor microenvironment. In the current review, the role of BER in cancer and innate immune cells and its impact on innate immune signaling within the tumor microenvironment is summarized. This is a special issue that focuses on DNA damage and cancer therapy to demonstrate how BER inhibitor or aberrant repair modulates innate inflammatory response and impact immunotherapy approaches. Overall, the review provides substantial evidence to understand the impact of BER in innate immune response dynamics within the current immune-based therapeutic strategy.

PD-L1 promotes myofibroblastic activation of hepatic stellate cells by distinct mechanisms selective for TGF-β receptor I versus II.

SUMMARYIntrahepatic cholangiocarcinoma (ICC) contains abundant myofibroblasts derived from hepatic stellate cells (HSCs) through an activation process mediated by TGF-β. To determine the role of programmed death-ligand 1 (PD-L1) in myofibroblastic activation of HSCs, we disrupted PD-L1 of HSCs by shRNA or anti-PD-L1 antibody. We find that PD-L1, produced by HSCs, is required for HSC activation by stabilizing TGF-β receptors I (TβRI) and II (TβRII). While the extracellular domain of PD-L1 (amino acids 19–238) targets TβRII protein to the plasma membrane and protects it from lysosomal degradation, a C-terminal 260-RLRKGR-265 motif on PD-L1 protects TβRI mRNA from degradation by the RNA exosome complex. PD-L1 is required for HSC expression of tumor-promoting factors, and targeting HSC PD-L1 by shRNA or Cre/loxP recombination suppresses HSC activation and ICC growth in mice. Thus, myofibroblast PD-L1 can modulate the tumor microenvironment and tumor growth by a mechanism independent of immune suppression.

Alantolactone: A Natural Plant Extract as a Potential Therapeutic Agent for Cancer.

Alantolactone (ALT) is a natural compound extracted from Chinese traditional medicine Inula helenium L. with therapeutic potential in the treatment of various diseases. Recently, in vitro and in vivo studies have indicated cytotoxic effects of ALT on various cancers, including liver cancer, colorectal cancer, breast cancer, etc. The inhibitory effects of ALT depend on several cancer-associated signaling pathways and abnormal regulatory factors in cancer cells. Moreover, emerging studies have reported several promising strategies to enhance the oral bioavailability of ALT, such as combining ALT with other herbs and using ALT-entrapped nanostructured carriers. In this review, studies on the anti-tumor roles of ALT are mainly summarized, and the underlying molecular mechanisms of ALT exerting anticancer effects on cells investigated in animal-based studies are also discussed.

Design and Characterization of a Cell-Penetrating Peptide Derived from the SOX2 Transcription Factor.

SOX2 is an oncogenic transcription factor overexpressed in nearly half of the basal-like triple-negative breast cancers associated with very poor outcomes. Targeting and inhibiting SOX2 is clinically relevant as high SOX2 mRNA levels are positively correlated with decreased overall survival and progression-free survival in patients affected with breast cancer. Given its key role as a master regulator of cell proliferation, SOX2 represents an important scaffold for the engineering of dominant-negative synthetic DNA-binding domains (DBDs) that act by blocking or interfering with the oncogenic activity of the endogenous transcription factor in cancer cells. We have synthesized an interference peptide (iPep) encompassing a truncated 24 amino acid long C-terminus of SOX2 containing a potential SOX-specific nuclear localization sequence, and the determinants of the binding of SOX2 to the DNA and to its transcription factor binding partners. We found that the resulting peptide (SOX2-iPep) possessed intrinsic cell penetration and promising nuclear localization into breast cancer cells, and decreased cellular proliferation of SOX2 overexpressing cell lines. The novel SOX2-iPep was found to exhibit a random coil conformation predominantly in solution. Molecular dynamics simulations were used to characterize the interactions of both the SOX2 transcription factor and the SOX2-iPep with FGF4-enhancer DNA in the presence of the POU domain of the partner transcription factor OCT4. Predictions of the free energy of binding revealed that the iPep largely retained the binding affinity for DNA of parental SOX2. This work will enable the future engineering of novel dominant interference peptides to transport different therapeutic cargo molecules such as anti-cancer drugs into cells.

MicroRNA-324-5p-CUEDC2 Axis Mediates Gain-of-Function Mutant p53-Driven Cancer Stemness.

Regulation of cancer stemness has recently emerged as a new gain-of-function (GOF) property of mutant p53. In this study, we identify miR-324-5p as a critical epigenetic regulator of cancer stemness and demonstrate its role in mediating GOF-mutant p53-driven stemness phenotypes. We report that miR-324-5p is upregulated in human cancer cell lines and non-small cell lung carcinoma (NSCLC) tumors carrying TP53 GOF mutations. Mechanistically, we show that GOF mutant p53 upregulates miR-324-5p expression via c-Myc, an oncogenic transcription factor in cancer cells. Our experimental results suggest that miR-324-5p-induced CSC phenotypes stem from the downregulation of CUEDC2, a downstream target gene of miR-324-5p. Accordingly, CUEDC2 complementation diminishes elevated CSC marker expression in miR-324-5p-overexpressing cancer cells. We further demonstrate that mutant p53 cancer cells maintain a low level of CUEDC2 that is rescued upon miR-324-5p inhibition. Importantly, we identify CUEDC2 downregulation as a novel characteristic feature of TP53-mutated human cancers. We show that activation of NF-κB due to downregulation of CUEDC2 by miR-324-5p imparts stemness in GOF mutant p53 cancer cells. Finally, we provide evidence that TP53 mutations coupled with high miR-324-5p expression predict poor prognosis in patients with lung adenocarcinoma. Thus, our study delineates an altered miR-324-5p-CUEDC2-NF-κB pathway as a novel regulator of GOF mutant p53-driven cancer stemness. IMPLICATIONS: Our findings implicate miRNA-324-5p as a novel epigenetic modifier of human cancer stemness.

Effects of Lysophosphatidic Acid Receptor and Free Fatty Acid Receptor Agonists on CCN1/Cyr61 Expression in Prostate Cancer Cells

Expression of CCN1/Cyr61, a member of the CCN family of matricellular proteins, can be induced by growth factors in cancer cells. CCN1 participates in cell migration and other regulatory processes in various cancers. However, the role of CCN1 in prostate cancer remains poorly understood. In previous work we observed that lysophosphatidic acid (LPA), a bioactive lipid, stimulates proliferation and migration in prostate cancer cells via the G protein-coupled receptor (GPCR) LPAR1, and also induces CCN1 expression. We also showed that activation of free fatty acid receptor 4 (FFAR4), another GPCR, inhibits LPA-induced proliferation and migration. For the current study, we hypothesized that CCN1 contributes to adhesion and migration of prostate cancer cells. We first confirmed that CCN1 protein levels were increased by LPA after 2-5 hours in PC-3 human prostate cancer cells. This response was inhibited by TUG-891, an FFA4 agonist. When serum-starved PC-3 cells were treated with LPA in an extended time course from 2 minutes to 24 hours, Erk, Akt, and FAK were initially activated within 30 minutes of LPA addition, as detected by immunoblotting for the phosphorylated kinases. However, all three kinases exhibited a later phase of activation after ~6 hours. Thus, CCN1 induction is downstream of early signals initiated by LPAR1, but potentially upstream of later events. For adhesion assays, serum-starved PC-3 cells were incubated with and without 10 μM LPA ± 1 μM TUG-891. Enhanced adhesion was prominent 1 hour after LPA addition, and was sustained for at least 8 hours. For migration assays, serum-starved cells were incubated with the same agents in modified Boyden chambers. Enhanced cell migration was observed by 2 hours after LPA addition, and continued for at least 24 hours. LPA-induced adhesion and migration were blocked by TUG-891. In other experiments, extracellular matrix was subjected to immunoblotting and to immunofluorescence microscopy for CCN1. In the immunoblotting experiments, CCN1 protein was increased in the extracted extracellular matrix 2-5 hours after LPA addition. In the immunohistochemistry experiments, CCN1 was detected in the extracellular space of PC-3 cells treated with LPA for 0-5 hours. In conclusion, the results suggest a role for LPA-induced CCN1 in adhesion and migration in prostate cancer cells, making CCN1 a potential therapeutic target. In addition, the results further indicate that FFAR activation is a strategy to suppress both short- and long-term LPA responses.

An exonuclease protection and CRISPR/Cas12a integrated biosensor for the turn-on detection of transcription factors in cancer cells

Transcription factors (TFs) are critical proteins that regulate the expression of genes, and the abnormal change of TFs levels is directly related to physical dysfunctions. Herein, we developed a clustered regularly interspaced short palindromic repeats (CRISPR)-based biosensor for the measurement of TFs level with the assistance of exonuclease protection assay. A dsDNA (activator) with the ability to activate Cas12a was engineered to contain TFs binding domain, and the binding between TFs and the activator can protect the dsDNA from being digested by exonuclease III (Exo III). The reserved activator then triggered a CRISPR/Cas12a reporting reaction to produce fluorescent signal for detection. In the detection of nuclear factor-kappa B (NF-κB) p50 subunit, the limit of detection of 0.2 pM and limit of quantification of 0.6 pM were obtained respectively, and the performance of this biosensor has been challenged by cell nucleoprotein extracts. Additionally, this method can be applied in the screening and evaluation of TFs inhibitors, calculating the IC50 of oridonin. Integrating merits including high sensitivity, low cost, and good portability, this method may enrich the arsenal for TFs-related applications.

Screening of novel alkaloid inhibitors for vascular endothelial growth factor in cancer cells: an integrated computational approach.

Vascular endothelial growth factor (VEGF) is expressed at elevated levels by most cancer cells, which can stimulate vascular endothelial cell growth, survival, proliferation as well as trigger angiogenesis modulated by VEGF and VEGFR (a tyrosine kinase receptor) signaling. The angiogenic effects of the VEGF family are thought to be primarily mediated through the interaction of VEGF with VEGFR-2. Targeting this signaling molecule and its receptor is a novel approach for blocking angiogenesis. In recent years virtual high throughput screening has emerged as a widely accepted powerful technique in the identification of novel and diverse leads. The high-resolution X-ray structure of VEGF has paved the way to introduce new small molecular inhibitors by structure-based virtual screening. In this study using different alkaloid molecules as potential novel inhibitors of VEGF, we proposed three alkaloid candidates for inhibiting VEGF and VEGFR mediated angiogenesis. As these three alkaloid compounds exhibited high scoring functions, which also highlights their high binding ability, it is evident that these alkaloids can be taken to further drug development pipelines for use as novel lead compounds to design new and effective drugs against cancer.

Generation and identification of a conditional knockout allele for the PSMD11 gene in mice

BackgroundOur previous study have shown that the PSMD11 protein was an important survival factor for cancer cells except for its key role in regulation of assembly and activity of the 26S proteasome. To further investigate the role of PSMD11 in carcinogenesis, we constructed a conditional exon 5 floxed allele of PSMD11 (PSMD11flx) in mice.ResultsIt was found that homozygous PSMD11 flx/flx mice showed normal and exhibited a normal life span and fertility, and showed roughly equivalent expression of PSMD11 in various tissues, suggesting that the floxed allele maintained the wild-type function. Cre recombinase could induce efficient knockout of the floxed PSMD11 allele both in vitro and in vivo. Mice with constitutive single allele deletion of PSMD11 derived from intercrossing between PSMD11flx/flx and CMV-Cre mice were all viable and fertile, and showed apparent growth retardation, suggesting that PSMD11 played a significant role in the development of mice pre- or postnatally. No whole-body PSMD11 deficient embryos (PSMD11−/−) were identified in E7.5–8.5 embryos in uteros, indicating that double allele knockout of PSMD11 leads to early embryonic lethality. To avoid embryonic lethality produced by whole-body PSMD11 deletion, we further developed conditional PSMD11 global knockout mice with genotype Flp;FSF-R26CAG − CreERT2/+; PSMD11flx/flx, and demonstrated that PSMD11 could be depleted in a temporal and tissue-specific manner. Meanwhile, it was found that depletion of PSMD11 could induce massive apoptosis in MEFs.ConclusionsIn summary, our data demonstrated that we have successfully generated a conditional knockout allele of PSMD11 in mice, and found that PSMD11 played a key role in early and postnatal development in mice, the PSMD11 flx/flx mice will be an invaluable tool to explore the functions of PSMD11 in development and diseases.

Pluripotency Stemness and Cancer: More Questions than Answers.

Embryonic stem cells and induced pluripotent stem cells provided us with fascinating new knowledge in recent years. Mechanistic insight into intricate regulatory circuitry governing pluripotency stemness and disclosing parallels between pluripotency stemness and cancer instigated numerous studies focusing on roles of pluripotency transcription factors, including Oct4, Sox2, Klf4, Nanog, Sall4 and Tfcp2L1, in cancer. Although generally well substantiated as tumour-promoting factors, oncogenic roles of pluripotency transcription factors and their clinical impacts are revealing themselves as increasingly complex. In certain tumours, both Oct4 and Sox2 behave as genuine oncogenes, and reporter genes driven by composite regulatory elements jointly recognized by both the factors can identify stem-like cells in a proportion of tumours. On the other hand, cancer stem cells seem to be biologically very heterogeneous both among different tumour types and among and even within individual tumours. Pluripotency transcription factors are certainly implicated in cancer stemness, but do not seem to encompass its entire spectrum. Certain cancer stem cells maintain their stemness by biological mechanisms completely different from pluripotency stemness, sometimes even by engaging signalling pathways that promote differentiation of pluripotent stem cells. Moreover, while these signalling pathways may well be antithetical to stemness in pluripotent stem cells, they may cooperate with pluripotency factors in cancer stem cells - a paradigmatic example is provided by the MAPK-AP-1 pathway. Unexpectedly, forced expression of pluripotency transcription factors in cancer cells frequently results in loss of their tumour-initiating ability, their phenotypic reversion and partial epigenetic normalization. Besides the very different signalling contexts operating in pluripotent and cancer stem cells, respectively, the pronounced dose dependency of reprogramming pluripotency factors may also contribute to the frequent loss of tumorigenicity observed in induced pluripotent cancer cells. Finally, contradictory cell-autonomous and non-cell-autonomous effects of various signalling molecules operate during pluripotency (cancer) reprogramming. The effects of pluripotency transcription factors in cancer are thus best explained within the concept of cancer stem cell heterogeneity.