Mesenchymal Phenotype Research Articles

Gene expression-phenotype association study reveals the dual role of TNF-α/TNFR1 signaling axis in confined breast cancer cell migration

AimsWhile enhanced tumor cell migration is a key process in the tumor dissemination, mechanistic insights into causal relationships between tumor cells and mechanical confinement are still limited. Here we combine the use of microfluidic platforms to characterize confined cell migration with genomic tools to systematically unravel the global signaling landscape associated with the migratory phenotype of breast cancer (BC) cells. Meterials and methodsThe spontaneous migration capacity of seven BC cell lines was evaluated in 3D microfluidic devices and their migration capacity was correlated with publicly available molecular signatures. The role of identified signaling pathways on regulating BC migration capacity was determined by receptor stimulation through ligand binding or inhibition through siRNA silencing. Downstream effects on cell migration were evaluated in microfluidic devices, while the molecular changes were monitored by RT-qPCR. Key findingsExpression of 715 genes was correlated with BC cells migratory phenotype, revealing TNF-α as one of the top upstream regulators. Signal transduction experiments revealed that TNF-α stimulates the confined migration of triple negative, mesenchymal-like BC cells that are also characterized by high TNFR1 expression, but inhibits the migration of epithelial-like cells with low TNFR1 expression. TNFR1 was strongly associated with the migration capacity and triple-negative, mesenchymal phenotype. Downstream of TNF/TNFR1 signaling, transcriptional regulation of NFKB seems to be important in driving cell migration in confined spaces. SignificanceTNF-α/TNFR1 signaling axis reveals as a key player in driving BC cells confined migration, emerging as a promising therapeutic strategy in targeting dissemination and metastasis of triple negative, mesenchymal BC cells.

DDX19A promotes gastric cancer cell proliferation and migration by activating the PI3K/AKT pathway

BackgroundDEAD-box RNA helicase 19 A (DDX19A) is overexpressed in cervical squamous cell carcinoma. However, its role in gastric cancer remains unclear. The present study aimed to explore the role and underlying mechanism of DDX19A in the development of gastric cancer.MethodsThe expression of DDX19A in gastric cancer and paracancerous tissues was evaluated through quantitative polymerase chain reaction, western blotting, and immunohistochemical staining. The biological functions of DDX19A in gastric cancer were determined using CCK8, plate colony-forming, and Transwell migration assays. The specific mechanism of DDX19A in gastric cancer cells was studied using western blotting, RNA-binding protein immunoprecipitation, mRNA half-life detection, and nuclear and cytoplasmic RNA isolation.ResultsDDX19A was highly expressed in gastric cancer and positively associated with malignant clinicopathological features and poor prognosis. Additionally, DDX19A promoted gastric cancer cell proliferation, migration, and epithelial–mesenchymal transition phenotypes. Mechanistically, DDX19A activated the PI3K/AKT pathway by upregulating phosphatidylinositol-3-kinase (PIK3CA) expression. Furthermore, DDX19A interacted with PIK3CA mRNA, stabilized it, and facilitated its export from the nucleus.ConclusionsOur study reveals a novel mechanism whereby DDX19A promotes the proliferation and migration of gastric cancer cells by enhancing the stability and nuclear export of PIK3CA mRNA, thereby activating the PI3K/AKT pathway.

Microtubule-associated NAV3 regulates invasive phenotypes in glioblastoma cells.

Glioblastomas are aggressive brain tumors for which effective therapy is still lacking, resulting in dismal survival rates. These tumors display significant phenotypic plasticity, harboring diverse cell populations ranging from tumor core cells to dispersed, highly invasive cells. Neuron navigator 3 (NAV3), a microtubule-associated protein affecting microtubule growth and dynamics, is downregulated in various cancers, including glioblastoma, and has thus been considered a tumor suppressor. In this study, we challenge this designation and unveil distinct expression patterns of NAV3 across different invasion phenotypes. Using glioblastoma cell lines and patient-derived glioma stem-like cell cultures, we disclose an upregulation of NAV3 in invading glioblastoma cells, contrasting with its lower expression in cells residing in tumor spheroid cores. Furthermore, we establish an association between low and high NAV3 expression and the amoeboid and mesenchymal invasive phenotype, respectively, and demonstrate that overexpression of NAV3 directly stimulates glioblastoma invasive behavior in both 2D and 3D environments. Consistently, we observed increased NAV3 expression in cells migrating along blood vessels in mouse xenografts. Overall, our results shed light on the role of NAV3 in glioblastoma invasion, providing insights into this lethal aspect of glioblastoma behavior.

Trabecular meshwork cell differentiation in response to collagen and TGFβ-2 spatial interactions

Primary open-angle glaucoma (POAG) is currently the most prevalent cause of irreversible blindness globally. To date, few in vitro models that can faithfully recapitulate the complex architecture of the trabecular meshwork (TM) and the specialised trabecular meshwork cell (TMC) characteristics that are local to the structurally opposing regions. This study aimed to investigate the parameters that govern TMC phenotype by adapting the extracellular matrix structure to mimic the juxtacanalicular tissue (JCT) region of the TM. Initially, TMC phenotypic characteristics were investigated within type I collagen matrices of controlled fiber density and anisotropy, generated through confined plastic compression (PC). Notably, PC-collagen presented biophysical cues that induced JCT cellular characteristics (elastin, α-β-Crystallin protein expression, cytoskeletal remodelling, increased mesenchymal markers and JCT-specific genetic markers). In parallel, a pathological mesenchymal phenotype associated with POAG was induced through localised transforming growth factor -beta 2 (TGFβ-2) exposure. This resulted in a profile of alternative mesenchymal states (fibroblast/smooth muscle or myofibroblast) displayed by the TMC in vitro. Overall, the study provides an advanced insight into the biophysical cues that modulate TMC fate, inducing a JCT-specific phenotype and transient mesenchymal characteristics that reflect healthy and pathological scenarios. Statement of significanceGlaucoma is a leading cause of blindness, with a lack of long-term efficacy within current drug candidates. Reliable trabecular meshwork (TM) in vitro models will be critical for enhancing the fields understanding of healthy and disease states for pre-clinical testing.Trabecular meshwork cells (TMCs) display heterogeneity throughout the hierarchical TM, however our understanding into recapitulating these phenotypes in vitro, remains elusive. This study hypothesizes the importance of specific matrix/growth factor spatial stimuli in governing TMCs phenotype. By emulating certain biophysical/biochemical in vivo parameters, we introduce an advanced profile of distinct TMC phenotypic states, reflecting healthy and disease scenarios. A notion that has not be stated prior and a fundamental consideration for future 3D TM in vitro modelling.

Pressure induces peritoneal fibrosis and inflammation through CD44 signaling

Peritoneal dialysis (PD) is a widely used sustainable kidney replacement therapy. Prolonged use of PD fluids is associated with mesothelial-mesenchymal transition, peritoneal fibrosis, and eventual ultrafiltration (UF) failure. However, the impact of pressure on the peritoneum remains unclear. In the present study, we hypothesized increased pressure is a potential contributing factor to peritoneal fibrosis and investigated the possible mechanisms. In vitro experiments found that pressurization led to a mesenchymal phenotype, the expression of fibrotic markers and inflammatory factors in human mesothelial MeT-5A cells. Pressure also increased cell proliferation and augmented cell migration potential in MeT-5A cells. The mouse PD model and human peritoneum equilibrium test (PET) data both showed a positive association between higher pressure and increased small solute transport, along with decreased net UF. Mechanistically, we found that significant upregulation of CD44 in mesothelial cells upon pressurization. Notably, the treatment of CD44 neutralizing antibodies prevented pressure-induced phenotypic changes in mesothelial cells, while a CD44 inhibitor oligo-fucoidan ameliorated pressure-induced peritoneal thickening, fibrosis, and inflammation in PD mice. To conclude, intraperitoneal pressure results in peritoneal fibrosis in PD via CD44-mediated mesothelial changes and inflammation. CD44 blockage can be utilized as a novel preventive approach for PD-related peritoneal fibrosis and UF failure.

Single-cell profiling of surface glycosphingolipids opens a new dimension for deconvolution of breast cancer intratumoral heterogeneity and phenotypic plasticity

Glycosylated sphingolipids (GSLs) are a diverse group of cellular lipids typically reported as being rare in normal mammary tissue. In breast cancer (BCa), GSLs have emerged as noteworthy markers associated with breast cancer stem cells, mediators of phenotypic plasticity, and contributors to cancer cell chemoresistance. GSLs are potential surface markers that can uniquely characterize the heterogeneity of the tumor microenvironment, including cancer cell subpopulations and epithelial–mesenchymal plasticity (EMP). In this study, mass spectrometry analyses of the total sphingolipidome in breast epithelial cells and their mesenchymal counterparts revealed increased levels of Gb3 in epithelial cells and significantly elevated GD2 levels in the mesenchymal phenotype. To elucidate if GSL-related epitopes on BCa cell surfaces reflect EMP and cancer status, we developed and rigorously validated a 12-color spectral flow cytometry panel. This panel enables the simultaneous detection of native GSL epitopes (Gb3, SSEA1, SSEA3, SSEA4, and GD2), epithelial–mesenchymal transition markers (EpCAM, TROP2, and CD9), and lineage markers (CD45, CD31, and CD90) at the single-cell level. Next, the established panel was used for the analysis of BCa primary tumors and revealed surface heterogeneity in SSEA1, SSEA3, SSEA4, GD2, and Gb3, indicative of native epitope presence also on non-tumor cells. These findings further highlighted the phenotype-dependent alterations in GSL surface profiles, with differences between epithelial and stromal cells in the tumor. This study provides novel insights into BCa heterogeneity, shedding light on the potential of native GSL-related epitopes as markers for EMP and cancer status in fresh clinical samples. The developed single-cell approach offers promising avenues for further exploration.

Phenotypic Plasticity and Cancer: A System Biology Perspective.

Epithelial-to-mesenchymal transition (EMT) is a major axis of phenotypic plasticity not only in diseased conditions such as cancer metastasis and fibrosis but also during normal development and wound healing. Yet-another important axis of plasticity with metastatic implications includes the cancer stem cell (CSCs) and non-CSC transitions. However, in both processes, epithelial (E) and mesenchymal (M) phenotypes are not merely binary states. Cancer cells acquire a spectrum of phenotypes with traits, properties, and markers of both E and M phenotypes, giving rise to intermediary hybrid (E/M) phenotypes. E/M cells play an important role in tumor initiation, metastasis, and disease progression in multiple cancers. Furthermore, the hybrid phenotypes also play a major role in causing therapeutic resistance in cancer. Here, we discuss how a systems biology perspective on the problem, which is implicit in the 'Team Medicine' approach outlined in the theme of this Special Issue of The Journal of Clinical Medicine and includes an interdisciplinary team of experts, is more likely to shed new light on EMT in cancer and help us to identify novel therapeutics and strategies to target phenotypic plasticity in cancer.

Preclinical spheroid models identify BMX as a therapeutic target for metastatic MYCN nonamplified neuroblastoma.

The development of targeted therapies offers new hope for patients affected by incurable cancer. However, multiple challenges persist, notably in controlling tumor cell plasticity in patients with refractory and metastatic illness. Neuroblastoma (NB) is an aggressive pediatric malignancy originating from defective differentiation of neural crest-derived progenitors with oncogenic activity due to genetic and epigenetic alterations and remains a clinical challenge for high-risk patients. To identify critical genes driving NB aggressiveness, we performed combined chromatin and transcriptome analyses on matched patient-derived xenografts (PDXs), spheroids, and differentiated adherent cultures derived from metastatic MYCN nonamplified tumors. Bone marrow kinase on chromosome X (BMX) was identified among the most differentially regulated genes in PDXs and spheroids versus adherent models. BMX expression correlated with high tumor stage and poor patient survival and was crucial to the maintenance of the self-renewal and tumorigenic potential of NB spheroids. Moreover, BMX expression positively correlated with the mesenchymal NB cell phenotype, previously associated with increased chemoresistance. Finally, BMX inhibitors readily reversed this cellular state, increased the sensitivity of NB spheroids toward chemotherapy, and partially reduced tumor growth in a preclinical NB model. Altogether, our study identifies BMX as a promising innovative therapeutic target for patients with high-risk MYCN nonamplified NB.

Zeb1 mediates EMT/plasticity-associated ferroptosis sensitivity in cancer cells by regulating lipogenic enzyme expression and phospholipid composition

Therapy resistance and metastasis, the most fatal steps in cancer, are often triggered by a (partial) activation of the epithelial–mesenchymal transition (EMT) programme. A mesenchymal phenotype predisposes to ferroptosis, a cell death pathway exerted by an iron and oxygen-radical-mediated peroxidation of phospholipids containing polyunsaturated fatty acids. We here show that various forms of EMT activation, including TGFβ stimulation and acquired therapy resistance, increase ferroptosis susceptibility in cancer cells, which depends on the EMT transcription factor Zeb1. We demonstrate that Zeb1 increases the ratio of phospholipids containing pro-ferroptotic polyunsaturated fatty acids over cyto-protective monounsaturated fatty acids by modulating the differential expression of the underlying crucial enzymes stearoyl-Co-A desaturase 1 (SCD), fatty acid synthase (FASN), fatty acid desaturase 2 (FADS2), elongation of very long-chain fatty acid 5 (ELOVL5) and long-chain acyl-CoA synthetase 4 (ACSL4). Pharmacological inhibition of selected lipogenic enzymes (SCD and FADS2) allows the manipulation of ferroptosis sensitivity preferentially in high-Zeb1-expressing cancer cells. Our data are of potential translational relevance and suggest a combination of ferroptosis activators and SCD inhibitors for the treatment of aggressive cancers expressing high Zeb1.

An MITF- and mTOR-dependent FLCN pathway suppresses TFE3-driven metastasis in melanoma.

Cancer cells have remarkable plasticity allowing them to acquire many biological states. Melanoma cells have the ability to switch from a proliferative melanocytic state to an invasive mesenchymal state and back again resulting in intratumoral heterogeneity. While microphthalmia-associated transcription factor (MITF) promotes the melanocytic phenotype, it is unclear what transcription factors drive the mesenchymal phenotype, and what mechanisms regulate the switch from the proliferative state to the mesenchymal state. We show that nuclear localization of the MITF paralog TFE3 correlates positively with metastatic potential in melanoma cell lines and tumors, and that deletion of TFE3 in MITF-low melanoma cell lines eliminates migration and metastatic ability. Further, we find that MITF suppresses the mesenchymal phenotype by activating expression of FNIP2, which encodes a component of an mTORC1-stimulated pathway promoting cytoplasmic retention and lysosomal degradation of TFE3. These findings point to the mTOR pathway and TFE3 as key regulators of melanoma plasticity.

Unveiling the role of TAGLN2 in glioblastoma: From proneural-mesenchymal transition to Temozolomide resistance

Glioblastoma (GBM) presents a daunting challenge due to its resistance to temozolomide (TMZ), a hurdle exacerbated by the proneural-to-mesenchymal transition (PMT) from a proneural (PN) to a mesenchymal (MES) phenotype. TAGLN2 is prominently expressed in GBM, particularly in the MES subtype compared to low-grade glioma (LGG) and the PN subtype. Our research reveals TAGLN2's involvement in PMT and TMZ resistance through a series of in vitro and in vivo experiments. TAGLN2 knockdown can restrain proliferation and invasion, trigger DNA damage and apoptosis, and heighten TMZ sensitivity in GBM cells. Conversely, elevating TAGLN2 levels amplifies resistance to TMZ in cellular and intracranial xenograft mouse models. We demonstrate the interaction relationship between TAGLN2 and ERK1/2 through co-immunoprecipitation (Co-IP) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) spectrometry analysis. Knockdown of TAGLN2 results in a decrease in the expression of p-ERK1/2, whereas overexpression of TAGLN2 leads to an increase in p-ERK1/2 expression within the nucleus. Subsequently, the regulatory role of TAGLN2 in the expression and control of MGMT has been demonstrated. Finally, the regulation of TAGLN2 by NF-κB has been validated through chromatin immunoprecipitation and ChIP-PCR assays. In conclusion, our results confirm that TAGLN2 exerts its biological functions by interacting with the ERK/MGMT axis and being regulated by NF-κB, thereby facilitating the acquisition of promoting PMT and increased resistance to TMZ therapy in glioblastoma. These results provide valuable insights for the advancement of targeted therapeutic approaches to overcome TMZ resistance in clinical treatments.

Acupuncture improve proteinuria in diabetic kidney disease rats by inhibiting ferroptosis and epithelial-mesenchymal transition

ObjectiveTo explore the mechanism of acupuncture to relieve diabetic proteinuria in Diabetic Kidney Disease (DKD). MethodsA total of 10 male Sprague-Dawley rats were randomly selected as the negative control group (NC), and a further 30 rats were fed a high-fat diet (HFD) and intraperitoneal streptozotocin (STZ). The DKD model rats in the model group (DKD) and the acupuncture group (DKD + Acu) were randomly assigned. After 4 weeks of intervention, collected urine, peripheral blood, and renal tissues from all rats, and assessed blood urea nitrogen, serum creatinine, triglyceride, 24-h urine protein quantification, and blood glucose, left kidney weight, kidney body ratio index, then observe changes in renal histology in the rats. The renal cortex tissues of three rats from each group were sent for transcriptomic analysis. According to the results of transcriptomic analysis, various kits were used to detect SOD, MDA, GSH, GSH-px, and iron concentration. The expression levels of GPX4 and System Xc-, members of the antioxidative stress pathway, and TfR 1, SLC39A14, FTH 1, and SLC40A1, involved in iron metabolism, in the kidney tissues were measured by western blotting and reverse transcription-quantitative PCR. The expression of mesenchymal phenotype markers and podocyte-specific markers were evaluated by immunofluorescence. ResultsAcupuncture promoted the levels of GSH, GSH-px, and SOD, decreased the level of MDA (P < 0.05), promoted the expression of GPX4 and System Xc- (P < 0.05), decreased the expression of TfR1 and SLC39A14 (P < 0.01), and increased the expression of FTH 1 and SLC40A1 (P < 0.05), inhibited the expression of TGF-β1, desmin, FSP-1, and α-SMA (P < 0.05), promoted the expression of Nephrin, Podocin, and CD2AP (P < 0.05). ConclusionImproving the ability of podocytes to prevent oxidative stress and restoring iron ion homeostasis, can improve Ferroptosis and block epithelial-mesenchymal transition, improve podocyte injury, restore filtration function, and reduce proteinuria in DKD rats.

Red light-activated depletion of drug-refractory glioblastoma stem cells and chemosensitization of an acquired-resistant mesenchymal phenotype.

Glioblastoma stem cells (GSCs) are potent tumor initiators resistant to radiochemotherapy, and this subpopulation is hypothesized to re-populate the tumor milieu due to selection following conventional therapies. Here, we show that 5-aminolevulinic acid (ALA) treatment-a pro-fluorophore used for fluorescence-guided cancer surgery-leads to elevated levels of fluorophore conversion in patient-derived GSC cultures, and subsequent red light-activation induces apoptosis in both intrinsically temozolomide chemotherapy-sensitive and -resistant GSC phenotypes. Red light irradiation of ALA-treated cultures also exhibits the ability to target mesenchymal GSCs (Mes-GSCs) with induced temozolomide resistance. Furthermore, sub-lethal light doses restore Mes-GSC sensitivity to temozolomide, abrogating GSC-acquired chemoresistance. These results suggest that ALA is not only useful for fluorescence-guided glioblastoma tumor resection, but that it also facilitates a GSC drug-resistance agnostic, red light-activated modality to mop up the surgical margins and prime subsequent chemotherapy.

KLF5 inhibition initiates epithelial-mesenchymal transition in non-transformed human squamous epithelial cells

The transcriptional regulator Krüppel-like factor 5 (KLF5) is highly expressed in squamous epithelial cells of the esophagus. Increased KLF5 activity induces tumorigenesis and promotes metastasis in several cancers, although this function appears to be context-dependent. Here, we demonstrate that acute KLF5 inhibition, both genetically and with the potent KLF5 inhibitor ML264, causes non-transformed human primary esophageal squamous epithelial cells to enter the epithelial to mesenchymal transition (EMT). Moreover, chronic KLF5 inhibition with ML264 leads to the development of cells with a mesenchymal phenotype characterized by the expression of mesenchymal markers and functionally by reduced cell growth and increased migration and cellular invasion. This EMT resulting from chronic KLF5 inhibition is not driven by β-Catenin or TGF-β signaling. Pharmacologically, ML264 inhibits KLF5 by promoting proteasomal-mediated degradation. Taken together, we demonstrate that reduced KLF5 activity reprograms epithelial cells towards a mesenchymal phenotype and enhances their migratory and invasive potential. These findings have potential implications not only for esophageal cancers but also for normal processes such as esophageal tissue repair following injury.

Identification of New Chemoresistance-Associated Genes in Triple-Negative Breast Cancer by Single-Cell Transcriptomic Analysis.

Triple-negative breast cancer (TNBC) is a particularly aggressive mammary neoplasia with a high fatality rate, mainly because of the development of resistance to administered chemotherapy, the standard treatment for this disease. In this study, we employ both bulk RNA-sequencing and single-cell RNA-sequencing (scRNA-seq) to investigate the transcriptional landscape of TNBC cells cultured in two-dimensional monolayers or three-dimensional spheroids, before and after developing resistance to the chemotherapeutic agents paclitaxel and doxorubicin. Our findings reveal significant transcriptional heterogeneity within the TNBC cell populations, with the scRNA-seq identifying rare subsets of cells that express resistance-associated genes not detected by the bulk RNA-seq. Furthermore, we observe a partial shift towards a highly mesenchymal phenotype in chemoresistant cells, suggesting the epithelial-to-mesenchymal transition (EMT) as a prevalent mechanism of resistance in subgroups of these cells. These insights highlight potential therapeutic targets, such as the PDGF signaling pathway mediating EMT, which could be exploited in this setting. Our study underscores the importance of single-cell approaches in understanding tumor heterogeneity and developing more effective, personalized treatment strategies to overcome chemoresistance in TNBC.

Cadherin Expression Profiles Define Glioblastoma Differentiation and Patient Prognosis.

Cadherins are cell-cell adhesion proteins which have been strongly implicated in cancer invasion, dissemination and metastasis capacity; thus, they are key players in the epithelial-to-mesenchymal transition (EMT) program. However, their role in glioblastoma (GBM), a primary central nervous system aggressive tumor, remains to be clarified. N-, E- and P-cadherin expression was analyzed on a large series of GBMs, characterized with clinical, imaging and neuropathological parameters, as well as with patients' survival data. In addition, cadherins' expression was studied in match-recurrent cases. Using TCGA data, cadherin expression profiles were also evaluated according to GBM transcription subtypes. N-cadherin expression was observed in 81.5% of GBM, followed by E-cadherin in 31% and P-cadherin in 20.8%. Upon tumor recurrence, P-cadherin was the only significantly upregulated cadherin compared with the primary tumor, being positive in 65.8% of the cases. Actually, P-cadherin gain was observed in 51.4% of matched primary-recurrent cases. Cadherins' co-expression was also explored. Interestingly, E- and N-cadherin co-expression identified a GBM subgroup with frequent epithelial differentiation and a significant survival benefit. On the other hand, subgroups with P-cadherin expression carried the worse prognosis. P- and N-cadherin co-expression correlated with the presence of a mesenchymal phenotype. Expressions of isolated P-cadherin or E- and P-cadherin co-expression were associated with imaging characteristics of aggressiveness, to highly heterogeneous tumors, an d to worse patient survival. Classical cadherins co-expression subgroups present consistent clinical, imaging, neuropathological and survival differences, which probably reflect different states of an EMT-like program in GBM.

Anti-PEc: Development of a novel monoclonal antibody against prostate cancer.

The Ec peptide (PEc) that defines the IGF-1Ec isoform, is associated with prostate cancer progression by inducing proliferation, metastases, and tumour repair. On these grounds, an anti-PEc monoclonal antibody (MAb) was developed. Our objective is to examine the effects of this antibody on prostate cancer and its possible side effects. The effects of the obtained MAb were examined in cancer and non-cancerous cell lines (unmodified and modified either to overexpress or silence PEc) and in tumours in SCID mice injected with unmodified prostate cancer cells. The investigation was obtained with respect to cellular proliferation, migration, invasion, toxicity to tumours, effects on the cell cycle, immune response activation, effects on mesenchymal stem cell mobilisation leading to tumour repair, tissue distribution, and toxicity to mice. Anti-PEc MAb treatment led to a significant decrease in cellular proliferation, migration, and invasion compared to the untreated cell lines (p < 0.0005 in every case). Mechanistically, these effects were associated with the downregulation of pERK1/2 and vimentin and the upregulation of E-Cadherin. In vivo, anti-PEc MAb treatment was associated with a significant decrease in tumour size and metastases rate (p < 0.0005 in every case) by reversing the tumours mesenchymal phenotype. It also inhibited host stem cell mobilisation towards the tumour, leading to apoptosis. Anti-PEc MAb assessment in respect to distribution and toxicity, indicated its tumour specificity and lack of toxicity. These data indicate that the therapeutic targeting of PEc with the anti-PEc MAb may have considerable clinical benefit for prostate cancer patients.

Dissecting gastric cancer heterogeneity and exploring therapeutic strategies using bulk and single-cell transcriptomic analysis and experimental validation of tumor microenvironment and metabolic interplay.

Gastric cancer, the fifth most prevalent cancer worldwide, is often diagnosed in advanced stages with limited treatment options. Examining the tumor microenvironment (TME) and its metabolic reprogramming can provide insights for better diagnosis and treatment. This study investigates the link between TME factors and metabolic activity in gastric cancer using bulk and single-cell RNA-sequencing data. We identified two molecular subtypes in gastric cancer by analyzing the distinct expression patterns of 81 prognostic genes related to the TME and metabolism, which exhibited significant protein-level interactions. The high-risk subtype had increased stromal content, fibroblast and M2 macrophage infiltration, elevated glycosaminoglycans/glycosphingolipids biosynthesis, and fat metabolism, along with advanced clinicopathological features. It also exhibited low mutation rates and microsatellite instability, associating it with the mesenchymal phenotype. In contrast, the low-risk group showed higher tumor content and upregulated protein and sugar metabolism. We identified a 15-gene prognostic signature representing these characteristics, including CPVL, KYNU, CD36, and GPX3, strongly correlated with M2 macrophages, validated through single-cell analysis and an internal cohort. Despite resistance to immunotherapy, the high-risk group showed sensitivity to molecular targeted agents directed at IGF-1R (BMS-754807) and the PI3K-mTOR pathways (AZD8186, AZD8055). We experimentally validated these promising drugs for their inhibitory effects on MKN45 and MKN28 gastric cells. This study unveils the intricate interplay between TME and metabolic pathways in gastric cancer, offering potential for enhanced diagnosis, patient stratification, and personalized treatment. Understanding molecular features in each subtype enriches our comprehension of gastric cancer heterogeneity and potential therapeutic targets.

IFIT1 + neutrophil is a causative factor of immunosuppressive features of poorly cohesive carcinoma (PCC)

The importance of the immune microenvironment in poorly cohesive carcinoma (PCC) has been highlighted due to its limited response rate to conventional therapy and emerging treatment resistance. A combination of clinical cohorts, bioinformatics analyses, and functional/molecular experiments revealed that high infiltration of Interferon Induced Protein with Tetratricopeptide Repeats 1 (IFIT1) + tumor-associated neutrophils (TANs) is a distinguishing feature of PCC patients. Upregulation of IFIT1 + TANs promote migration and invasion of gastric cancer (GC) cell lines (MKN45 and MKN74) and stimulates the growth of cell-derived xenograft models. Besides, by promoting macrophage secreted phosphoprotein 1 (SPP1) expression and facilitating cancer-associated fibroblast and endothelial cell recruitment and activation through TANs, IFIT1 promotes a mesenchymal phenotype, which is associated with a poor prognosis. Importantly, compared to non-PCC (NPCC), PCC tumors is more immunosuppressive. Mechanistically, IFIT1 can be stimulated by IFN-γ and contributes to the expression of Programmed Cell Death 1 Ligand (PDL1) in TANs. We demonstrated in mouse models that IFIT1 + PDL1 + TANs can induce acquired resistance to anti-PD-1 immunotherapy, which may be responsible for the difficulty of PCC patients to benefit from immunotherapy. This work highlights the role of IFIT1 + TANs in mediating the remodeling of the tumor immune microenvironment and immunotherapeutic resistance and introduces IFIT1 + TANs as a promising target for precision therapy of PCC.

Identification and characterization of intermediate states in mammalian neural crest cell epithelial to mesenchymal transition and delamination.

Epithelial to mesenchymal transition (EMT) is a cellular process that converts epithelial cells to mesenchymal cells with migratory potential in developmental and pathological processes. Although originally considered a binary event, EMT in cancer progression involves intermediate states between a fully epithelial and a fully mesenchymal phenotype, which are characterized by distinct combinations of epithelial and mesenchymal markers. This phenomenon has been termed epithelial to mesenchymal plasticity (EMP), however, the intermediate states remain poorly described and it's unclear whether they exist during developmental EMT. Neural crest cells (NCC) are an embryonic progenitor cell population that gives rise to numerous cell types and tissues in vertebrates, and their formation and delamination is a classic example of developmental EMT. However, whether intermediate states also exist during NCC EMT and delamination remains unknown. Through single-cell RNA sequencing of mouse embryos, we identified intermediate NCC states based on their transcriptional signature and then spatially defined their locations in situ in the dorsolateral neuroepithelium. Our results illustrate the importance of cell cycle regulation and functional role for the intermediate stage marker Dlc1 in facilitating mammalian cranial NCC delamination and may provide new insights into mechanisms regulating pathological EMP.