Cell Epithelial-mesenchymal Transition Research Articles

NAT10 promotes ovarian cancer cell migration, invasion, and stemness via N4-acetylcytidine modification of CAPRIN1

Ovarian cancer (OC) is the most lethal gynecological tumor. N4-acetylcytidine (ac4C) modification, catalyzed by the acetyltransferase NAT10, is involved in the occurrence and development of cancers. This study aimed to investigate the role of NAT10 in OC and the underlying molecular mechanisms. The expression of NAT10 and CAPRIN1 in OC cells lines were measured using quantitative real-time polymerase chain reaction and immunoblotting. Biological behaviors of OC cells were evaluated using EdU, Transwell, sphere formation, and immunoblotting assays. The molecular mechanism of NAT10 function was analyzed using bioinformatics, ac4C- RNA immunoprecipitation, and actinomycin D treatment assay. The effect of NAT10 on OC progression in vivo was evaluated using xenograft tumor model. The results indicated that NAT10 and CAPRIN1 were highly expressed in OC cells. NAT10 knockdown suppressed OC cell proliferation, migration, invasiveness, stemness, and epithelial-mesenchymal transition in vitro, and impeded tumor growth in vivo. Additionally, CAPRIN1 expression was found to be positively related to NAT10 expression in OC. Silencing of NAT10 inhibited ac4C levels of CAPRIN1 and reduced its RNA stability. Moreover, overexpression of CAPRIN1 reversed the suppression of migration, invasion, and stemness caused by NAT10 knockdown, while knockdown of CAPRIN1 alone inhibited these malignant behaviors of OC cells. In conclusion, NAT10 promotes OC progression by promoting cellular migration, invasion, and stemness via upregulating CAPRIN1 expression. Mechanistically, NAT10 stabilizes CAPRIN1 by promoting its ac4C modification. These findings suggest that NAT10 may be a promising therapy target for OC.Graphical abstract

3,5,6,7,8,3’,4’- Heptamethoxyflavonoid inhibits TGF-β1-induced epithelial–mesenchymal transition by regulating oxidative stress and autophagy through MEK/ERK/PI3K/AKT/mTOR signaling pathway

Epithelial–mesenchymal transition (EMT) is a crucial pathological process in the pathogenesis of fibrosis. 3,5,6,7,8,3′,4′-hepmethoxyflavone (HMF), the main active ingredient extracted from the Chinese herb Breynia fruticosa (L.) Hook. f., has been shown to have beneficial effects on regulating apoptosis and inhibiting collagen deposition. However, it remains unclear whether and how HMF alleviates transforming growth factor-β1 (TGF-β1)-induced EMT. The objective of this study was to investigate the impact of HMF on TGF-β1-induced EMT in human alveolar Type II epithelial cells (A549) and its underlying mechanism. In vitro culture of TGF-β1-induced EMT in A549 cells revealed that HMF reduced cell viability and migration, inhibited collagen deposition, decreased expression levels of mesenchymal cell markers and fibrosis markers α-SMA, MMP2, TIMP1, β-catenin, and Snail. Meanwhile, the expression level of E-cadherin increased as an epithelial cell marker. Additionally, we discussed the effects of HMF on oxidative stress and autophagy. Various experiments confirmed that HMF regulated the expression levels of Nrf2, keap-1, HO-1, ROS, MDA, SOD, GSH, and played a role in reducing oxidative stress. At the same time, HMF significantly activated autophagy by increasing expressions of Beclin-1 and LC3B as well as enhancing autophagosome content. The addition 3-MA, an autophagy inhibitor attenuated these beneficial effects. Furthermore, HMF significantly inhibited phosphorylation levels of MEK, ERK, PI3K, AKT, and mTOR through various pathways. In conclusion, HMF effectively inhibits TGF-β1-induced EMT in A549 cells by targeting the MEK/ERK/PI3K/AKT/mTOR signaling pathway. Moreover, it exhibits a close correlation with the suppression of oxidative stress and induction of autophagy.

Endoplasmic reticulum stress-related CLIP4 plays a procarcinogenic role in hepatocellular carcinoma: an integrated analysis

ObjectiveTo explore the potential of endoplasmic reticulum stress (ERS)-associated protein CLIP4 as a biomarker for hepatocellular carcinoma (HCC) and the underlying mechanism.MethodsTCGA public database and a tissue microarray were used to investigate the molecular characteristics of CLIP4 and its association with disease. TCGA-LIHC dataset was used for single-gene differential expression analysis, single-gene correlation analysis, functional enrichment analysis, immune infiltration analysis, and DNA methylation analysis. RNA-seq, immunohistochemistry, western blotting, and RT-qPCR were used to verify the effect of ERS on CLIP4 expression. Public databases and miRNA-seq data were used to explore the TF-miRNA-CLIP4 regulatory network. CCK-8, colony formation, EdU staining, wound-healing, Transwell, western blotting and RT-qPCR were used to detect the effects of CLIP4 on the proliferation, migration and epithelial-mesenchymal transition (EMT) of HCC cells.ResultsAnalysis of TCGA datasets and tissue microarrays demonstrated that elevated CLIP4 expression was associated with poor prognosis in HCC. Enrichment analysis revealed that CLIP4 is involved in the immune response, cell adhesion, and EMT. There was a positive correlation between CLIP4 expression and the infiltration of the majority of immune cells, immunomodulators, and chemokines. Furthermore, the DNA methylation pattern of CLIP4 was found to have significant prognostic value. ERS was found to significantly upregulate CLIP4 expression. In addition, the ERS-RELA-miR-222-5p-CLIP4 transcriptional network was constructed to clarify the role of CLIP4. Cell function experiments confirmed that it promotes the proliferation, migration, and EMT of HCC cells.ConclusionsCLIP4 is a potential immune-related oncogenic molecule in HCC. ERS regulates the expression of CLIP4, and CLIP4 promotes the proliferation, migration, and EMT of HCC cells.

The FUS/COL11A1/ZEB1 Axis Contributes to the Development of Oral Squamous Cell Carcinoma.

Oral squamous cell carcinoma (OSCC) is a common type of head and neck cancer with high metastasis rate and poor prognosis. This study aimed to explore the role of collagen type XI alpha 1 (COL11A1) and its detailed mechanisms in OSCC progression. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) analysis was used to measure the level of COL11A1, fused in sarcoma/translocated in liposarcoma (FUS) and zinc finger E-box binding homeobox1 (ZEB1). The protein expression of COL11A1, E-cadherin, N-cadherin, FUS, and ZEB1 were detected using Western blot. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and 5-Ethynyl-2'-deoxyuridine (EdU) staining assays were conducted to measure cell proliferation. Cell migration and invasion were evaluated via wound-healing assay and transwell migration and invasion assays, respectively. Moreover, epithelial mesenchymal transition (EMT) was determined via detecting the expression of EMT-related proteins, including E-cadherin and N-cadherin. RNA immunoprecipitation assay was conducted to evaluate the relationship between ZEB1 and COL11A1. Invivo animal experiments were carried out to explore the role of COL11A1 in OSCC mice, and immunohistochemistry assay was performed to detect the level of ZEB1 and EMT-related proteins in tumor tissues from mice. COL11A1 was augmented in OSCC tissues and cells. COL11A1 knockdown significantly inhibited cell proliferation, migration, invasion, and EMT in OSCC cells. FUS was increased in OSCC tissues. FUS stabilized the mRNA level of COL11A1 and positively regulated the protein expression of COL11A1. ZEB1 was highly expressed in OSCC tissues, and COL11A1directly targeted ZEB1. The inhibition effects of COL11A1 knockdown on cell proliferation, invasion, migration, and EMT were reversed by ZEB1 overexpression in OSCC cells. Additionally, COL11A1 depletion markedly repressed tumor growth through decreasing ZEB1 expression invivo. FUS stabilized COL11A1 to promote the malignant progression of OSCC via regulating ZEB1 expression.

GULP1 as a Novel Diagnostic and Predictive Biomarker in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is characterized by high recurrence and mortality, necessitating the identification of reliable biomarkers. In this study, we aimed to identify the predictive gene signatures for HCC recurrence and evaluate the efficiency of GULP PTB domain-containing engulfment adaptor 1 (GULP1) as a predictive and diagnostic marker and therapeutic target for HCC. We analyzed genomic datasets from The Cancer Genome Atlas and Gene Expression Omnibus databases via least absolute shrinkage and selection operator Cox regression and 10-fold cross-validation, leading to the development of a 15-gene risk score model, which was validated using three independent datasets. Serum GULP1 and α-fetoprotein levels were assessed to determine the diagnostic accuracy of the model. Using clinical cohorts and patient sera, GULP1 roles were examined, and functional assays in vitro and in vivo were used to evaluate its effects on cell growth, epithelial-mesenchymal transition (EMT), ADP-ribosylation factor 6 activation, and β-catenin signaling. Our newly developed risk-score model accurately predicted recurrent HCC in all datasets. Among the 15 genes in the risk score model, GULP1 was overexpressed in patients with HCC and independently predicted HCC recurrence. Its expression modulation influenced cell growth and EMT, with observed effects on ADP-ribosylation factor 6 activation and β-catenin signaling pathways. GULP1 is a crucial biomarker for HCC, serving as a non-invasive diagnostic and predictive tool. It also plays key roles in HCC progression. Our findings highlight the potential use of GULP1 in treatment strategies targeting EMT and HCC recurrence to improve the personalized care and patient outcomes.

KDM1A Acts as an Oncogene and Facilitates the Epithelial Mesenchymal Transition Process in Gastric Cancer.

This study is performed to research the biological role of KDM1A in the epithelial mesenchymal transition (EMT) of gastric cancer and investigate the mechanism involved. The KDM1A, Vimentin and E-cadherin levels were studied, as well as the correlation among them in gastric cancer samples. Gastric cancer cells were transfected with KDM1A overexpression and knockdown, and the cellular infiltration, motility, morphology and F-actin expression were subsequently identified. For the protein level assessment of EMT, the western blot analysis combined with immunofluorescence was employed. The effect of KDM1A on TGF-β/Notch signaling was also detected. KDM1A was overexpressed in gastric cancer tumor tissues. In the clinical gastric carcinoma samples, the level of KDM1A was linked negatively to the expression of E-cadherin, while positively to the expression of Vimentin. Among the gastric carcinoma population, the expression of KDM1A was linked to the lymph node metastasis, TNM stage and tumor differentiation. The KDM1A downregulation prohibited the cellular motility, infiltration and F-actin expression, and suppressed EMT process. KDM1A overexpression exhibited promoting effect on EMT in gastric cancer cells. KDM1A regulated TGF-β/Notch signaling to affect EMT in gastric cancer cells. KDM1A acts as an oncogene and facilitates the epithelial mesenchymal transition process by regulating TGF-β/Notch signal pathway in gastric cancer cells.

SMYD3 plays a pivotal role in mediating the epithelial-mesenchymal transition process in breast cancer.

In previous reports, we highlighted the significant involvement of SMYD3, a histone methyltransferase (HMT), in various aspects of cancer progression, including cell adhesion, migration, and invasion. In this study, we delved deeper into understanding the relationship between SMYD3 and epithelial-mesenchymal transition (EMT) both in cell lines and clinical samples. Our investigation uncovered a notable correlation between heightened SMYD3 expression and the presence of EMT markers in human breast cancer tissues. We found that the induction of SMYD3 expression is facilitated by transforming growth factor beta 1 (TGF-β1), which achieves this by suppressing miR-124, an inhibitor that targets SMYD3, through alterations in DNA methylation. Conversely, our experiments demonstrated that reducing SMYD3 levels through RNA interference impeded TGF-β1-induced EMT in breast cancer cells. Furthermore, our results revealed that SMYD3 alone has the capability to modulate the expression of markers associated with EMT. An intriguing aspect of our study is the revelation that SMYD3 influences the activation of vimentin by binding to its response elements within the core promoter region. Notably, this effect is independent of SMYD3's histone methyltransferase activity. These findings collectively underscore the pivotal role of SMYD3 in driving EMT, both in cell lines and primary cancer tissues, particularly emphasizing its significance in TGF-β1-induced EMT in breast cancer.

Hypoxia enhances IL-8 signaling through inhibiting miR-128-3p expression in glioblastomas

Glioblastoma multiforme (GBM) is an aggressive type of brain tumor known for its hypoxic microenvironment. Understanding the dysregulated mechanisms in hypoxic GBM is crucial for its effective treatment. Through data mining of The Cancer Genome Atlas (TCGA) with hypoxia enrichment scores and in vitro experiments, miR-128-3p was negatively correlated with hypoxia signaling and the epithelial-mesenchymal transition (EMT). Additionally, lower miR-128-3p levels existed in hypoxic GBM, leading to desensitizing temozolomide (TMZ)'s efficacy, a first-line therapeutic drug for GBM. Overexpressing miR-128-3p enhanced both the in vitro and in vivo sensitivity of hypoxic gliomas to TMZ treatment. Mechanistically, HIF-1α suppressed miR-128-3p expression in hypoxic GBM. Through establishing miR-128-3p-mediated transcriptomic profiles and data mining, interleukin (IL)-8 was selected. IL-8 respectively showed positive and negative correlations with hypoxia and miR-128-3p, and was associated with poor TMZ therapeutic results in GBM. Elevated miR-128-3p, which targets both the 3′-untranslated region (UTR) and 5′UTR of IL-8, resulted in suppression of IL-8 expression. Moreover, IL-8 was validated to be involved in HIF-1α/miR-128-3p-regulated TMZ sensitivity and the EMT in hypoxic GBM cells. Collectively, the HIF-1α/miR-128-3p/IL-8 signaling pathway plays a critical role in promoting the progression of hypoxic GBM. Targeting this signaling pathway holds promise as a potential therapeutic strategy.

The role and molecular mechanism of GDF5 in regulating cell migration and angiogenesis via the Hippo-YAP signaling pathway in colorectal cancer.

292 Background: Distant metastasis is a leading cause of poor prognosis and mortality in colorectal cancer (CRC) patients, with angiogenesis playing a crucial role in tumor progression. The TGF-β superfamily has been shown to significantly influence tumor angiogenesis, yet the biological function and molecular mechanism of Growth Differentiation Factor 5 (GDF5), a member of this superfamily, remain unexplored in CRC. Given that the efficacy of current anti-angiogenic therapies is limited to a subset of patients, identifying new therapeutic targets involved in tumor angiogenesis is of great clinical importance. GDF5, with its potential role in angiogenesis regulation, presents a promising candidate to bridge this therapeutic gap. Methods: In this study, we analyzed GDF5 expression in CRC tissue samples using both the TCGA dataset and samples from our center, revealing significantly lower GDF5 expression in tumors compared to adjacent normal tissues. Moreover, GDF5 downregulation correlated with lymph node metastasis and disease recurrence. Functional assays in vitro demonstrated that GDF5 overexpression reduced tumor cell migration and invasion, while its knockdown produced the opposite effect. Co-culture assays showed that GDF5 knockdown significantly enhanced endothelial cell migration and angiogenesis, whereas overexpression diminished these effects. Additionally, F-actin cytoskeletal staining indicated that GDF5 overexpression led to cytoskeletal remodeling in endothelial cells. Mechanistically, we found that GDF5 overexpression in tumor cells downregulated the epithelial-mesenchymal transition (EMT) marker Slug and upregulated E-cadherin, while downregulating N-cadherin. In endothelial cells, GDF5 inhibited the Hippo-YAP signaling pathway, increasing YAP phosphorylation and reducing nuclear YAP levels. Conversely, GDF5 knockdown suppressed YAP phosphorylation and promoted its nuclear translocation. Results: Our findings demonstrate that GDF5 functions as a tumor suppressor in CRC by regulating cell migration and angiogenesis. This study reveals that GDF5 exerts its anti-cancer effects by modulating both EMT in tumor cells and cytoskeletal changes in endothelial cells, leading to the inhibition of the Hippo-YAP signaling pathway and reduced angiogenic potential. These results identify GDF5 as a promising new target for anti-angiogenic therapy in CRC, offering potential avenues for improving treatment strategies in patients resistant to current therapies. Conclusions: GDF5 acts as a tumor suppressor in colorectal cancer by inhibiting cell migration and angiogenesis through modulation of the Hippo-YAP signaling pathway, making it a promising therapeutic target for anti-angiogenic treatment in patients resistant to current therapies.

Hypoxia-induced CTCF mediates alternative splicing via coupling chromatin looping and RNA Pol II pause to promote EMT in breast cancer.

Hypoxia influences the epithelial-mesenchymal transition (EMT) through the remodeling of the chromatin structure, epigenetics, and alternative splicing. Hypoxia drives CCCTC-binding factor (CTCF) induction through hypoxia-inducible factor 1-alpha (HIF1α), which promotes EMT, although the underlying mechanisms remain unclear. We find that hypoxia significantly increases CTCF occupancy at various EMT-related genes. We present a CTCF-mediated intricate mechanism promoting EMT wherein CTCF binding at the collagen type V alpha 1 chain (COL5A1) promoter is crucial for COL5A1 upregulation under hypoxia. Additionally, hypoxia drives exon64A inclusion in a mutually exclusive alternative splicing event of COL5A1exon64 (exon64A/64B). Notably, CTCF mediates COL5A1 promoter-alternatively spliced exon upstream looping that regulates DNA demethylation at distal exon64A. This further regulates the CTCF-mediated RNA polymerase II pause at COL5A1exon64A, leading to its inclusion in promoting the EMT under hypoxia. Genome-wide study indicates the association of gained CTCF occupancy with the alternative splicing of many cancer-related genes, similar to the proposed model. Specifically, disrupting the HIF1α-CTCF-COL5A1exon64A axis through the dCas9-DNMT3A system alleviates the EMT in hypoxic cancer cells and may represent a novel therapeutic target in breast cancer.

PER3 suppresses tumor metastasis of oral squamous cell carcinoma by promoting HIF-1α degradation.

The low expression of period circadian regulator 3 (PER3) in head and neck squamous cell carcinoma is closely correlated with tumor size and invasion depth. Hypoxia-inducible factor 1 subunit alpha (HIF-1α) regulates epithelial-mesenchymal transition (EMT) transcription factors, activates EMT, and promotes tumor metastasis. Here, we investigated the role and molecular mechanism of PER3 in regulating HIF-1α and metastasis in oral squamous cell carcinoma (OSCC) by using bioinformatics analyses and in vitro and in vivo experiments. PER3 expression was decreased in OSCC, and PER3 expression was significantly negatively correlated with T stage, N stage, clinical classification, and survival time. PER3 overexpression inhibited, while PER3 knockdown prompted EMT and metastasis of OSCC cells. HIF-1α reversed the effects of alterations in PER3 expression on OSCC metastasis. Mechanistically, PER3 bound to HIF-1α via the Per-ARNT-Sim 1 domain and promoted its ubiquitination degradation. Hypermethylation at CpG site cg12258811 of PER3 promoter inhibited PER3 expression and prognosis of OSCC. Decitabine combined with LW6 upregulated PER3, downregulated HIF-1α, and inhibited lymph node metastasis of OSCC in nude mice. Our findings reveal the role and mechanism of HIF-1α regulation by PER3 and support the potential clinical application of targeting PER3 in treating OSCC metastasis.

Anticancer and therapeutic efficacy of XPO1 inhibition in pancreatic ductal adenocarcinoma through DNA damage and modulation of miR-193b/KRAS/LAMC2/ERK/AKT signaling cascade.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and grave malignancies with confined and ineffective therapeutic options. XPO1 is a critical regulator of nuclear export and activation of tumor suppressor proteins. The present study evaluated the therapeutic potential and molecular mechanisms of XPO1 inhibition against PDAC. Firstly, we observed significant overexpression of XPO1 transcript in 179 PDAC patients than 171 normal pancreatic tissues in TCGA transcriptomic dataset. Higher XPO1 transcript levels displayed worse overall and disease-free survival. Further, we confirmed significant upregulation of XPO1 in a panel of PDAC cells. Eltanexor treatment resulted in significant inhibition of cell viability, clonogenic growth, migration, and epithelial-mesenchymal transition (EMT), along with the induction of cell cycle arrest. Mechanistically, eltanexor modulated the expression of key proteins including p21, p27, p53, cyclin B1, cyclin D1, c-Myc, N-cadherin, vimentin, E-cadherin associated with the cell viability, growth, cell cycle and EMT. Additionally, the eltanexor treatment resulted in marked increase in expression of γH2AX, and cleaved PARP, cleaved caspase-9 leading to induction of DNA damage and apoptosis of PDAC cells, respectively. Moreover, eltanexor treatment regulated the expression of key non-coding RNAs including miR193b, DINO, MALAT-1, H19, and SOX21-AS1 linked with tumorigenesis. Our results revealed a correlation among miR193b/KRAS/LAMC2, XPO1/KRAS, and LAMC2/KRAS. The findings also revealed that eltanexor treatment rescued the expression of miR193b which acts as a sponge for LAMC2 and KRAS resulting in the suppression of AKT/ERK downstream signaling cascade in PDAC. Interestingly, the combination of eltanexor with gemcitabine showed significant anticancer activity in PDAC cells. Altogether, our findings revealed the crucial role of XPO1 in modulating the expression of oncogenic proteins, ncRNAs, and DNA damage during PDAC progression as well as identified novel therapeutic miR-193b/KRAS/LAMC2/ERK/AKT axis.

Expression pattern and prognostic significance of aldehyde dehydrogenase 2 in lung adenocarcinoma as a potential predictor of immunotherapy efficacy.

The incidence of alcohol-associated cancers is higher within Asian populations having an increased prevalence of an inactivating mutation in aldehyde dehydrogenase 2 (ALDH2), a mitochondrial enzyme required for the clearance of acetaldehyde, a cytotoxic metabolite of ethanol. The role of alcohol consumption in promoting lung cancer is controversial, and little attention has been paid to the association between alcohol drinking and pulmonary ALDH2 expression. We performed a comprehensive bioinformatic analysis of multi-omics data available in public databases to elucidate the role of ALDH2 in lung adenocarcinoma (LUAD). Transcriptional and proteomic data indicate a substantial pulmonary expression of ALDH2, which is functional for the metabolism of alcohol diffused from the bronchial circulation. ALDH2 expression is higher in healthy lung tissue than in LUAD and inhibits cell cycle, apoptosis, and epithelial-mesenchymal transition pathways. Moreover, low ALDH2 mRNA levels predict poor prognosis and low overall survival in LUAD patients. Interestingly, ALDH2 expression correlates with immune infiltration in LUAD. A better understanding of the role of ALDH2 in lung tumor progression and immune infiltration might support its potential use as a prognostic marker and therapeutic target for improving immunotherapeutic response.

Clinical and molecular characterization of FAP and SPP1 in colorectal cancer (CRC), CALGB (Alliance)/SWOG 80405 and real-world data.

276 Background: FAP and SPP1 contribute to immune modulation in the CRC tumor microenvironment (TME). FAP is expressed by cancer associated fibroblasts, aiding in tissue remodeling and tumor invasion. SPP1 is an integrin-binding protein expressed by tumor associated macrophages that promotes tumor growth, adhesion, and metastasis. We present a clinical and molecular characterization of FAP and SPP1 in CRC. Methods: We analyzed 24,257 CRC samples tested at Caris Life Sciences (Phoenix, AZ) with WTS (Illumina NovaSeq), NextGen DNA sequencing (NextSeq), and PD-L1 expression (SP142, positive ≥ 2+, 5%). RNA deconvolution analysis estimated cell infiltration in the TME. Data from the phase 3 CALGB/SWOG 80405 trial (NCT00265850) on 433 metastatic CRC patients treated with bevacizumab (Bev, n = 226) or cetuximab (Cet, n = 207) in combination with first-line chemotherapy were also evaluated. RNA isolated from FFPE tumor samples were sequenced with HiSeq 2500 (Illumina). Overall survival (OS) and progression-free survival (PFS) were compared using Cox regression in categorical gene expression tertiles (high (T3), medium (T2), and low (T1)) for FAP and SPP1 . Results: FAP -T3 and SPP1 -T3 had increased PD-L1 positivity (q<0.05), while SPP1 -T3 also demonstrated increased MSI-H (8.4% vs 5.3%) and TMB-H(>10 mt/mb, 15.0% vs 10.1%) status relative SPP1 -T1 (all q<0.001). T3 of both genes of correlated with increased M1/M2 macrophages, NK cells and T cell inflamed score (q<0.001); dendritic cells and neutrophils were increased in SPP1 -T3 and FAP -T1 tumors (q<0.05). FAP -T3 and SPP1 -T3 had increased pathway activation of epithelial-mesenchymal transition (EMT), inflammatory response, TNF-a signaling, angiogenesis and KRAS signaling (all q < .005). In Caris cohorts, FAP -T3 demonstrated worse OS in Cet/panitumumab treated CRC (T3: 23.6 vs T1: 21.0 months [mo], P = .005; HR 0.85, 95% CI [0.77-0.95]), but SPP1 did not correlate with OS. In 80405, FAP -T3 showed shorter PFS (T3: 9.5 vs T2: 11.5 vs T1: 12.6 mo, T3 vs T1 (reference) adjusted HR 1.27 [1.07-1.51]) and OS (25.2 vs 29.4 vs 35.5 mo, adjusted HR 1.31 [1.10-1.57]). Similarly, SPP1 -T3 demonstrated worse PFS (9.0 vs 12.7 vs 14.0 mo, adjusted HR 1.29 [1.12-1.48]) and OS (20.9 vs 34.0 vs 36.3 mo, HR 1.24 [1.07-1.44], P < 0.001). Treatment interaction tests noted FAP -T1 CRC benefited from Cet over Bev with respect to PFS ( P = 0.003) and OS ( P = 0.044), while SPP1-T1 tumors demonstrated a PFS benefit with Cet ( P = 0.009). Conclusions: Our results indicate that increased FAP and SPP1 expression is associated with immune cell infiltration, EMT, and inflammatory signaling. Additionally, their expression may be prognostic and predictive of targeted therapy. These data support the evaluation of FAP and SPP1 as predictive markers and therapeutic targets in CRC.

Hypoxic stress promotes astrocyte infiltration-like growth via HIF-1α/GDNF/LOXL2 axis.

Elevated levels of glial cell line-derived neurotrophic factor (GDNF) are implicated in the transformation of astrocytes into astrogliomas, but the underlying mechanisms are not fully understood. In this study, we found that hypoxia led to a significant increase in GDNF expression in primary rat astrocytes from various brain regions, including the cortex, hippocampus, and corpus callosum. This was accompanied by the activation of astrocytes, particularly those of the A2 subtype, and a concurrent increase in hypoxia-inducible factor 1-alpha (HIF-1α) expression. The elevated levels of HIF-1α enhanced its binding to the GDNF promoter, resulting in increased GDNF expression. Interestingly, this process formed a positive feedback loop, as elevated GDNF further activated HIF-1α in primary rat and human astrocytes. Furthermore, lysyl oxidase-like protein 2 (LOXL2), a novel downstream oncogene of GDNF, showed a significant increase following hypoxia treatment and exhibited a positive correlation with GDNF expression. Inhibiting GDNF signaling effectively suppressed this expression. Hypoxia-induced GDNF also increased the phosphorylation of ERK, P38, and CREB through the classical GDNF receptors, GFRα1 and RET. This led to increased binding of phosphorylated CREB to the LOXL2 promoter, resulting in enhanced LOXL2 expression. Consequently, rat astrocytes under hypoxic stress exhibited increased cell viability, migration, and epithelial-mesenchymal transition, which were mitigated by inhibiting GDNF signaling or silencing LOXL2. This phenomenon was also observed in C6 cells. Our findings suggest that hypoxia induces astrocyte activation and upregulates LOXL2 expression through the HIF-1α/GDNF/P-CREB signaling axis, facilitating the infiltration-like growth of astrocytes and the infiltrative growth of C6 astroglioma cells.

Polymer-based nanodrugs enhance sonodynamic therapy through epigenetic reprogramming of the immunosuppressive tumor microenvironment.

While sonodynamic therapy (SDT) has shown promise in treating triple-negative breast cancer (TNBC) due to its non-invasive nature, deep tissue penetration, and induction of immunogenic cell death (ICD), its efficacy remains limited by the complex immunosuppressive tumor microenvironment (TME). In this study, we developed tumor microenvironment-responsive nanoparticles (GdNPs) to enhance SDT effectiveness through epigenetic reprogramming of the TME by encapsulating the sonosensitizer chlorin e6 (Ce6) and the histone deacetylase 6 (HDAC6) inhibitor Ricolinostat (Ric) (GdNPs/Ce6-Ric). GdNPs/Ce6-Ric effectively accumulate at tumor sites via the enhanced permeability and retention (EPR) effect and release Ce6 and Ric in response to the acidic TME. Upon ultrasound stimulation, GdNPs/Ce6-Ric induce cancer cell apoptosis and trigger ICD by generating reactive oxygen species (ROS), which activate cytotoxic T cells and promote tumor cell elimination. Notably, the epigenetic modulation by Ric within the immunosuppressive TME increased the proportion of natural killer (NK) cells and cytotoxic T cells while decreasing the population of immunosuppressive regulatory T (Treg) cells. This modulation synergistically enhanced the anti-tumor effects of SDT by downregulating the HDAC6/p-STAT3/PD-L1 pathway. Furthermore, GdNPs/Ce6-Ric minimized lung metastases by not only improving systemic immune responses but also inhibiting TGFβ-induced epithelial-mesenchymal transition (EMT) of tumor cells through the blockade of α-tubulin deacetylation. Thus, GdNPs/Ce6-Ric-based epigenetic modulation of the immunosuppressive TME offers a promising approach to enhance the efficacy of SDT in treating TNBC.

Estimation of miRNA 208 Effects in Inducing Epithelial-Mesenchymal Transition by Targeting CDH2 in Breast Cancer Patients of Iraqi Population

Breast cancer is the most frequently diagnosed disease in women and remains a leading cause of cancer-related deaths worldwide. The small non-coding RNA molecule, MicroRNA-208 (miR-208), plays a crucial role in the development and advancement of numerous malignancies, including breast cancer. This study aimed to investigate miR-208 potential as an oncogene in breast cancer by examining its impact on cell motility and epithelial-mesenchymal transition (EMT). Tissue samples from 25 Iraqi women with malignant breast cancer and 25 with benign breast cancer were analysed. Total RNA was extracted from the samples and real-time reverse transcription-polymerase chain reaction (RT-PCR) was performed using SYBR Green technology to measure miR-208 expression levels. Tissue samples from both benign and malignant tumours were analysed to determine U6 gene expression. The analysis showed that the typical Ct values for benign and malignant samples were 29.83 and 29.67 respectively. No significant difference in U6 expression levels was detected between benign and malignant tumours. However, upon comparing the initial malignant and the benign tumours samples, miRNA-208 ΔΔCT values were found to be substantially lower (-7.105 and -0.781 respectively) with a p-value of less than 0.05. The study found that malignant tumours had a significantly higher miRNA-208 folding value (2.896±) compared to benign tumours which had substantially lower miRNA-208 folding values (1.591 ±0.37). The study also found that the differences were statistically significant (p-value = 1.292*). In conclusion, the study found evidence linking miR-208 to the CDH2 gene, indicating its role in the development and susceptibility to breast cancer in certain Iraqi women.

Phytochemicals in Cancer Therapy: Modulating Cell Cycle, Angiogenesis, and Epithelial-Mesenchymal Transition

Phytochemicals in Cancer Therapy: Modulating Cell Cycle, Angiogenesis, and Epithelial-Mesenchymal Transition

SHP2 promotes the epithelial-mesenchymal transition in triple negative breast cancer cells by regulating β-catenin

PurposeGrowing evidence suggests that the tyrosine phosphatase SHP2 is pivotal for tumor progression. Triple-negative breast cancer (TNBC) is the most lethal subtype of breast cancer, characterized by its high recurrence rate, aggressive metastasis, and resistance to chemotherapy. Understanding the mechanisms of tumorigenesis and the underlying molecular pathways in TNBC could aid in identifying new therapeutic targets.MethodsIn this study, we conducted bioinformatics analysis of the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases to examine PTPN11 (encoding SHP2) expression levels and perform survival analysis in TNBC. Additionally, we analyzed SHP2 levels in four TNBC cell lines and a normal breast epithelial cell line using Western blot. Furthermore, we knocked down SHP2 expression via RNA interference in three TNBC cell lines. To assess the impact of SHP2 on invasion and migration, we conducted transwell assays and wound healing experiments. An in vivo experiment utilizing a mouse xenograft model was also performed to evaluate tumor metastasis. Moreover, we detected the expression levels of epithelial-mesenchymal transition (EMT) biomarkers and investigated the mechanism between SHP2 and β-catenin using Western blot and immunofluorescence experiments.ResultsWe found that high SHP2 expression was associated with a poor prognosis in patients with TNBC. The migratory and invasive abilities of TNBC cells in vitro, as well as the metastatic potential of TNBC in mouse xenograft models, were reduced after SHP2 depletion. Downregulation of SHP2 also decreased the expression of mesenchymal markers but induced upregulation of the epithelial marker E-cadherin. Additionally, SHP2 promoted β-catenin stability by inhibiting its degradation via the proteasome. Furthermore, c-Myc expression and GSK3β and AKT phosphorylation, which are involved in β-catenin signaling, were decreased in SHP2-depleted TNBC cells. ConclusionOur study demonstrates that SHP2 is involved in migration, invasion, and EMT in TNBC cells by modulating β-catenin. Manipulating SHP2 expression or its target protein β-catenin may offer a novel approach to TNBC therapy.

Perilipin2-dependent lipid droplets accumulation promotes metastasis of oral squamous cell carcinoma via epithelial-mesenchymal transition

Emerging evidence shows that lipid metabolic reprogramming plays a vital role in tumor metastasis. The effect and mechanism of fatty acids and lipid droplets (LDs), the core products of lipid metabolism, on the metastasis of oral squamous cell carcinoma (OSCC), need further exploration. In this study, the influence of palmitic acid (PA) and oleic acid (OA) on the migration and invasion ability of OSCC cells was determined by in vitro experiments. Genetic manipulation of PLIN2 was performed to explore its effect on the accumulation of LDs and OSCC metastasis. Possible mechanisms of these biological effects were clarified by detecting the levels of epithelial-mesenchymal transition (EMT) markers and phosphatidylinositol 3-kinase (PI3K) pathway proteins as well as conducting various bioinformatics analyses. The results indicated that PA/OA promoted the migration and invasion of OSCC cells and induced PLIN2-dependent LDs accumulation in vitro. Knockdown of PLIN2 inhibited the LDs accumulation and the migration and invasion of OSCC cells in vitro, while overexpression of PLIN2 enhanced those of OSCC cells in vitro and also promoted the metastasis of OSCC in vivo. Besides, PLIN2 up-regulation activated the PI3K pathway and subsequently enhanced EMT in OSCC cells in vitro. OSCC patients with higher PLIN2 expression possessed poorer prognosis and higher sensitivity to chemotherapy drugs (1S,3 R)-RSL3 and ML-210. In conclusion, PLIN2-dependent LDs accumulation could promote the metastasis of OSCC cells by regulating EMT. PLIN2 might be a potential therapeutic target for OSCC patients, especially those with obesity.