Paeoniae radix overcomes resistance to EGFR-TKIs via aurora B pathway suppression in lung adenocarcinoma.

Targeted therapies using epidermal growth factor receptor (EGFR) inhibitors have markedly improved survival rates and quality of life for patients with EGFR-mutant lung adenocarcinoma (LUAD). Despite these advancements, resistance to EGFR inhibitors remains a significant challenge, limiting the overall effectiveness of the treatment. This study explored the synergistic effects of combining Paeoniae Radix (PR) with first-generation EGFR-tyrosine kinase inhibitors (TKIs), erlotinib and gefitinib, to overcome this resistance. Transcriptomic analysis of EGFR-mutant LUAD cell lines revealed that PR treatment could potentially reverse the gene signatures associated with resistance to EGFR-TKIs, primarily through the suppression of the Aurora B pathway. Experimental validation demonstrated that combining PR with erlotinib and gefitinib enhanced drug responsiveness by inhibiting Aurora kinase activity and inducing apoptosis in LUAD cells. Additionally, gene expression changes confirmed these combined effects, with the suppression of the Aurora B pathway and upregulation of the apoptotic pathway, which was accompanied by increased expression of multiple pro-apoptotic genes. Our findings contribute to the development of natural product-based therapeutic strategies to mitigate drug resistance in LUAD.

Discovery of N-Aryl-N'-[4-(aryloxy)cyclohexyl]squaramide-Based Inhibitors of LXR/SREBP-1c Signaling Pathway Ameliorating Steatotic Liver Disease: Navigating the Role of SIRT6 Activation.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is primarily attributed to the abnormal upregulation of hepatic lipogenesis, which is especially caused by the overactivation of the liver X receptor/sterol regulatory element-binding protein-1c (LXR/SREBP-1c) pathway in hepatocytes. In this study, we report the rational design and synthesis of a novel series of squaramides via bioisosteric replacement, which was evaluated for its inhibitory activity on the LXR/SREBP-1c pathway using dual cell-based assays. Compound 31 was found to significantly downregulate LXR, SREBP-1c, and their target genes associated with lipogenesis. Further investigation revealed that compound 31 may indirectly inhibit the LXR/SREBP-1c pathway by activating the upstream regulator sirtuin 6 (SIRT6). Encouragingly, compound 31 substantially attenuated lipid accumulation in HepG2 cells and in the liver of high-fat-diet-fed mice. These findings suggest that compound 31 holds promise as a candidate for the development of treatments for MASLD and other lipid metabolism-related diseases.

Tumor necrosis factor-inducible gene 6 protein and its derived peptide ameliorate liver fibrosis by repressing CD44 activation in mice with alcohol-related liver disease.

BACKGROUND: Alcohol-related liver disease (ALD) is a major health concern worldwide, but effective therapeutics for ALD are still lacking. Tumor necrosis factor-inducible gene 6 protein (TSG-6), a cytokine released from mesenchymal stem cells, was shown to reduce liver fibrosis and promote successful liver repair in mice with chronically damaged livers. However, the effect of TSG-6 and the mechanism underlying its activity in ALD remain poorly understood. METHODS: To investigate its function in ALD mice with fibrosis, male mice chronically fed an ethanol (EtOH)-containing diet for 9 weeks were treated with TSG-6 (EtOH + TSG-6) or PBS (EtOH + Veh) for an additional 3 weeks. RESULTS: Severe hepatic injury in EtOH-treated mice was markedly decreased in TSG-6-treated mice fed EtOH. The EtOH + TSG-6 group had less fibrosis than the EtOH + Veh group. Activation of cluster of differentiation 44 (CD44) was reported to promote HSC activation. CD44 and nuclear CD44 intracellular domain (ICD), a CD44 activator which were upregulated in activated HSCs and ALD mice were significantly downregulated in TSG-6-exposed mice fed EtOH. TSG-6 interacted directly with the catalytic site of MMP14, a proteolytic enzyme that cleaves CD44, inhibited CD44 cleavage to CD44ICD, and reduced HSC activation and liver fibrosis in ALD mice. In addition, a novel peptide designed to include a region that binds to the catalytic site of MMP14 suppressed CD44 activation and attenuated alcohol-induced liver injury, including fibrosis, in mice. CONCLUSIONS: These results demonstrate that TSG-6 attenuates alcohol-induced liver damage and fibrosis by blocking CD44 cleavage to CD44ICD and suggest that TSG-6 and TSG-6-mimicking peptide could be used as therapeutics for ALD with fibrosis.

Systematic omics analysis identifies CCR6 as a therapeutic target to overcome cancer resistance to EGFR inhibitors.

Epidermal growth factor receptor inhibitors (EGFRi) have exhibited promising clinical outcomes in the treatment of various cancers. However, their widespread application has been limited by low patient eligibility and the emergence of resistance. Leveraging a multi-omics approach (>1000 cancer cell lines), we explored molecular signatures linked to EGFRi responsiveness and found that expression signatures involved in the estrogen response could recapitulate cancer cell dependency on EGFR, a phenomenon not solely attributable to EGFR-activating mutations. By correlating genome-wide function screening data with EGFRi responses, we identified chemokine receptor 6 (CCR6) as a potential druggable target to mitigate EGFRi resistance. In isogenic cell models, pharmacological inhibition of CCR6 effectively reversed acquired EGFRi resistance, disrupting mitochondrial oxidative phosphorylation, a cellular process commonly associated with therapy resistance. Our data-driven strategy unveils drug-response biomarkers and therapeutic targets for resistance, thus potentially expanding EGFRi applicability and efficacy.

SIRT1 ISGylation accelerates tumor progression by unleashing SIRT1 from the inactive state to promote its deacetylase activity.

ISG15 is an interferon-stimulated ubiquitin-like protein (UBL) with multifaceted roles as a posttranslational modifier in ISG15 conjugation (ISGylation). However, the mechanistic consequences of ISGylation in cancer have not been fully elucidated, largely due to a lack of knowledge on the ISG15 target repertoire. Here, we identified SIRT1, a nicotinamide adenine dinucleotide (NAD+)-dependent protein deacetylase, as a new target for ISGylation. SIRT1 ISGylation impairs the association of SIRT1 with its negative regulator, deleted in breast cancer 1 (DBC1), which unleashes SIRT1 from its inactive state and leads to an increase in its deacetylase activity. Importantly, SIRT1 ISGylation promoted lung cancer progression and limited lung cancer cell sensitivity to DNA damage-based therapeutics in vivo and in vitro models. The levels of ISG15 mRNA and protein were significantly higher in lung cancer tissues than in adjacent normal tissues. Accordingly, elevated expression of SIRT1 and ISG15 was associated with poor prognosis in lung cancer patients, a finding that could be translated for lung cancer patient stratification and disease outcome evaluation. Taken together, our findings provide a mechanistic understanding of the regulatory effect of SIRT1 ISGylation on tumor progression and therapeutic efficacy in lung cancer.

Overexpression of Interleukin-8 Promotes the Progression of Fatty Liver to Nonalcoholic Steatohepatitis in Mice.

Nonalcoholic steatohepatitis (NASH) is an advanced stage of fatty liver disease characterized by liver damage, inflammation, and fibrosis. Although neutrophil infiltration is consistently observed in the livers of patients with NASH, the precise role of neutrophil-recruiting chemokines and infiltrating neutrophils in NASH pathogenesis remains poorly understood. Here, we aimed to elucidate the role of neutrophil infiltration in the transition from fatty liver to NASH by examining hepatic overexpression of interleukin-8 (IL8), a major chemokine responsible for neutrophil recruitment in humans. Mice fed a high-fat diet (HFD) for 3 months developed fatty liver without concurrent liver damage, inflammation, and fibrosis. Subsequent infection with an adenovirus overexpressing human IL8 for an additional 2 weeks increased IL8 levels, neutrophil infiltration, and liver injury in mice. Mechanistically, IL8-induced liver injury was associated with the upregulation of components of the NADPH oxidase 2 complex, which participate in neutrophil oxidative burst. IL8-driven neutrophil infiltration promoted macrophage aggregate formation and upregulated the expression of chemokines and inflammatory cytokines. Notably, IL8 overexpression amplified factors associated with fibrosis, including collagen deposition and hepatic stellate cell activation, in HFD-fed mice. Collectively, hepatic overexpression of human IL8 promotes neutrophil infiltration and fatty liver progression to NASH in HFD-fed mice.

FSP1 confers ferroptosis resistance in KEAP1 mutant non-small cell lung carcinoma in NRF2-dependent and -independent manner.

Ferroptosis, a type of cell death induced by lipid peroxidation, has emerged as a novel anti-cancer strategy. Cancer cells frequently acquire resistance to ferroptosis. However, the underlying mechanisms are poorly understood. To address this issue, we conducted a thorough investigation of the genomic and transcriptomic data derived from hundreds of human cancer cell lines and primary tissue samples, with a particular focus on non-small cell lung carcinoma (NSCLC). It was observed that mutations in Kelch-like ECH-associated protein 1 (KEAP1) and subsequent nuclear factor erythroid 2-related factor 2 (NRF2, also known as NFE2L2) activation are strongly associated with ferroptosis resistance in NSCLC. Additionally, AIFM2 gene, which encodes ferroptosis suppressor protein 1 (FSP1), was identified as the gene most significantly correlated with ferroptosis resistance, followed by multiple NRF2 targets. We found that inhibition of NRF2 alone was not sufficient to reduce FSP1 protein levels and promote ferroptosis, whereas FSP1 inhibition effectively sensitized KEAP1-mutant NSCLC cells to ferroptosis. Furthermore, we found that combined inhibition of FSP1 and NRF2 induced ferroptosis more intensely. Our findings imply that FSP1 is a crucial suppressor of ferroptosis whose expression is partially dependent on NRF2 and that synergistically targeting both FSP1 and NRF2 may be a promising strategy for overcoming ferroptosis resistance in cancer.

Integrative Transcriptomic Analysis Reveals Upregulated Apoptotic Signaling in Wound-Healing Pathway in Rat Liver Fibrosis Models.

Liver fibrosis, defined by the aberrant accumulation of extracellular matrix proteins in liver tissue due to chronic inflammation, represents a pressing global health issue. In this study, we investigated the transcriptomic signatures of three independent liver fibrosis models induced by bile duct ligation, carbon tetrachloride, and dimethylnitrosamine (DMN) to unravel the pathological mechanisms underlying hepatic fibrosis. We observed significant changes in gene expression linked to key characteristics of liver fibrosis, with a distinctive correlation to the burn-wound-healing pathway. Building on these transcriptomic insights, we further probed the p53 signaling pathways within the DMN-induced rat liver fibrosis model, utilizing western blot analysis. We observed a pronounced elevation in p53 protein levels and heightened ratios of BAX/BCL2, cleaved/pro-CASPASE-3, and cleaved/full length-PARP in the livers of DMN-exposed rats. Furthermore, we discovered that orally administering oligonol-a polyphenol, derived from lychee, with anti-oxidative properties-effectively countered the overexpressions of pivotal apoptotic genes within these fibrotic models. In conclusion, our findings offer an in-depth understanding of the molecular alterations contributing to liver fibrosis, spotlighting the essential role of the apoptosis pathway tied to the burn-wound-healing process. Most importantly, our research proposes that regulating this pathway, specifically the balance of apoptosis, could serve as a potential therapeutic approach for treating liver fibrosis.

In Silico Discovery of 5'-Modified 7-Deoxy-7-ethynyl-4'-thioadenosine as a HASPIN Inhibitor and Its Synergistic Anticancer Effect with the PLK1 Inhibitor.

Despite genetic perturbations resulting in embryo lethality for most mitotic kinases, loss of the histone H3 mitotic kinase HASPIN reveals no adverse effect in mice models, establishing HASPIN as a promising target for anticancer therapy. However, developing a HASPIN inhibitor from conventional pharmacophores poses a technical challenge as this atypical kinase shares slight similarities with eukaryotic protein kinases. Chemically modifying a cytotoxic 4'-thioadenosine analogue through high genotoxicity yielded several novel nongenotoxic kinase inhibitors. In silico apporoaches utilizing transcriptomic and chemical similarities with known compounds and KINOMEscan profiles unveiled the HASPIN inhibitor LJ4827. LJ4827's specificity and potency as a HASPIN inhibitor were verified through in vitro kinase assay and X-ray crystallography. HASPIN inhibition by LJ4827 reduced histone H3 phosphorylation and impeded Aurora B recruitment in cancer cell centromeres but not in noncancer cells. Through transcriptome analysis of lung cancer patients, PLK1 was determined as a druggable synergistic partner to complement HASPIN inhibition. Chemical or genetic PLK1 perturbation with LJ4827 effectuated pronounced lung cancer cytotoxicity in vitro and in vivo. Therefore, LJ4827 is a novel anticancer therapeutic for selectively impeding cancer mitosis through potent HASPIN inhibition, and simultaneous HASPIN and PLK1 interference is a promising therapeutic strategy for lung cancer.

TPX2 Amplification-Driven Aberrant Mitosis in Culture Adapted Human Embryonic Stem Cells with gain of 20q11.21.

BACKGROUND: Despite highly effective machinery for the maintenance of genome integrity in human embryonic stem cells (hESCs), the frequency of genetic aberrations during in-vitro culture has been a serious issue for future clinical applications. METHOD: By passaging hESCs over a broad range of timepoints (up to 6 years), the isogenic hESC lines with different passage numbers with distinct cellular characteristics, were established. RESULT: We found that mitotic aberrations, such as the delay of mitosis, multipolar centrosomes, and chromosome mis-segregation, were increased in parallel with polyploidy compared to early-passaged hESCs (EP-hESCs) with normal copy number. Through high-resolution genome-wide approaches and transcriptome analysis, we found that culture adapted-hESCs with a minimal amplicon in chromosome 20q11.21 highly expressed TPX2, a key protein for governing spindle assembly and cancer malignancy. Consistent with these findings, the inducible expression of TPX2 in EP-hESCs reproduced aberrant mitotic events, such as the delay of mitotic progression, spindle stabilization, misaligned chromosomes, and polyploidy. CONCLUSION: These studies suggest that the increased transcription of TPX2 in culture adapted hESCs could contribute to an increase in aberrant mitosis due to altered spindle dynamics.

Epigenetic Activation of Tensin 4 Promotes Gastric Cancer Progression.

Gastric cancer (GC) is a complex disease influenced by multiple genetic and epigenetic factors. Chronic inflammation caused by Helicobacter pylori infection and dietary risk factors can result in the accumulation of aberrant DNA methylation in gastric mucosa, which promotes GC development. Tensin 4 (TNS4), a member of the Tensin family of proteins, is localized to focal adhesion sites, which connect the extracellular matrix and cytoskeletal network. We identified upregulation of TNS4 in GC using quantitative reverse transcription PCR with 174 paired samples of GC tumors and adjacent normal tissues. Transcriptional activation of TNS4 occurred even during the early stage of tumor development. TNS4 depletion in GC cell lines that expressed high to moderate levels of TNS4, i.e., SNU-601, KATO III, and MKN74, reduced cell proliferation and migration, whereas ectopic expression of TNS4 in those lines that expressed lower levels of TNS4, i.e., SNU-638, MKN1, and MKN45 increased colony formation and cell migration. The promoter region of TNS4 was hypomethylated in GC cell lines that showed upregulation of TNS4. We also found a significant negative correlation between TNS4 expression and CpG methylation in 250 GC tumors based on The Cancer Genome Atlas (TCGA) data. This study elucidates the epigenetic mechanism of TNS4 activation and functional roles of TNS4 in GC development and progression and suggests a possible approach for future GC treatments.

TPX2 prompts mitotic survival via the induction of BCL2L1 through YAP1 protein stabilization in human embryonic stem cells.

Genetic alterations have been reported for decades in most human embryonic stem cells (hESCs). Survival advantage, a typical trait acquired during long-term in vitro culture, results from the induction of BCL2L1 upon frequent copy number variation (CNV) at locus 20q11.21 and is one of the strongest candidates associated with genetic alterations that occur via escape from mitotic stress. However, the underlying mechanisms for BCL2L1 induction remain unknown. Furthermore, abnormal mitosis and the survival advantage that frequently occur in late passage are associated with the expression of BCL2L1, which is in locus 20q11.21. In this study, we demonstrated that the expression of TPX2, a gene located in 20q11.21, led to BCL2L1 induction and consequent survival traits under mitotic stress in isogenic pairs of hESCs and human induced pluripotent stem cells (iPSCs) with normal and 20q11.21 CNVs. High Aurora A kinase activity by TPX2 stabilized the YAP1 protein to induce YAP1-dependent BCL2L1 expression. A chemical inhibitor of Aurora A kinase and knockdown of YAP/TAZ significantly abrogated the high tolerance to mitotic stress through BCL2L1 suppression. These results suggest that the collective expression of TPX2 and BCL2L1 from CNV at loci 20q11.21 and a consequent increase in YAP1 signaling promote genome instability during long-term in vitro hESC culture.

Systematic transcriptome analysis reveals molecular mechanisms and indications of bupleuri radix.

Pharmacogenomic analysis based on drug transcriptomic signatures is widely used to identify mechanisms of action and pharmacological indications. Despite accumulating reports on the efficacy of medicinal herbs, related transcriptome-level analyses are lacking. The aim of the present study was to elucidate the underlying molecular mechanisms of action of Bupleuri Radix (BR), a widely used herbal medicine, through a systematic transcriptomic analysis. We analyzed the drug-responsive transcriptome profiling of A549 lung cancer cell line after treating them with multiple doses of BR water (W-BR) and ethanol (E-BR) extracts and their phytochemicals. In vitro validation experiments were performed using both A549 and the immortalized human keratinocyte line HaCaT. Pathway enrichment analysis revealed the anti-cancer effects of BR treatment via inhibition of cell proliferation and induction of apoptosis. Enhanced cell adhesion and migration were observed with the W-BR but not with the E-BR. Comparison with a disease signature database validated an indication of the W-BR for skin disorders. Moreover, W-BR treatment showed the wound-healing effect in skin and lung cells. The main active ingredients of BR showed only the anti-cancer effect of the E-BR and not the wound healing effect of the W-BR, suggesting the need for research on minor ingredients of BR.

Activation of the HSP27-AKT axis contributes to gefitinib resistance in non-small cell lung cancer cells independent of EGFR mutations.

PURPOSE: Although epidermal growth factor receptor (EGFR)-activating mutations in non-small cell lung cancer (NSCLC) usually show sensitivity to first-generation EGFR-tyrosine kinase inhibitors (TKIs), most patients relapse because of drug resistance. Heat shock protein 27 (HSP27) has been reported to be involved in the resistance of EGFR-TKIs, although the underlying mechanism is unclear. Here, we explore the mechanisms of HSP27-mediated EGFR TKI resistance and propose novel therapeutic strategies. METHODS: To determine the mechanism of HSP27 associated gefitinib resistance, differences were assessed using gefitinib-sensitive and -resistant NSCLC cell lines. In vivo xenograft experiments were conducted to elucidate the combinatorial effects of J2, a small molecule HSP27 inhibitor, and gefitinib. Analyses of human NSCLC tissues and PDX tissues were also used for comparison of HSP27 and phosphorylated AKT expression. RESULTS: Large-scale cohort analysis of NSCLC cases revealed that HSP27 expression correlated well with the incidence of EGFR mutations and affected patient survival. Increased pAKT and HSP27 was observed in gefitinib-resistant cells compared with gefitinib-sensitive cells. Moreover, increased phosphorylation of HSP27 by gefitinib augmented its protein stability and potentiated its binding activity with pAKT, which resulted in increased gefitinib resistance. However, in gefitinib-sensitive cells, stronger binding activity between EGFR and HSP27 was observed. Moreover, these phenomena occurred regardless of EGFR mutation including secondary mutations, such as T790M. AKT knockdown switched HSP27-pAKT binding to HSP27-EGFR, which promoted gefitinib sensitivity in gefitinib-resistant cells. Functional inhibition of HSP27 yielded sensitization to gefitinib in gefitinib-resistant cells by inhibiting the interaction between HSP27 and pAKT. CONCLUSIONS: Our results indicate that combination of EGFR-TKIs with HSP27 inhibitors may represent a good strategy to overcome resistance to EGFR-TKIs, especially in cancers exhibiting AKT pathway activation.

Identification of a novel anticancer mechanism of Paeoniae Radix extracts based on systematic transcriptome analysis.

Paeoniae Radix (PR) has a great therapeutic value in many clinical applications; however, the presence of various bioactive compounds and its complicated effects on human health makes its precise mechanisms of action unclear. This study investigated the effects of PR at the molecular pathway level by profiling genome-wide gene expression changes following dose-dependent treatment of human lung cancer cells (A549) with PR water extract (WPR), PR ethanol extracts (EPR), as well as their individual components. We found that PR exerts anticancer effects in A549 cells by regulating numerous pathways. Specifically, EPR and two compounds, namely, hederagenin (HG) and oleanolic acid (OA), significantly downregulate the Aurora B pathway. Furthermore, we generated an integrated PR extracts-compounds-target genes network in the Aurora B pathway to understand their interactions. Our findings reinforce that inhibiting Aurora kinase activity is a therapeutic target for treating cancers, providing the potential for novel mechanisms of action for PR and its components against lung cancer.

BAP1 Downregulates NRF2 Target Genes and Exerts Anti-Tumorigenic Effects by Deubiquitinating KEAP1 in Lung Adenocarcinoma.

KELCH-ECH-associated protein 1 (KEAP1) is an adaptor protein of Cullin 3 (CUL3) E3 ubiquitin ligase that targets a redox sensitive transcription factor, NF-E2-related factor 2 (NRF2). BRCA1-associated protein 1 (BAP1) is a tumor suppressor and deubiquitinase whose mutations increase the risk of several types of familial cancers. In the present study, we have identified that BAP1 deubiquitinates KEAP1 by binding to the BTB domain. Lentiviral transduction of BAP1 decreased the expression of NRF2 target genes, suppressed the migration and invasion, and sensitized cisplatin-induced apoptosis in human lung adenocarcinoma (LUAD) A549 cells. Examination of the lung tissues in KrasG12D/+ mice demonstrated that the level of Bap1 and Keap1 mRNAs progressively decreases during lung tumor progression, and it is correlated with NRF2 activation and the inhibition of oxidative stress. Supporting this observation, lentiviral transduction of BAP1 decreased the growth of A549 xenografts in athymic nude mice. Transcriptome analysis of human lung tissues showed that the levels of Bap1 mRNA are significantly higher in normal samples than LUAD samples. Moreover, the expression of Bap1 mRNA is associated with a better survival of LUAD patients. Together, our study demonstrates that KEAP1 deubiquitination by BAP1 is novel tumor suppressive mechanism of LUAD.

Necrostatin-1 Prevents Ferroptosis in a RIPK1- and IDO-Independent Manner in Hepatocellular Carcinoma.

Ferroptosis is caused by the iron-mediated accumulation of lipid peroxidation, which is distinct from apoptosis and necroptosis. Necrostatin-1 inhibits receptor-interacting serine/threonine-protein kinase 1 (RIPK1) to initiate necroptosis; it also inhibits indoleamine 2,3-dioxygenase (IDO) to regulate tumor immunity. However, few studies have examined the off-target effect of necrostatin-1 on the ferroptosis pathway. The present study examined whether necrostatin-1 could interrupt ferroptosis induced by system xc- inhibitors (sulfasalazine and erastin) and a glutathione peroxidase 4 inhibitor (RSL3) in Huh7 and SK-HEP-1 cells. Necrostatin-1 completely prevented decreases in cell viability induced by sulfasalazine and erastin; it partially blunted decreases in cell viability induced by RSL3. Necrostatin-1, ferrostatin-1, and deferoxamine repressed sulfasalazine-provoked membrane permeabilization, as detected by 7-aminoactinomycin D staining and lipid peroxidation measured using a C11-BODIPY probe. However, other RIPK1 inhibitors (necrostatin-1s and GSK2982772) and an IDO inhibitor (1-methyl-D-tryptophan) did not recover the decrease in cell viability induced by sulfasalazine. Necrostatin-1 potentiated sulfasalazine-induced expression of xCT, a catalytic subunit of system xc- in these cells. These results demonstrated that necrostatin-1 blocked ferroptosis through a mechanism independent from RIPK1 and IDO inhibition in Huh7 and SK-HEP-1 cells, indicating that its antioxidant activity should be considered when using necrostatin-1 as a RIPK1 inhibitor.

Crosstalk between YAP and TGFß regulates SERPINE1 expression in mesenchymal lung cancer cells.

Serpin family E member 1 (SERPINE1), a serine proteinase inhibitor, serves as an important regulator of extracellular matrix remodeling. Emerging evidence suggests that SERPINE1 has diverse roles in cancer and is associated with poor prognosis. However, the mechanism via which SERPINE1 is induced in cancer has not been fully determined. In order to examine the molecular mechanism of SERPINE1 expression, the present study took advantage of the isogenic pair of lung cancer cells with epithelial or mesenchymal features. Using genetic perturbation and following biochemical analysis, the present study demonstrated that SERPINE1 expression was upregulated in mesenchymal lung cancer cells and promoted cellular invasiveness. Yes­associated protein (YAP)­dependent SERPINE1 expression was modulated by treatment with a Rho­associated protein kinase inhibitor, Y27632. Moreover, TGFß treatment supported YAP­dependent SERPINE1 expression, and an enhanced TGFß response in mesenchymal lung cancer cells promoted SERPINE1 expression. TGFß­mediated SERPINE1 expression was significantly attenuated by knockdown of YAP or transcriptional co­activator with PDZ­binding motif, suggesting that crosstalk between the TGFß and YAP pathways underlies SERPINE1 expression in mesenchymal cancer cells.

TGFß promotes YAP-dependent AXL induction in mesenchymal-type lung cancer cells.

The acquisition of chemoresistance remains a major cause of cancer mortality due to the limited accessibility of targeted or immune therapies. However, given that severe alterations of molecular features during epithelial-to-mesenchymal transition (EMT) lead to acquired chemoresistance, emerging studies have focused on identifying targetable drivers associated with acquired chemoresistance. Particularly, AXL, a key receptor tyrosine kinase that confers resistance against targets and chemotherapeutics, is highly expressed in mesenchymal cancer cells. However, the underlying mechanism of AXL induction in mesenchymal cancer cells is poorly understood. Our study revealed that the YAP signature, which was highly enriched in mesenchymal-type lung cancer, was closely correlated to AXL expression in 181 lung cancer cell lines. Moreover, using isogenic lung cancer cell pairs, we also found that doxorubicin treatment induced YAP nuclear translocation in mesenchymal-type lung cancer cells to induce AXL expression. Additionally, the concurrent activation of TGFß signaling coordinated YAP-dependent AXL expression through SMAD4. These data suggest that crosstalk between YAP and the TGFß/SMAD axis upon treatment with chemotherapeutics might be a promising target to improve chemosensitivity in mesenchymal-type lung cancer.