Pathological cardiac hypertrophy develops as a maladaptive response to sustained pressure overload, transitioning from compensatory adaptation to dysfunction. Understanding its molecular mechanisms is crucial for developing therapeutic strategies. Here, we identified ovarian tumor (OTU) domain-containing ubiquitin aldehyde-binding protein 2 (OTUB2) as a key regulator of pathological cardiac hypertrophy. OTUB2 expression was significantly upregulated at both transcriptional and translational levels in transverse aortic constriction (TAC)-induced hypertrophic hearts and phenylephrine (PE)-stimulated cardiomyocytes. In vivo, cardiomyocyte-specific OTUB2 overexpression via AAV9 exacerbated TAC-induced cardiac remodeling, manifested by increased heart weight/body weight ratio, impaired ejection fraction, ventricular dilatation, and enhanced fibrosis (as shown by Picrosirius red staining). In neonatal rat cardiomyocytes (NRCMs), OTUB2 overexpression aggravated while its knockdown attenuated PE-induced cardiomyocytes hypertrophy. Mechanistically, OTUB2 upregulated both total and GTP-bound active Rac1, thereby activating the downstream MEK/ERK pathway. Notably, pharmacological inhibition of Rac1 activation with NSC23766 abolished OTUB2-mediated hypertrophic responses in PE-treated cardiomyocytes. Our findings establish the OTUB2/Rac1 axis as a novel regulator of pathological cardiac hypertrophy and a potential therapeutic target for cardiac remodeling.

This study aims to screen for differentially expressed mRNAs (DEmRNAs) and lncRNAs (DELncRNAs) related to autophagy in bone nonunion through transcriptome sequencing, and explore their molecular mechanisms at the cellular level. In this study, 5 pairs of bone nonunion and bone healing tissue samples were collected for RNA sequencing to obtain DEmRNAs and DElncRNAs. The protein-protein interaction (PPI) network of autophagy-related differential genes was constructed. In addition, an lncRNAs-mRNAs interaction network was constructed. qRT-PCR was used to verify key autophagy-related genes. shBNIP3 was transfected into Wistar rat bone marrow mesenchymal stem cells (BMSCs), and 14 days after osteoinduction, cell migration and osteoblast differentiation were detected by transwell, alizarin red staining, and ALP staining. A total of 1108 DEmRNAs and 46 DElncRNAs were identified, and further screening resulted in 15 autophagy-related DEmRNAs. GO and KEGG analysis showed that autophagy-related DEmRNAs are mainly involved in autophagy-related pathways. In addition, the lncRNAs network included 356 relationship pairs, and 5 central autophagy-related mRNAs (BNIP3, DDIT3, SIRT2, PINK1, and BAG3) were identified. QRT-PCR verified that the expressions of BNIP3, SIRT2, PINK1, and BAG3 were up-regulated, while DDIT3 was down-regulated in patients with bone nonunion compared with patients with normal bone union. Besides, sh2BNIP3 inhibited BMSCs cell migration and osteogenic differentiation compared with Mock and BNIP3-NC groups in vitro. In this study, central autophagy-related DEmRNA may play an important role in bone nonunion, especially BNIP3, which will help us to further understand the pathogenesis of bone nonunion.

Relapsing and remitting mucosal inflammation can potentially spread throughout the colon in cases of ulcerative colitis (UC). N6-methyladenosine (mA) is the most prevalent epigenetic change in RNA, and it is essential to numerous biological processes and diseases, including UC. This study aims to investigate the roles and mechanisms of Wilms tumor 1-associated protein (WTAP) and protein tyrosine phosphatase non-receptor type 1 (PTPN1) in the etiology of UC and as potential therapeutic targets. PTP1B (gene name PTPN1) was significantly upregulated in UC samples and lipopolysaccharide (LPS)-stimulated NCM460 cells, influencing cell viability, DNA synthesis, and apoptosis. WTAP was identified as an upregulated methyltransferase in UC and was found to enhance PTPN1 expression through mA modification, thereby exacerbating LPS-induced inflammation and cellular dysfunction. The use of the mA modification inhibitor Cycloleucine and PTPN1 knockdown attenuated these effects, highlighting the effect of WTAP-mediated mA modification of PTPN1 on UC pathophysiology. Our findings indicate the significant role of the WTAP-mediated mA modification of PTPN1 in exacerbating UC-related inflammation and cellular dysfunction. Targeting the WTAP-PTPN1 axis could provide a novel therapeutic approach for managing UC, thereby addressing the unmet need for effective treatments in patients who are unresponsive to current treatment regimens.

The choroid plexus (ChP) is a key brain structure responsible for cerebrospinal fluid (CSF) production and forms a selective barrier that regulates brain homeostasis and immune surveillance. In vitro models of ChP are essential for studying CSF dynamics, viral entry, neuroinflammation, and CNS drug transport; yet current organoid protocols remain complex, slow, and difficult to reproduce. Here, we report a quick and robust method for the generation of human iPSC-derived ChP organoids that is xeno-free and serum-free, scalable, and reproducible. Early GSK3β inhibition and transient WNT modulation guide organoids toward cystic ChP-enriched structures, confirmed by ventricle-like morphology, and expression of canonical markers (TTR, ZO-1). This minimal workflow enables rapid production of ChP-like organoids that recapitulate ChP morphology and marker expression, providing a potential platform for studies of cerebrospinal fluid physiology, barrier modelling, and translational neuroscience.

TEX14 is essential for the formation and maintenance of the intercellular bridge (ICB) which is one of the major cellular junctions required for spermatogenesis. Previously, TEX14 was shown to block germ cell abscission prior to cytokinesis through the GPPX3Y (Gly-Pro-Pro-X-X-X-Tyr) motif. Although TEX14 could have the potential to inhibit cell proliferation, it is difficult for full-length TEX14 to be applied as anti-tumour agents because of its molecular size. In addition, the involvement of amino acids surrounding the GPPX3Y motif in the ICB formation as well as cell proliferation remains to be investigated for anti-tumour treatment. In this study, we show that partial peptides of TEX14, which include the GPPX3Y motif, are sufficient to inhibit the proliferation of a variety of cancer cells and efficiently induce apoptosis. In addition, the length and variation of amino acids surrounding the GPPX3Y motif might attenuate the efficiency of partial TEX14 peptides to inhibit cell proliferation as well as to induce apoptosis. Thus, our findings suggest that these TEX14 short peptides could be useful to suppress cell division in continuously proliferating cells such as cancer cells without affecting germ cell differentiation and have the potential as anti-tumour agents.

The lysyl oxidase family is a group of copper-containing amine oxidases involved in the remodeling of the extracellular matrix, and regulates cell signaling, metabolism, organ development, and immunity by modifying the cellular matrix. Among them, LOXL3 had been shown to regulate the proliferation, migration, and invasion of breast, colorectal, lung, melanoma, and gastric cancers. Differential expression of LOXL3 in tumor affects tumor progression by modulating immune microenvironment, where LOXL3 plays a role in regulating immune cell infiltration, immune checkpoints, and the establishment of tumor drug resistance. Here, we summarize the structure, mechanism, and functional role of LOXL3 in tumor and tumor immune microenvironment. Further, we explored strategies to improve the immunotherapy efficacy by targeting LOXL3 in tumor. Based on its properties in immunity, we explored strategies to improve the immune efficacy of LOXL3 in immunotherapy.

Dead-box RNA helicases (DDXs) are a family of proteins with roles in RNA metabolism, regulating processes such as RNA splicing, translation, and ribosome assembly. Recently, their functions have expanded to include essential roles in autophagy-a cellular degradation pathway crucial for maintaining homeostasis-and oncogenesis, notably in glioblastoma. Glioblastoma is characterized by rapid proliferation, invasiveness, and resistance to conventional treatments, making it a formidable clinical challenge. Emerging evidence suggests that specific DDXs may influence multiple key pathways that contribute to gliomagenesis, the process of glioma formation including cell cycle regulation, epithelial-to-mesenchymal transition (EMT), angiogenesis, immune modulation, anti-inflammatory signaling, and autophagy. Understanding the dual role of DDXs in autophagy and gliomagenesis may reveal potential therapeutic targets, as manipulating these helicases could disrupt cancer cell adaptation mechanisms and slow tumor progression. We have also explored the potential of autophagy inhibitors to enhance the efficacy of current therapeutics. This review aims to explore the implications of DDXs in glioblastoma, focusing on their interactions with cellular pathways, and highlights the need for further investigation into how these proteins could be leveraged for therapeutic benefit.

The prognosis of gastric cancer with peritoneal dissemination is poor because of its resistance to chemotherapy. To investigate the mechanism of drug resistance in peritoneal metastasis, cancer organoids were established from the ascites of a patient with peritoneal metastases of gastric cancer. The histological characteristics of the tumors were preserved in the organoids. A co-culture system was established by overlaying human-derived mesothelial cells on gastric cancer organoids embedded in type IA collagen, mimicking peritoneal dissemination foci. When co-cultured with mesothelial cells, the proliferation of ascites-derived gastric cancer organoids and other primary gastric cancer organoids was suppressed. Soluble factors derived from mesothelial cells were involved in suppressing cell proliferation. Organoids in co-culture showed reduced sensitivity to paclitaxel. This co-culture model may provide a useful platform for studying drug resistance mechanisms in the microenvironment of gastric cancer peritoneal metastases.

Dermatofibrosarcoma protuberans (DFSP) is a rare, locally aggressive cutaneous sarcoma characterized by high recurrence rates and the development of resistance to imatinib. The scarcity of preclinical models hinders research into DFSP pathogenesis and the development of novel therapeutic strategies. In this study, we established and characterized a novel DFSP cell line, designated DFSP-DPH1, derived from a 47-year-old male patient with an abdominal tumor. Comprehensive characterization confirmed that DFSP-DPH1 retains key features of the original tumor, including the fibroblast-like spindle morphology and expression of diagnostic markers CD34 and vimentin, with absence of factor XIIIa. Short tandem repeat profiling confirmed the cell line's origin and excluded cross-contamination. Sanger sequencing revealed a COL1A1 exon 46-PDGFB exon 2 fusion transcript, a breakpoint not previously reported in established DFSP cell lines. Functionally, DFSP-DPH1 exhibits robust proliferative capacity, forms three-dimensional spheroids under anchorage-independent conditions, and demonstrates significant migratory and invasive capabilities. Drug sensitivity screening of a panel of 48 PDGFR inhibitors confirmed its resistance to imatinib and identified several compounds with superior efficacy compared to imatinib. Transcriptomic analysis confirmed the dominance of the COL1A1::PDGFB fusion transcript and revealed enrichment of pathways related to cancer, viral infection, and neuroactive ligand-receptor interaction. This novel imatinib-resistant DFSP cell line, DFSP-DPH1 provides a valuable preclinical model for investigating the molecular mechanisms underlying DFSP pathogenesis, drug resistance, and tumor progression, and for developing and evaluating novel therapeutic strategies.

Pleural effusion (PE) is a common clinical manifestation associated with advanced stages of both malignant and non-malignant diseases. PE frequently occurs in advanced non-small cell lung cancer (NSCLC) and contributes to tumor progression. NSCLC accounts for more than 85% of the lung cancers and remains a problem worldwide due to its late diagnosis and low rate of response to treatment. Extracellular vesicles (EVs) present in PE are emerging as key mediators of intercellular communication, capable of transferring oncogenic signals through their molecular cargo. Among these molecules, microRNAs (miRNAs) are increasingly recognized as important drivers of cancer progression. miR-21 is a representative onco-miRNA, involved in lung cancer progression; moreover EV-miR-21 upregulation at the pre-dissemination stage promotes cancer cell survival in the pleural cavity. This study compares, for the first time, the functional role of EVs isolated from malignant PE in NSCLC patients (NSCLC-PE-EVs) with those isolated from PE in patients with congestive heart failure (CHF-PE-EVs), focusing on their ability to modulate lung cancer cell behavior. The effects of these EVs were evaluated on COLO699 lung adenocarcinoma cells with proliferation, migration, and gene expression assays. NSCLC-PE was found to contain approximately twice the amount of EVs compared to CHF-PE. NSCLC-PE-EVs were enriched in the oncogenic miR-21-5p, while CHF-PE-EVs had higher levels of the tumor-suppressive miR-126-3p. Only NSCLC-PE-EVs induced dose-dependent increases in COLO699 cell proliferation and migration, consistent with elevated miR-21-5p expression. Functional studies confirmed that miR-21-5p mediates these effects by downregulating PTEN and PDCD4, and by upregulating MMP9 expression. Our findings show that NSCLC-PE-EVs promote malignant phenotypes in lung cancer cells via the transfer of miR-21-5p.

In vitro models of acute leukemia are crucial for understanding its biology and developing effective treatments. The authors have established and characterized a novel cell line, ICH-BCPALL-3, which expresses the TCF3::HLF fusion from B cell precursor acute lymphoblastic leukemia (BCP-ALL). The karyotype of the cultured cells is 46,XY, der(1)(1qter- > 1q11::1p32- > 1q11::4q21- > 4qter), der(4)t(1;4)(q11;p32), add(8)(q24), del(17)(q24). Analysis of the diagnostic sample revealed deletions in RB1, VPREB1, and NR3C1. The cell line showed additional deletions of VPREB1, NR3C1, and CDKN2A/2B, as well as a gain of AKT1. The loci for PAX5 and BTG1 were retained. Exome and Sanger sequencing identified nucleotide variants of ARID5B and NCOR1 in the diagnostic sample, as well as a KRAS variant (p.Lys117Asn) in the first recurrent sample and another KRAS variant (p.Asp119Gly) in the second recurrent sample and the cell line. Transcriptome analysis and RT-PCR confirmed that all examined samples contained a TCF3::HLF chimeric transcript. However, molecular cytogenetics did not verify the juxtaposition of TCF3 and HLF loci. Further long-range PCR analyses confirmed that genomic material containing HLF exon 4 was inserted into TCF3 intron 16. Using dimensional reduction techniques, we found that the current cell line shares an expression pattern with other TCF3::HLF-positive BCP-ALL cell lines. The cytotoxicity assay indicated that the cell line is sensitive to Aurora Kinase B inhibitor, but not to BCL2 inhibitor. This cell line is the first TCF3::HLF-positive BCP-ALL model without the t(17;19) translocation, facilitating research into leukemogenesis and the development of novel treatments for patients with poor prognosis associated with TCF3::HLF-positive BCP-ALL.

Radioresistance of adenocarcinoma cells limits the efficiency of radiotherapy. In addition to the cell nucleus, ionizing radiation (IR) also induces damage to the mitochondria. Mitophagy, a selective degradation of impaired mitochondria via autophagy, has been found to respond to IR, but its role in the radiosensitivity of adenocarcinoma cells remains unclear. Using several different adenocarcinoma cell lines, we confirmed that exposing the adenocarcinoma cells to 5 Gy X-ray enhanced the expression of some mitophagy receptors and increased mitophagy activity. However, pharmacological inhibition of mitophagy by mdivi-1 did not significantly change the radiosensitivity of HCT116 and A549 cells. Similarly, molecular targeting inhibition of mitophagy by BNIP3L knockdown in HCT116 and A549 cells that showed significant IR-induced BNIP3L up-regulation did also not significantly affect the radiosensitivity of adenocarcinoma cells, although the IR-induced enhancement of mitophagy activity was effectively suppressed. According to our data, mitophagy is responsible to IR but plays a very limited role in the radiosensitivity of adenocarcinoma cells. Further in vivo studies are warranted to elucidate the radiosensitizing effect of targeting mitophagy on malignant tumors.

Clear cell renal cell carcinoma (ccRCC) is the most common type of renal cell carcinoma, and exploration of its molecular mechanism benefits for developing more effective molecular targeted drugs. CDC6 has been found to be highly expressed in a variety of malignancies and plays oncogenic role; however, its function in ccRCC has not been elucidated. In this work, immunohistochemical (IHC) staining was used to detect protein expression of genes in clinical tissues. qPCR and WB were used for expression detection of mRNA and protein levels in cells. The Celigo assay, plate cloning assay, flow cytometry, and wound-healing/Transwell assays were used to detect cell proliferation, colony formation, apoptosis, and migration, respectively. A subcutaneous xenograft model in nude mice was used to verify the function of CDC6 in vivo. The results of clinical sample-related detection as well as analysis showed that CDC6 was highly expressed in ccRCC and was significantly associated with higher tumor malignancy as well as worse patients' prognosis. Knockdown of CDC6 in ccRCC cells significantly inhibited cell proliferation and migration while promoting apoptosis, and inhibited in vivo growth of transplanted tumors in animal models. Mechanistically, RRM2 is identified as a potential downstream effector molecule that has co-expression characteristics with CDC6 and whose expression levels are regulated by it. More importantly, RRM2 knockdown mediated tumor suppression could partially reversed CDC6 overexpression induced tumor promotion. This study identified CDC6/RRM2 axis as a potential target for development of novel targeted therapy for ccRCC treatment.

N6-methyladenosine (mA) is the most prevalent mRNA internal modification in eukaryotic mRNAs and is frequently associated with progression and immune response in human cancers. This study delves into the function of mA reader insulin like growth factor 2 mRNA binding protein 3 (IGF2BP3) in the immune evasion and progression of laryngeal squamous cell carcinoma (LSCC), along with its underpinning mechanisms. We conducted integrated bioinformatics to examine mA-modifying regulators and their prognostic values in LSCC. IGF2BP3 was identified as a promising candidate, which was verified to be highly expressed in LSCC tissues and cell lines. Furthermore, the aberrant IGF2BP3 upregulation in the context of LSCC was found to be partly ascribed to DNA hypomethylation. IGF2BP3 was found to elevate RMND5A expression in an mA-dependent manner. Silencing either IGF2BP3 or RMND5A significantly decreased the viability, colony formation ability, and tumorigenicity of LSCC cells. Moreover, this intervention reduced the protein level of PD-L1 in cells while increasing CD8 T cell infiltration in xenograft tumors. However, further upregulation of RMND5A negated the tumor-suppressive and immune-enhancing effects observed upon IGF2BP3 silencing. In conclusion, this study demonstrates that IGF2BP3 elevates RMND5A expression through mA modification, thereby promoting malignant properties and immune evasion in LSCC cells.

We aimed to generate immortalized stromal cell lines from the ovarian and fallopian tube tissues of a single patient using Sendai virus (SeV) vectors and identify candidate stromal genes involved in ovarian carcinogenesis. Tissues were collected from a 48-year-old woman with endometrioid borderline tumors and endometriomas. Primary cultures were established from the right ovarian endometrioma, left ovarian surface, bilateral fallopian tube, and endometrial surface. Immortalization was achieved using SeV vectors encoding human telomerase reverse transcriptase (TERT), B lymphoma Mo-MLV insertion region 1 homolog (Bmi-1), and Simian virus 40 large T antigen (SV40T). Morphologically, the established cells exhibited spindle-shaped fibroblast-like features and expressed stromal markers (Vimentin-positive, Keratin-negative), confirming their stromal origin. Genetic and molecular changes associated with immortalization were evaluated via chromosomal analyses, transcriptome sequencing, and reverse transcription-polymerase chain reaction (RT-PCR). SeV-infected stromal cell lines retained their proliferative capacity for over 25 passages, whereas non-infected primary cells lost their epithelial characteristics and underwent senescence after five passages. Chromosomal abnormalities were more prevalent in stromal cells derived from the ovarian endometriomas, suggesting early genomic instability. Transcriptomic profiling and RT-PCR revealed upregulation of matrix metallopeptidase 1 (MMP1), pregnancy-associated plasma protein A (PAPPA), and C-X-C motif chemokine ligand 1 in cyst-derived stromal cells compared to those from the normal ovary and fallopian tube, implicating these genes in extracellular matrix remodeling and tumor-stroma crosstalk. We established immortalized ovarian and fallopian tube stromal cell lines using SeV-based vectors. The cyst-derived stromal cells exhibited early chromosomal instability and overexpression of MMP1 and PAPPA, supporting their potential role in ovarian carcinogenesis. These immortalized stromal cell lines provide a novel and stable platform for mechanistic studies and may contribute to biomarker discovery and therapeutic target development in ovarian cancer.

Mitochondrial dysfunction is a key contributor to septic cardiomyopathy, driving myocardial inflammation and apoptosis. This study found that Sushi-repeat containing protein X-linked 2 (Srpx2) is associated with mitochondrial damage and is downregulated in the myocytes of lipopolysaccharide (LPS)-treated rats. Sprague Dawley rats were intraperitoneally injected with LPS (10 mg/kg) to establish a septic cardiomyopathy model. Adeno-associated viruses (8.5 × 10 vg/mL) containing Srpx2-overexpressing plasmids were injected into rats through their tail vein. Srpx2 overexpression improved hemodynamics and decreased myocardial damage in LPS-treated rats. H9C2 cells were treated with LPS (10 μg/mL) to establish an in vitro septic model. The cells were then transfected with Srpx2-overexpressing plasmids. Srpx2 overexpression ameliorated mitochondrial damage, which was evidenced by restoring mitochondrial morphology, enhancing the complex activities, and elevating ATP and mitochondrial membrane potential levels in cardiomyocytes. Srpx2 overexpression reduced mitochondrial reactive oxygen species levels and superoxide generation in cardiomyocytes. Srpx2 overexpression decreased cleaved caspase-3 and 9 protein levels. Tumor necrosis factor-α and interleukin-1 beta levels were also reduced in cardiomyocytes with Srpx2 overexpression, suggesting reversal of inflammation. The RNA-sequencing data indicated that Srpx2 might regulate the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) pathway to protect cardiomyocytes. Srpx2 overexpression increased the p-PI3K and p-AKT protein levels. The treatment of H9C2 cells with 10 μM LY294002, a PI3K/AKT pathway inhibitor, reversed the protective effects of Srpx2 against mitochondrial damage and apoptosis. In conclusion, Srpx2 alleviates mitochondrial damage by activating the PI3K/AKT pathway, thereby reducing apoptosis and inflammation in septic cardiomyopathy.

The malignant progression of colorectal cancer (CRC) is closely related to cell stemness, but its regulatory mechanism has not been fully elucidated. This study found that ADAMTS14 was significantly highly expressed in CRC tissues and cell lines and was associated with poor prognosis in patients. Functional experiments have confirmed that ADAMTS14 enhances the stemness characteristics (such as upregulation of ALDH1A1, ALDH1A3, and CD133 expression) and spheroidization ability of CRC cells by activating the Wnt signaling pathway. Further mechanism studies have shown that the transcription factor ONECUT2 is also highly expressed in CRC and indicates a poor prognosis, and it can directly activate its transcription by binding to the ADAMTS14 promoter region. In conclusion, this study has revealed a novel mechanism by which the ONECUT2/ADAMTS14/Wnt axis regulates the stemness of CRC cells, providing a potential molecular target for targeted intervention.

Endothelin-1 (ET-1) plays a critical role in diabetic vasculopathy. Although clinical trials have shown promise for ET-1 receptor antagonists in treating diabetic nephropathy, their clinical use remains limited by adverse effects. MiR-454-3p targets ET-1. This study aimed to investigate the role of miR-454-3p in modulating ET-1 expression and related molecular changes in endothelial cells (ECs) under high glucose conditions using both bioinformatics and experimental approaches. Bioinformatics analysis identified 10 miR-454-3p target genes expressed in ECs previously implicated in diabetic vascular complications: ET-1, GJA1, IRF1, PIK3CB, TRPC3, SLMAP, ESR1, ITGB8, MAPK1, and PPARG. With the exception of PPARG, which protects ECs from hyperglycemia-induced damage, all have been reported to exacerbate endothelial dysfunction. Western blotting showed that high glucose increased ET-1 expression in human umbilical vein ECs (HUVECs) and human dermal microvascular endothelial cells (HDMECs), while miR-454-3p overexpression significantly suppressed this effect in both cell types. Conditioned medium (CM) from HUVECs transfected with miR-454-3p mimics enhanced eNOS expression in recipient cells, compared to control CM. Pre-treatment of HUVEC control CM with an anti-ET-1 antibody also increased eNOS expression, supporting that miR-454-3p promotes NOS production partly via ET-1 suppression. MiR-454-3p overexpression in HUVECs did not affect PPARG expression or cell proliferation. In conclusion, miR-454-3p overexpression inhibits high glucose-induced ET-1 expression in HUVECs and HDMECs, and promotes eNOS production without affecting PPARG expression in HUVECs. Our findings suggest that miR-454-3p modulates ET-1 expression under hyperglycemic conditions in vitro, which may provide a foundation for future studies exploring its potential application in managing diabetic vasculopathy.

A 62-year-old woman with EGFR exon 19-mutant lung adenocarcinoma developed resistance to osimertinib after 13 months of treatment. At progression, genetic analysis revealed persistence of the EGFR mutation and acquisition of a TPM3-NTRK1 fusion. Cells from the pleural effusion at this stage were cultured and established as a continuous cell line, LUNK1. LUNK1 cells displayed epithelial characteristics, with a population doubling time of 57.66 h in vitro and 100% tumorigenicity in vivo. Xenograft histopathology showed poorly differentiated lung adenocarcinoma exhibiting solid, acinar, and micropapillary patterns. Immunohistochemistry confirmed positivity for CK7, TTF-1, Napsin A, EGFR, and pan-TRK. EGFR exon 19 deletion and TPM3-NTRK1 fusion were confirmed by whole-exome and RNA sequencing, respectively, and further validated by PCR and Sanger sequencing. Drug sensitivity assays revealed an IC₅₀ of 64.0 nM for entrectinib and 806.8 nM for osimertinib, indicating reduced sensitivity to EGFR inhibition. NTRK1 knockdown significantly increased osimertinib sensitivity (124.5-264.5 fold) and reduced entrectinib sensitivity (8.7-19.0 fold), suggesting NTRK1 as a key regulator of drug response. We provided experimental evidence for that TPM3-NTRK1 fusions can mediate acquired resistance to osimertinib in a new lung adenocarcinoma cell line. LUNK1 represents a useful preclinical model for investigating resistance mechanisms and assessing dual-targeted treatment strategies.

Asthma is a chronic inflammatory disease characterized by airway hyperresponsiveness, airway remodeling, and persistent inflammation, with neutrophilic phenotypes often associated with severe and steroid-resistant cases. SerpinB1, a serine protease inhibitor, has been shown to modulate inflammatory processes through its interaction with neutrophil elastase (Elane); however, its specific role in asthma pathogenesis has not been fully clarified. In the present study, we investigated the function of SerpinB1 in an ovalbumin (OVA)-induced mouse model of asthma as well as in lipopolysaccharide (LPS)-stimulated BEAS-2B cells. Expression analysis revealed that SerpinB1 was downregulated in asthmatic mice. Overexpression of SerpinB1 markedly alleviated neutrophil-driven airway inflammation, reduced structural remodeling of the airways, and suppressed pyroptosis, as demonstrated by decreased expression of caspase-1, GSDMD, IL-1β, and NLRP3. Co-immunoprecipitation and immunofluorescence assays further confirmed that SerpinB1 directly interacts with Elane. Importantly, knockdown of Elane abolished the protective effects conferred by SerpinB1, indicating that the regulation of asthma-related pathology by SerpinB1 is mediated through Elane inhibition. In conclusion, SerpinB1 may mitigate airway inflammation and remodeling in asthma by targeting Elane and suppressing pyroptosis, supporting it as a potential therapeutic target for neutrophil-dominant asthma.