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Protein disulfide isomerase (PDI) regulates multiple protein functions by catalyzing the oxidation, reduction, and isomerization of disulfide bonds. The enzyme is considered a potential target for treating thrombosis. We previously developed a potent PDI inhibitor, CPD, which contains the propiolamide as a warhead targeting cysteine residue in PDI. To address its issues with undesirable off-target effects and weak metabolic stability, we replaced the propiolamide group with various electrophiles. Among these, compound 2d, which contains 2-trifluoromethyl acrylamide exhibited potent PDI inhibition compared to the reference PACMA31. Further structural optimization of compound 2d led to a novel series of 2-trifluoromethyl acrylamide derivatives. Several of these compounds displayed substantially improved enzyme inhibition. Notably, compound 14d demonstrated the strongest inhibition against PDI, with an IC 50 value of 0.48 ± 0.004 μM. Additionally, compound 14d was found to exhibit a reversible binding mode with PDI enzyme. Further biological evaluations show that 14d suppressed platelet aggregation and thrombus formation by attenuating GPIIb/IIIa activation without significantly causing cytotoxicity. Altogether, these findings suggest PDI inhibitors could be a potential strategy for anti-thrombosis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Lung cancer is the leading cause of cancer-related mortality globally. N6-methyladenosine (m6A) is the most abundant modification in mammalian mRNA and is involved in the biological regulation of tumors, including lung cancer. However, the role of m6A-related proteins, such as RNA-binding motif protein 15 (RBM15), in lung cancer progression remains largely unknown. Our study indicated that RBM15 is significantly overexpressed in lung adenocarcinoma, serving as an independent prognostic factor for poor outcomes and facilitating tumor cell proliferation and migration. RBM15 was markedly elevated in patients with EGFR mutations, correlating with a poorer prognosis, while it had negligible prognostic value in EGFR wild-type patients. As EGFR-tyrosine kinase inhibitors (TKIs) are the standard treatment for patients with EGFR mutations, we subsequently determined that RBM15 drives osimertinib resistance via a novel mechanism: enhancing m6A modification of cwcv- and kazal-like domains proteoglycan 1 (SPOCK1) mRNA, promoting epithelial-mesenchymal transition-mediated osimertinib resistance through a bypass activation pathway. These findings were validated in osimertinib-resistant H1975 cells and organoids from patients with osimertinib-resistant lung adenocarcinoma. Furthermore, the RBM15-SPOCK1 axis was activated in drug-tolerant persister cells, indicating that early targeting of RBM15 during EGFR-TKI treatment could dramatically extend the patient response and benefit from TKI therapy. Our results emphasize the critical role of RBM15 in reversing EGFR-TKI resistance and propose it as a promising therapeutic target for prolonging TKI treatment benefits in patients with lung adenocarcinoma., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Overcoming acquired resistance to Osimertinib remains a critical challenge in treating NSCLC. This research indicates that Osimertinib-resistant cells exhibit a strong dependence on glutamine metabolism. However, targeting GLS1 shows limited anticancer effects, probably because it cannot fully block the glutamine metabolic pathway. The investigation reveals that a more effective strategy involves simultaneously inhibiting both ASCT2 and GLS1. After confirming the efficacy of this dual-targeting approach against Osimertinib-resistant cells in preclinical models, the potential of utilizing a broad-spectrum glutamine metabolism antagonist is further explored to achieve superior antitumor efficacy. DON, broad-spectrum glutamine antagonist, presents toxicity issues. Herein, the high NQO1 expression in Osimertinib-resistant NSCLC cells is leveraged to design an NQO1-responsive DON prodrug, 10e (LBJ-10e). This prodrug demonstrates superior safety compared to natural DON and greater antitumor activity against resistant tumors compared to the clinical phase II drug DRP104. These findings may address the clinical limitations of GLS1 allosteric inhibitors and underscore prodrug strategies in effectively treating Osimertinib-resistant lung cancer, providing a foundation for future clinical trials., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

In this study, a patient with lung adenocarcinoma harboring an EGFR mutation exhibited primary resistance to the targeted EGFR inhibitor Osimertinib after 2 months of treatment. As the disease advanced, further genetic analysis revealed the emergence of additional mutations in ARID1A, NTRK1, and ZRSR2, alongside the existing EGFR mutation. Subsequent treatment with Pemetrexed resulted in a significant reduction in liver metastases. Protein mass spectrometry sequencing and immunohistochemical analysis collectively indicated that the PI3K/mTOR pathway mediates the mechanism through which these gene mutations confer primary drug resistance. Evidence demonstrates that the co-occurrence of EGFR and ARID1A mutations diminishes the efficacy of EGFR tyrosine kinase inhibitors (EGFR TKIs). Consequently, it is hypothesized that mutations in NTRK1 and ZRSR2, which are implicated in the PI3K/mTOR pathway, contribute to the primary resistance observed with Osimertinib treatment. In this case, the illness was effectively managed through prompt adjustments to the treatment regimen and the rapid administration of chemotherapy drugs. This finding also constitutes the first evidence that mutations in NTRK1 and ZRSR2 are pivotal in the development of primary resistance to Osimertinib. Consequently, it is imperative to conduct genetic testing at the earliest opportunity and modify the treatment plan accordingly., (© 2025 The Author(s). IUBMB Life published by Wiley Periodicals LLC on behalf of International Union of Biochemistry and Molecular Biology.)

The management of diabetic wounds faces significant challenges due to the excessive activation of reactive oxygen species (ROS), dysregulation of the inflammatory response, and impaired angiogenesis. A substantial body of evidence suggests that the aforementioned diverse factors contributing to the delayed healing of diabetic wounds may be associated with impaired autophagy. Impaired autophagy leads to endothelial and fibroblast dysfunction and impedes macrophage phenotypic transformation. This disruption hinders angiogenesis and extracellular matrix deposition, ultimately culminating in delayed wound healing. Therefore, biomaterials possessing autophagy regulatory functions hold significant potential for clinical applications in enhancing the healing of diabetic wounds. A hybrid multifunctional hydrogel (GelMa@SIS-Qu) has been developed, comprising methacrylamide gelatin (GelMa), a small intestine submucosal acellular matrix (SIS), and quercetin nanoparticles, which demonstrates the capability to promote autophagy. The promotion of autophagy not only reduces ROS levels in endothelial cells and enhances their antioxidant activity but also mitigates ROS-induced endothelial cell dysfunction and apoptosis, thereby promoting angiogenesis. Furthermore, the promotion of autophagy facilitates the phenotypic transformation of macrophages from the M1 phenotype to the M2 phenotype. This study investigates the distinctive mechanisms of the GelMa@SIS-Qu hydrogel and proposes a promising therapeutic strategy for treating diabetes-related wounds.

Background: Afatinib and Osimertinib are first-line treatments for EGFR-mutated advanced non-small cell lung cancer (NSCLC), but their comparative efficacies and the patient groups that benefit the most remain unclear. This multicenter retrospective study evaluated the efficacy of first-line Afatinib and Osimertinib in NSCLC patients with EGFR 19del and no brain metastases at diagnosis., Methods: The primary endpoints were time on treatment (ToT) and overall survival (OS). Survival analyses were performed for three groups: Afatinib followed by Osimertinib, Afatinib followed by other therapies, and Osimertinib (alone or followed by other therapies). Rebiopsy practices, including T790M mutation detection, were also analyzed in patients with disease progression on Afatinib., Results: Among 97 Afatinib-treated and 60 Osimertinib-treated patients, Osimertinib showed a significantly longer ToT (23.3 vs. 16.5 months; p = 0.007). Median OS was numerically higher for Afatinib with sequential Osimertinib (40.5 vs. 34.6 months for Osimertinib; p = 0.473). Osimertinib demonstrated advantages, with fewer brain metastases upon progression and fewer adverse effects. In the Afatinib group, 64% of patients with disease progression underwent rebiopsy, with 39% testing positive for T790M mutation and subsequently receiving Osimertinib. Rebiopsy was most frequently performed on the lung parenchyma using non-surgical methods., Conclusions: In this real-world study, Osimertinib achieved a significantly longer ToT compared to Afatinib in NSCLC patients with EGFR 19del and no brain metastases. The sequential use of Afatinib followed by Osimertinib showed a trend toward improved OS, highlighting the importance of rebiopsy for identifying T790M mutations to guide subsequent therapy.

The epidermal growth factor receptor (EGFR) signaling pathway is crucial for skin barrier integrity and immune response. This study explores the impact of EGFR inhibitors, osimertinib and afatinib, on keratinocyte function, focusing on keratin (KRT1, KRT17) and tight junction protein (CLDN1, CLDN2, CLDN4) expression in HaCaT cells. Osimertinib significantly increased the mRNA and protein levels of keratins and inflammatory markers, IL-6 and TNF-α, via activation of the EGFR-STAT3 signaling pathway. Co-treatment with recombinant human EGF reversed these changes, suggesting the pathway's modulatory role. These findings underscore the potential therapeutic applications of targeting the EGFR-STAT3 axis in skin barrier dysfunction and inflammatory skin disorders., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Various studies have reported resistance mechanisms and treatment methods after epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor treatment; however, treatment policies have not yet been established, and few cases have reported transformation to pleomorphic carcinoma (PC) as the resistance mechanism. Herein, we report the case of a 66-year-old woman who was diagnosed with Stage 4A lung adenocarcinoma (cT2bN0M1b) through bronchoscopic biopsy. Genetic profiling revealed an EGFR L858R mutation; therefore, osimertinib was administered as the first-line therapy and achieved a partial response. After 46 months of osimertinib treatment, the metastases remained under control; however, the primary tumor enlarged and was therefore resected. Pathological examination confirmed the diagnosis of PC. Genetic testing of the surgical pathology specimen showed that the EGFR mutation L858R was retained, and the patient was considered drug-resistant owing to the histologic transformation to PC. The patient continued osimertinib therapy and had no recurrence at 9 months postoperatively. Transformation to PC following osimertinib administration is rare, and we report this unique case. This study was approved by the Jichi Medical University Saitama Medical Center Ethics Committee (S24-073), and written informed consent was obtained from the patient., (© 2024 The Author(s). Thoracic Cancer published by John Wiley & Sons Australia, Ltd.)

In this study, we investigated the synergistic effect of co-administration of osimertinib and HAD-B1 using gefitinib-resistant non-small cell lung cancer cells, HCC827-GR. HAD-B1 is composed of 4 natural drugs, Panax Notoginseng Radix, Panax ginseng C. A. Meyer, Cordyceps militaris, and Boswellia carterii Birdwood, and has been reported to have therapeutic effects on patients with advanced non-small cell lung cancer in several studies. Resistance to gefitinib in HCC827 cells was acquired through MET activity. Co-treatment with osimertinib and HAD-B1 reduced the cell viability of HCC827-GR cells. In addition, phosphorylation of MET and ERK were effectively suppressed for HCC827-GR cells. And, compared to when osimertinib and HAD-B1 were administered alone, cell proliferation was significantly inhibited and apoptosis was effectively induced when osimertinib and HAD-B1 were co-administered to HCC827-GR cells. We found that the synergistic effect of osimertinib and HAD-B1 combination therapy resulted in cancer cell death and cell cycle arrest by targeting the ERK and mTOR signaling pathways. In conclusion, this study confirmed that the combination of osimertinib, a third-generation anticancer drug, and HAD-B1, a natural anticancer drug, had a potentially synergistic effect on non-small cell lung cancer resistant to EGFR-targeted anticancer drugs., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

HDAC3 has been demonstrated to play a crucial role in the progression of various tumors and the differentiation and development of T cells. However, its impact on peripheral T cells in the development of murine lung cancer remains unclear. In this experiment, a subcutaneous lung tumor model was established in C57BL/6 mice, and tumor-bearing mice were treated with the specific inhibitor of HDAC3, RGFP966, at different doses to observe changes in tumor size. Additionally, a lung tumor model was established using hdac3 fl/fl cd4cre +/+ mice to investigate its mechanism. Mice injected with 10 mg/kg RGFP966 had the smallest tumor volume, while those injected with 30 mg/kg RGFP966 had the largest tumors. Flow cytometry analysis revealed that the expression of HDAC3 in splenic T cells was reduced in all groups of mice, while IFN-γ and IL-17 A were increased. Moreover, the expression of granzyme B and perforin in splenic CD8 + T cells was increased in all groups of mice. Compared to the use of 30 mg/kg RGFP966 alone, the combination with anti-IL-17 A mAb reduced the infiltration of Neutrophils and exhausted T cells in mouse tumors, thereby impeding tumor development. These findings demonstrate that the use of RGFP966 or T cell-specific loss of hdac3 promotes the expression of IL-17 A in splenic T cells, leading to tumor resistance and providing insights for clinical treatment., Competing Interests: Declarations. Competing interests: The authors declare no competing interests. Accordance: We confirmed that all experiments in this study were performed in accordance with the relevant guidelines and regulations. Arrive: All the procedure of the study is followed by the ARRIVE guidelines., (© 2024. The Author(s).)

Lung adenocarcinoma (LUAD) is one of the deadliest cancers. Epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI)-targeted therapy is an important approach for treating LUAD. However, the development of acquired resistance poses a serious clinical challenge. Our objective was to explore the differentially expressed genes (DEGs) associated with EGFR and detect biomarkers for diagnosing and treating osimertinib resistance in LUAD patients. LUAD datasets were downloaded from public databases. Differential expression analysis was performed to screen DEGs, and prognostic modules were constructed by Cox regression. Enrichment analysis, gene regulatory network analysis and immune microenvironment analysis were employed to explore the underlying mechanisms in LUAD. Finally, the expression of prognosis module genes (PMGs) was validated in 8 LUAD tissue specimens and 5 cell lines by qRT-PCR. In total, 13 differential module genes (BIRC3, CCT6A, CPLX2, GLCCI1, GSTA1, HLA-DQB2, ID1, KCTD12, MUC15, NOTUM, NT5E, TCIM, and TM4SF4) were screened for the construction of a prognostic module. Notably, CCT6A and KCTD12 demonstrated excellent accuracy in the diagnosis of LUAD. Immune dysregulation and BIRC3, HLA-DQB2, KCTD12, and NT5E expression were significantly associated with invasive immune cells in LUAD patients. The expression level of CCT6A was highest in PC9-OR and H1975-OR cells, while the expression level of KCTD12 was highest in paracancerous tissue and HBE cells. The constructed prognostic model showed promise in predicting the survival of LUAD patients. Notably, KCTD12 and CCT6A might be candidate biomarkers for improving diagnostic performance and guiding individualized therapy for EGFR-TKI-resistant LUAD patients., Competing Interests: Declarations. Ethics approval and consent to participate: The studies involving human participants were reviewed and approved by the Ethics Committee of the Affiliated Guangdong Medical University (reference number: PJKT2024-023). The patients/participants provided written informed consent to participate in this study. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Objective: The combination of epithelial growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) and immune checkpoint inhibitors (ICIs) leads to an increased incidence of severe immune-related adverse events (irAEs). However, the mechanisms underlying macrophages in irAEs have not been elucidated., Methods: An osimertinib and ICI-induced irAE mouse model was constructed. Lung micro-CT scans were used to assess the degree of inflammatory infiltration. Hematoxylin-eosin staining was used to analyze the histopathologic inflammatory infiltration in mouse liver and lung tissues. Flow cytometry was used to detect the percentages of T cells, NK cells, and macrophages and the expression of EGFR. Enzyme-linked immunosorbent assay (ELISA) was used to detect the serum interleukin (IL)-6, alanine transaminase (ALT), ferritin, and tumor necrosis factor (TNF)-α levels. Total RNA extracted from mouse liver macrophages was analyzed by RNA-seq. Simple Western blot analysis was used to detect the IL-6/JAK/STAT3 pathway activation state., Results: Osimertinib combined with ICIs upregulated EGFR expression on macrophages with increased serum IL-6, ALT, and ferritin levels. RNA-seq and simple Western blot analysis of mouse liver macrophages confirmed that that the IL-6/JAK/STAT3 pathway was activated in the combination treatment group. Ruxolitinib blocked the IL-6/JAK/STAT3 pathway and significantly decreased the serum IL-6, ALT, and ferritin levels in the combination treatment group., Conclusions: An osimertinib and ICI-induced irAE mouse model was constructed that showed osimertinib combined with ICIs inhibited EGFR phosphorylation and activated the IL-6/JAK/STAT3 signaling pathway in mouse liver macrophages, which led to the release of relevant cytokines., Competing Interests: No potential conflicts of interest are disclosed., (Copyright © 2024 The Authors.)

Osimertinib, a third generation epidermal growth factor receptor tyrosine kinase inhibitor, is approved as a first-line therapy in patients with advanced non-small cell lung carcinoma (NSCLC) with EGFR-activating mutations or the T790M resistance mutation. However, the efficacy of osimertinib is limited due to acquired resistance, highlighting the need to elucidate resistance mechanisms to facilitate the development of improved treatment strategies. Here, we screened for significantly upregulated genes encoding protein kinases in osimertinib-resistant NSCLC cells and identified NUAK1 as a pivotal regulator of osimertinib resistance. NUAK1 was highly expressed in osimertinib-resistant NSCLC and promoted the emergence of osimertinib resistance. Genetic or pharmacological blockade of NUAK1 restored the sensitivity of resistant NSCLC cells to osimertinib in vitro and in vivo. Mechanistically, NUAK1 directly interacted with and phosphorylated nicotinamide adenine dinucleotide kinase (NADK) at serine 64 (S64), which mitigated osimertinib-induced accumulation of reactive oxygen species (ROS) and contributed to the acquisition of osimertinib resistance in NSCLC. Furthermore, virtual drug screening identified T21195 as an inhibitor of NADK-S64 phosphorylation, and T21195 synergized with osimertinib to reverse acquired resistance by inducing ROS accumulation. Collectively, these findings highlight the role of the NUAK1-NADK axis in governing osimertinib resistance in NSCLC and indicate the potential of targeting this axis as a strategy for circumventing resistance. Significance: Phosphorylation of NADK by NUAK1 diminishes ROS accumulation and confers resistance to osimertinib, identifying NUAK1-NADK signaling as a potential therapeutic target for improving the response to EGFR inhibition in lung cancer., (©2024 American Association for Cancer Research.)

HER2-positive breast cancer (HER2+ BC) is distinguished by its poor prognosis, propensity for early onset, and high risk of recurrence and metastasis. Consequently, anti-HER2-targeted therapy has emerged as a principal strategy in the treatment of this form of breast cancer. Pyrotinib, a novel irreversible pan-HER2 tyrosine kinase inhibitor, has brought fresh hope to patients with advanced HER2+ breast cancer. In this study, we conducted a comprehensive exploration of pyrotinib's antitumor mechanism. The in vitro results showed that pyrotinib significantly inhibited SKBR3 cells viability and induced apoptosis by promoting HER2 endocytosis and ubiquitylation, leading to HER2 degradation through the displacement of HSP90 from HER2. Beyond targeting the HER2 signaling pathway, pyrotinib also induced DNA damage, which was mediated by the activation of the reactive oxygen species/heat shock factor 1 signaling pathway and the downregulation of proliferating cell nuclear antigen expression. Furthermore, the in vivo results demonstrated a pronounced anticancer effect of pyrotinib in the SKBR3 xenograft mouse model, concomitant with a reduction in HER2 expression. In summary, our findings provide novel insights into the mechanism of pyrotinib in the treatment of HER2+ BC., Competing Interests: Declarations of interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Integrins, the receptors of the extracellular matrix, are critical in the proliferation and metastasis of cancer cells. GMI, a Ganoderma microsporum immunomodulatory protein, possesses anticancer and antivirus abilities. The object of this study is to investigate the role of GMI in the integrins signaling pathway in lung cancer cells that harbor the EGFR L858R/T790M double mutation and osimertinib-resistance. Liquid chromatography-mass spectrometry and western blot assay were used to investigate the effect of GMI on inhibiting the protein expressions of integrins in H1975 cells. The migration ability and xenograft tumor growth of H1975 were suppressed by GMI. To elucidate the role of the integrin family in lung cancer resistant to osimertinib (AZD-9291, Tagrisso), H1975 cells were used to establish the osimertinib-resistant cells, named H1975/TR cells. The expressions of Integrin αV and stemness markers were much higher in H1975/TR cells than in H1975 cells. GMI suppressed cell viability, tumor spheroid growth, and the expressions of integrin αV and β1 in H1975/TR cells. Furthermore, GMI suppressed the expressions of stemness markers and formation of tumor spheres via blocking integrin αV signaling cascade. This is the first study to reveal the novel function of GMI in constraining cancer stem cells and migration by abolishing the integrin αV-related signaling pathway in EGFR-mutated and osimertinib-resistant lung cancer cells., (© 2024 Wiley Periodicals LLC.)

Uterine leiomyoma (UL), commonly referred to as benign tumors, is characterized by excessive cell proliferation, extracellular matrix (ECM) accumulation, and the presence of stem cell-like properties. Nicotinamide adenine dinucleotide (NAD + ) metabolism, regulated in part by nicotinamide phosphoribosyltransferase (NAMPT), plays a crucial role in these pathological processes and has emerged as a potential therapeutic target. Additionally, redox signaling pathways are integral to the pathogenesis of UL, influencing the dynamics of NAD + metabolism. This study sought to elucidate the regulatory functions of NAMPT and NAD + metabolism, in conjunction with redox signaling, in the progression of UL, and to explore potential therapeutic strategies targeting these pathways. Evaluation of NAMPT expression in human UL tissues revealed a positive correlation between elevated NAMPT levels and increased ECM deposition, as well as the expression of stemness markers. The use of FK866 and nicotinamide (NAM), to inhibit NAMPT significantly suppressed UL cell viability and attenuated stem cell-like characteristics. Redox signaling pathways, including those associated with DNA damage, lysosomal function homeostasis, and redox-sensitive phagophore formation, were implicated in the regulation of ECM dynamics, particularly through ECM-targeted inhibition. This study highlights the pivotal roles of NAMPT, NAD + metabolism, and redox signaling in the pathophysiology of UL. Targeting NAMPT, particularly through the use of inhibitors FK866 and NAM, represents a promising therapeutic approach for mitigating UL progression by modulating redox and ECM dynamics. These findings offer novel insights into UL pathogenesis and establish NAMPT as a compelling target for future clinical investigation., Competing Interests: Declaration of competing interest The authors have declared no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Visfatin is expressed in the testis of chicken, humans and rodents; however, direct role of visfatin in the adult testis has not been studied. We investigated testicular responses after intra-testicular injection of FK866. The effects of visfatin inhibition were accessed at 24 hrs and 1 week post FK866 treatment. The testicular histoarchitecture were degenerated after 24 hrs of FK866 treatment along with supressed testosterone and proliferating markers and resumption in these parameters showed after 1 week. The expression of AR and ERα were down-regulated after 1 week of FK866 treatment. The expression of BCl2 was down-regulated along with a slight elevation of caspase3 after 24 hrs; however, both proteins still showed suppressed expression after 1 week. Furthermore, ERβ expression, 3βHSD, and 17βHSD were down-regulated in both groups compared to the control. Despite the down-regulation of some factors, the testicular proliferation and histoarchitecture showed resumption in the testis after 1 week of FK866 treatment. This could be due to increased testosterone secretion by suppressing aromatase expression. In conclusion, our result is the first report on the direct role of visfatin in the adult testis. Visfatin has a stimulatory role in testosterone synthesis and proliferation in the testis. Moreover, some deregulated factors in the testis after 1 week of FK866 treatment, despite normal histoarchitecture treatment, could be a compensatory mechanism after visfatin inhibitions., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2024 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

Death Receptor 5 (DR5) targeted therapies offer significant promise due to their pivotal role in mediating the extrinsic pathway of apoptosis. Despite DR5 overexpression in various malignancies and the potential of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), clinical applications of anti-DR5 monoclonal antibodies (mAbs) have been hampered by suboptimal outcomes potentially due to lack of receptor clustering. To address the limitation, we developed N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-based conjugates integrating multiple copies of DR5-targeting peptide (cyclic WDCLDNRIGRRQCVKL; cDR5) to enhance receptor clustering and apoptosis. Three conjugates with variable number of cDR5 were prepared and denoted as P H -cDR5 (high valence), P M -cDR5 (medium valence) and P L -cDR5 (low valence). Our studies in TRAIL-sensitive and resistant cancer cell lines demonstrated that the HPMA copolymer-peptide conjugates (P-cDR5) significantly improved DR5 receptor clustering and induced apoptosis effectively. In TRAIL-sensitive colon cancer cells (COLO205, HCT-116), P-cDR5 showed efficacy comparable to anti-DR5 mAb Drozitumab (DRO), but P-cDR5 outperformed DRO in TRAIL-resistant cells (HT-29), highlighting the importance of efficient receptor clustering. In COLO205 cells P M -cDR5 exhibited an IC50 of 94 pM, while P H -cDR5 had an even lower IC50 of 15 pM (based on cDR5 equivalent concentration), indicating enhanced potency of the multivalent HPMA copolymer-based system with a flexible polymer backbone in comparison with the IC 50 for TRAIL at 0.12 nM. Combining P-cDR5 with valproic acid, a histone deacetylase inhibitor, resulted in further enhancement of apoptosis inducing efficacy, along with destabilizing mitochondrial membranes and increased sensitivity of TRAIL-resistant cells. These findings suggest that attaching multiple cDR5 peptides to a flexible water-soluble polymer carrier not only overcomes the limitations of previous designs but also offers a promising avenue for treating resistant cancers, pointing toward the need for further preclinical exploration and validation of this innovative strategy., Competing Interests: Declaration of competing interest The authors declare no competing financial interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Introduction: Osimertinib resistance inevitably occurs in EGFR mutated NSCLC. We aimed to identify resistance mechanisms (RM) using paired plasma and tumor samples in patients that progressed on 2nd/3rd line osimertinib., Methods: From 09 - 2019 to 02 - 2021, 51 patients were enrolled. Plasma sequencing used AVENIO Expanded Panel (research use only), tumor biopsies underwent DNA and RNA sequencing and histological evaluation. Sequencing was regarded successful when the driver mutation was confirmed with a variant allele frequency of ≥0.10%. Concordance between modalities was calculated for the driver mutation and RMs covered in both modalities. The Molecular Tumor Board formulated a treatment advice., Results: The driver mutation was detected in 42/51 plasma samples (82%) and in 50/51 tumor samples (98%), concordance rate was 80%. In 41/51 (80%) patients ≥1 RM was identified. Thirty-two RMs covered in both modalities were found in plasma (61.5%), 39 in tumor (75%), nineteen in both. RM concordance rate was 36.5%., Conclusion: Combined analysis of plasma and tumor samples post 2nd/3rd line osimertinib identifies additional RMs regardless of the comparative approach used. Plasma sequencing identified 61.5% of RMs, tumor analysis identified 75%. Combined, they provide a superior overview of osimertinib resistance, enabling more tailored treatment options., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Baas reports a grant from BMS, unrestricted to the hospital, as well as consulting fees paid to the hospital by BMS, MSD, AMheart, payment or honoraria for lectures, presentations, speakers bureaus, manuscript writing or educational events to the hospital from BMS. Burgers reports a grant from MSD as support for an investigator initiated study, payment or honoraria for lectures, presentations, speakers bureaus, manuscript writing or educational events to the hospital from MSD, participation on a data safety monitoring board or advisory board with Roche, paid to the hospital, and receipt of free drugs for an investigator initiated study from MSD. Hashemi reports grants from AstraZeneca, Boehringer Ingelheim, BMS, Lilly, Janssen, GSK, MSD, Novartis, Roche and Takeda, all to the institution. De Langen reports grants from BMS, grants from MSD, grants from Boehringer, non-financial support from Merck Serono, grants from AstraZeneca, non-financial support from Roche, outside the submitted work. Monkhorst reports consulting fees from Lilly, Bayer and Amgen, all paid to the institution. Smit reports consulting fees from Eli Lilly, AstraZeneca, MSD, Merck, BMS, DSI, Takeda and Boehringer Ingelheim, all paid to the institution, payment or honoraria for lectures, presentations, speakers bureaus, manuscript writing or educational events from Boehringer Ingelheim, and participation on a data safety monitoring board or advisory board with DSI. Theelen reports grants from Sanofi/Regeneron and MSD to the institution. Boelens, Bosch, Buikhuisen, van den Broek, Jebbink, van der Noort, Steinbusch, Ruiter, Vermeulen, van der Wel declare no potential conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Acquired resistance to first-line treatments in various cancers both promotes cancer recurrence as well as limits effective treatment. This is true for epidermal growth factor receptor (EGFR) mutations, for which secondary EGFR mutations are one of the principal mechanisms conferring resistance to the covalent inhibitor osimertinib. Thus, it is very important to develop a deeper understanding of the secondary mutational resistance mechanisms associated with EGFR mutations arising in tumors treated with osimertinib to expedite the development of innovative therapeutic drugs to overcome acquired resistance. This work uses all-atom molecular dynamics (MD) simulations to investigate the conformational variation of two reported EGFR mutants (L858R/L718Q and L858R/L792H) that resist osimertinib. The wild-type EGFR kinase domain and the L858R mutant are used as the reference. Our MD simulation results revealed that both the L718Q and L792H secondary mutations induce additional hydrogen bonds between the residues in the active pocket and the residues with the water molecules. These additional hydrogen bonds reduce the exposure area of C797, the covalent binding target of osimertinib. The additional hydrogen bonds also influence the binding affinity of the EGFR kinase domain by altering the secondary structure and flexibility of the amino acid residues in the domain. Our work highlights how the two reported mutations may alter both residue-residue and residue-solvent hydrogen bonds, affecting protein binding properties, which could be helpful for future drug discovery., (© 2024 Wiley Periodicals LLC.)

EGFR tyrosine kinase inhibitors (EGFR-TKIs) are the first-line treatment for EGFR-mutant non-small cell lung cancer (NSCLC) patients, which remarkably improve the clinical outcomes. However, drug resistance has greatly impaired the efficacy of EGFR-TKIs and contributes to cancer treatment failure. DUSP1, a negative regulator of MAPK signaling pathway, was discovered to mediate drug resistance in multiple types of cancers. Our study aimed to explore the role of DUSP1 in NSCLC cell resistance to osimertinib, a third-generation EGFR-TKI. Human NSCLC cell lines PC-9 and HCC827 were exposed to increasing concentrations of osimertinib for over 6 months to generate osimertinib resistant cells (PC-9-OR and HCC827-OR). The viabilities of osimertinib-resistant and parental sensitive NSCLC cells in response to osimertinib stimulation were detected by MTS assay and the IC50 values for osimertinib were obtained. The differentially expressed genes in osimertinib-resistant and sensitive NSCLC cells were identified by analyzing the GEO dataset GSE106765 using bioinformatic tools. DUSP1 expression was knocked down by using the short hairpin RNAs (shRNAs). Then, the effects of DUSP1 silencing on osimertinib-resistant and sensitive NSCLC cell resistance to osimertinib, viability, proliferation and apoptosis were assessed through loss-of-function experiments. The expression of key molecules (JNK, ERK, and p38 MAPK) in the MAPK signaling pathway was detected through western blotting analysis. DUSP1 was overexpressed in osimertinib-resistant NSCLC cells versus parental sensitive cells. DUSP1 silencing attenuated the resistance of NSCLC cells to osimertinib. DUSP1 silencing markedly inhibited osimertinib-resistant and sensitive NSCLC cell proliferation but enhanced cell apoptosis. Mechanically, DUSP1 knockdown increased phosphorylated-JNK, ERK, and p38 MAPK levels in NSCLC cells. Treatment with SB203580, the p38 MAPK inhibitor, reversed the effects of DUSP1 silencing on osimertinib-resistant NSCLC cell resistance to osimertinib, cell proliferation and apoptosis. DUSP1 downregulation restores the sensitivity of NSCLC cells to osimertinib via activating the MAPK signaling pathway., Competing Interests: Declarations. Competing Interests: The authors declare that they have no competing interests. Ethical Approval: Our study did not require an ethical board approval because it did not contain human or animal trials., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Overcoming clinical resistance to osimertinib mediated by the tertiary C797S mutation remains an unmet medical need. To date, there are no effective drugs that have been approved for patients who harbor EGFR T790M/C797S mutations. Herein, we applied a structure-based drug design strategy to discover a series of potent and selective diaminopyrimidine macrocycles as novel EGFR T790M/C797S inhibitors. The representative compound 21v potently inhibited EGFR 19del/T790M/C797S and EGFR L858R/T790M/C797S mutants with IC 50 values of 2.3 nM and 12.5 nM, respectively, and exhibited antiproliferative activity against Ba/F3-EGFR 19del/T790M/C797S and Ba/F3-EGFR L858R/T790M/C797S cells with IC 50 values of 41 and 52 nM, respectively. Further, 21v inhibited proliferation of the EGFR 19del/T790M/C797S mutant PC-9-OR NSCLC cell line with an IC 50 value of 56 nM and displayed selectivity over parental Ba/F3 and A431 cells. Moreover, 21v exhibited antitumor efficacy in a Ba/F3-EGFR 19del/T790M/C797S xenograft model. This study provides a promising macrocyclic lead for anticancer drug discovery overcoming EGFR C797S mutation mediated resistance in NSCLC patients.

Background: Overcoming resistance to Osimertinib in epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer (NSCLC) is clinically challenging because the underlying mechanisms are not fully understood. The murine double minute 2 (MDM2) has been extensively described as a tumor promotor in various malignancies, mainly through a negative regulatory machinery on the p53 tumor suppressor. However, the significance of MDM2 on the sensitivity to Osimertinib has not been described., Methods: Osimertinib resistant cells were generated by standard dose escalation strategy and individual resistant clones were isolated for MDM2 testing. The MDM2 and its mutant constructs (ΔPBD, ΔRING, C464A) were introduced into PC-9, HCC827 and H1975 cells and evaluated for the sensitivity to Osimertinib by MTT assay, colony formation, EdU assay and TUNEL assay. MDM2 expression in resistant cells was manipulated by pharmacological and molecular approaches, respectively. Proteins that were implicated in PI3K/Akt, MAPK/Erk and apoptosis signaling were measured by Western blot analysis. Candidate proteins that interacted with MDM2 were captured by immunoprecipitation and probed with indicated antibodies., Results: In comparison with parental PC-9 cells, the PC-9 OR resistant cells expressed high level of MDM2. Ectopic expression of MDM2 in PC-9, HCC827 and H1975 sensitive cells generated an Osimertinib resistant phenotype, regardless of p53 status. MDM2 promoted resistance to Osimertinib through a PI3K/Akt and MAPK/Erk-independent machinery, in contrast, MDM2 selectively stabilized MCL-1 protein to arrest Osimertinib-induced cancer cell apoptosis. Mechanistically, MDM2 acted as a E3 ligase to ubiquitinate FBW7, a well-established E3 ligase for MCL-1, at Lys412 residue, which resulted in FBW7 destruction and MCL-1 stabilization. Targeting MDM2 to augment MCL-1 protein breakdown overcame resistance to Osimertinib in vitro and in vivo. Finally, the clinical relevance of MDM2-FBW7-MCL-1 regulatory axis was validated in mouse xenograft tumor model and in NSCLC specimen., Conclusion: Overexpression of MDM2 is a novel resistant mechanism to Osimertinib in EGFR mutant NSCLC. MDM2 utilizes its E3 ligase activity to provoke FBW7 destruction and sequentially leads to MCL-1 stabilization. Cancer cells with aberrant MDM2 state are refractory to apoptosis induction and elicit a resistant phenotype to Osimertinib. Therefore, targeting MDM2 would be a feasible approach to overcome resistance to Osimertinib in EGFR mutant NSCLC., Competing Interests: Declarations Consent for publication Written consent was obtained from each participate. Ethical approval and consent to participate Animal care and experiments were reviewed and approved by the Institutional Animal Care and Use Committee (#2023JLHGZRDWLS-00032). The handling and processing of patient tumor samples were done in accordance with Institutional Ethics Committee Review Board Approved protocols (#2023DZGZR-030). Competing interest Wei Liu is an employee of Liaoning Kanghui Biotechnology. Liu analyzed the NGS data and did not have access to interpret the bioinformatic and experimental results. The remaining authors did not have any financial association with Liaoning Kanghui Biotechnology or other biomedical companies., (© 2024. The Author(s).)

Ethnopharmacological Relevance: Acquired resistance to osimertinib limits its clinical efficacy in non-small cell lung cancer (NSCLC) with EGFR mutations. The widespread recognition of Taxus chinensis var. Mairei (Lemée et Lévl) Cheng et L.K. Fu (Chinese yew) as a natural anti-cancer medication is well-established. However, the specific contribution of Taxus chinensis var. Mairei (Lemée et Lévl) Cheng et L.K. Fu in addressing resistance to osimertinib is still uncertain., Aim of the Study: Based on the biological behaviors and lipid metabolism, we investigated whether aqueous extract of Taxus chinensis var. Mairei (Lemée et Lévl) Cheng et L.K. Fu (AETC) could enhance the antitumor effect of osimertinib in NSCLC with an investigation on the precise mechanisms., Materials and Methods: The effect of AETC on enhancing osimertinib sensitivity was assessed via cell viability measurements, levels of reactive oxygen species (ROS), apoptosis, and lipid levels. Western blotting was used to verify the mechanisms of AETC responsible for overcoming the resistance to osimertinib via ERK1/2 overexpression and knockdown models. In vivo validation was conducted using subcutaneous xenografts from osimertinib-resistant cells in nude mice., Results: Osimertinib-resistant cells exhibited altered cholesterol biosynthesis, which was induced by ERK1/2 activation. The combination of AETC and osimertinib can synergistically decrease the levels of ROS in cells, enhance apoptosis, and inhibit the growth of osimertinib-resistant cells. Mechanistic experiments demonstrated that AETC can downregulate the key regulators of cholesterol biosynthesis by regulating ERK1/2, inhibiting the endogenous synthesis rate of cholesterol, and suppressing the level of lipids in osimertinib-resistant cells and xenograft tumors when combined with osimertinib, ultimately reversing resistance to osimertinib., Conclusions: The resistance to osimertinib is significantly influenced by cholesterol biosynthesis, highlighting its pivotal role in this context. AETC can enhance osimertinib sensitivity via ERK/SREBP-2/HMGCR-mediated cholesterol biosynthesis. These results provide a promising therapeutic target and potential treatment option for resistance to osimertinib., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The effective treatment of inflammatory diseases, particularly their chronic forms, is a key task of modern medicine. Herein, we report the synthesis and evaluation of biocompatible polymer conjugates based on N-2-(hydroxypropyl)methacrylamide copolymers enabling the controlled release of acetylsalicylic acid (ASA)-based anti-inflammatory drugs under specific stimuli. All polymer nanotherapeutics were proposed as water-soluble drug delivery systems with a hydrodynamic size below 10 nm ensuring suitability for the parenteral application and preventing opsonization by the reticuloendothelial system. The nanotherapeutics bearing an ester-bound ASA exhibited long-term release of the ASA/salicylic acid mixture, while the nanotherapeutics carrying salicylic acid hydrazide (SAH) ensured the selective release of SAH in the acidic inflammatory environment thanks to the pH-sensitive hydrazone bond between the polymer carrier and SAH. The ASA- and SAH-containing nanotherapeutics inhibited both cyclooxygenase isoforms and/or the production of pro-inflammatory mediators. Thanks to their favorable design, they can preferentially accumulate in the inflamed tissue, resulting in reduced side effects and lower dosage, and thus more effective and safer treatment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Evaluating the steady-state protein level of the EGFR in live cells presents significant challenges compared to measuring its kinase activity. Traditional testing methods, such as immunoblotting, ELISA, and immunofluorescence assays, are generally restricted to fixed cells or cell lysates. Despite their utility, these methods are cumbersome and provide only intermittent snapshots of EGFR levels at specific time points. With emerging trends in drug development shifting toward engineering novel agents that promote protein degradation, rather than simply inhibiting kinase activity, a tool that enables real-time, quantitative detection of drug effects in live cells could catalyze advances in the field. Such an innovation would expedite the drug development process, enhancing the translation of research findings into effective, patient-centered therapies. The NanoLuc-EGFR cell line, created through CRISPR genome editing, allows for the continuous tracking and analysis of EGFR protein levels and their degradation within live cells. This approach provides quantitative monitoring of protein dynamics in real time, offering insights that go beyond absolute protein levels to include aspects such as protein stability and degradation rate. Using this cell line model, we observed that AT13387 and H84T BanLec induce EGFR degradation in A549-HiBiT cells, with the results confirmed by immunoblotting. In contrast, Erlotinib, Osimertinib, and Cetuximab inhibit EGFR phosphorylation without altering total EGFR levels, as validated by the HiBiT luciferase assay. The NanoLuc-EGFR cell line marks a significant advancement in understanding protein regulation and serves as an instrumental platform for investigating targeted therapies that modulate protein kinases, especially those that induce protein degradation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Patient-derived xenograft (PDX) is currently considered a preferred preclinical model to evaluate drug sensitivity, explore drug resistance mechanisms, and select individualized treatment regimens., Methods: Histopathological examination, immunohistochemistry and whole-exome sequencing confirmed similarity between our PDX tumors and primary tumors in terms of morphology and genetic characteristics. The drug reactivity of the PDX tumor was validated in vivo. The mechanisms of acquired resistance to Osimertinib PDX tumors were investigated by WES and WB., Results: We successfully established 13 NSCLC-PDXs derived from 62 patients, including eight adenocarcinomas, four squamous-cell carcinoma, and one large-cell neuroendocrine carcinoma. Histological subtype and clinical stage were significant factors affecting the successful PDXs establishment. The treatment responses to conventional chemotherapy in PDXs were entirely consistent with that of their corresponding patients. According to the genetic status of tumors, more appropriate targeted agents were selected in PDXs for their corresponding patients as alternative treatment options. In addition, a PDX model with acquired resistance to osimertinib was induced, and the overactivation of RAS mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase (ERK) signaling pathway caused by the dual-specificity phosphatase 6 (DUSP6) M62I mutation was found to play a key role in the development of osimertinib resistance. Trametinib, a specific inhibitor of the MAPK-ERK pathway significantly slowed down the tumor growth in osimertinib-resistant PDX models, providing an alternative treatment in patients after osimertinib failure., Competing Interests: Declarations Ethics approval and consent to participate This study was approved by the ethics committee board of the First Affiliated Hospital of Chongqing Medical University (Approval number: 2022-K432), and was registered with the Chinese Clinical Trial Registry (No. ChiCTR2200066004). All animal studies were carried out in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals, and approved by the Animal Ethics and Experimental Committee of the Chongqing Medical University. Informed consent was obtained from all subjects involved in the study. Consent for publication Consent for publication was obtained from the participants. Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

The inevitable acquired resistance to osimertinib (AZD9291), an FDA-approved third-generation EGFR tyrosine kinase inhibitor (EGFR-TKI) for the treatment of patients with advanced non-small cell lung cancer (NSCLC) harboring EGFR activating or T790M resistant mutations, limits its long-term clinical benefit. Telomere maintenance via telomerase reactivation is linked to uncontrolled cell growth and is a cancer hallmark and an attractive cancer therapeutic target. Our effort toward understanding the action mechanisms, including resistance mechanisms, of osimertinib has led to the identification of a novel and critical role in maintaining c-Myc-dependent downregulation of hTERT, a catalytic subunit of telomerase, and subsequent inhibition of telomerase/telomere and induction of telomere dysfunction in mediating therapeutic efficacy of osimertinib. Consequently, osimertinib combined with the telomere inhibitor, 6-Thio-dG, which is currently tested in a phase II trial, effectively inhibited the growth of osimertinib-resistant tumors, regressed EGFRm NSCLC patient-derived xenografts, and delayed the emergence of acquired resistance to osimertinib, warranting clinical validation of this strategy to manage osimertinib acquired resistance., (© 2024 Chen et al.)

Surgical resection followed by radiotherapy (RT) is recommended for malignant meningioma, but poor outcome is unavoidable. To improve the efficacy of RT in malignant meningioma, a targeted radiosensitizer can be added. Nicotinamide phosphoribosyltransferase (NAMPT), highly expressed in high-grade meningiomas, may play a role in determining the radioresponse. Herein, we evaluated the impact of NAMPT inhibition on radiosensitivity in malignant meningioma in vivo and in vitro. IOMM-Lee and TTMM705 cells were treated with NAMPT inhibition (FK866 or shRNA NAMPT) before irradiation. The subsequent clonogenic assay demonstrated significantly increased radiosensitivity. Combination treatment with FK866 and irradiation significantly increased the number of G2/M-phase cells, percentage of apoptotic cells, and γ-H2A.X level compared with FK866 or RT alone. We examined the effect of NAMPT inhibition on NMI and p53 expression in IOMM-Lee and TTMM705 cells. NAMPT inhibition by FK866 and shRNA treatment increased NMI, p53, CDKN1A and BAX expression. Additionally, we assessed the efficacy of FK866/RT combination treatment in vivo. The combination treatment exhibited increased antitumor efficacy compared with either treatment alone. The Ki67 level was significantly lower, and the p53 and γ-H2A.X levels were significantly higher in the combination treatment group than in the other three groups. In conclusion, these results indicate that FK866 improves radiosensitivity in malignant meningioma, an effect that may be attributed to the increase in p53 expression., (©2024 American Association for Cancer Research.)

Resistance to osimertinib represents a significant challenge for the successful treatment of non-small cell lung cancer (NSCLC) harboring activating mutations in EGFR. N6-methyladenosine (m6A) on mRNAs is critical for various biological processes, yet whether m6A regulates osimertinib resistance of NSCLC remains unknown. In this study, we demonstrated that developing osimertinib-resistant phenotypes depends on m6A reduction resulting from downexpression of m6A methyltransferase METTL14 in EGFR-mutant NSCLCs. Both in vitro and in vivo assays showed that specific knockdown of METTL14 was sufficient to confer osimertinib resistance and that elevated expression of METTL14 rescued the efficacy of osimertinib in the resistant NSCLC cells. Mechanistically, METTL14 promoted m6A methylation of pro-apoptotic Bim mRNA and increased Bim mRNA stability and expression, resulting in activating the Bim-dependent pro-apoptotic signaling and thereby promoting osimertinib-induced cell apoptosis. Analysis of clinical samples revealed that decreased expression of METTL14 was observed in osimertinib-resistant NSCLC tissues and significantly associated with a poor prognosis. In conclusion, our study reveals a novel regulatory mechanism by which METTL14-mediated m6A methylation of Bim mRNA inhibited osimertinib resistance of NSCLC cells. It offers more evidences for the involvement of m6A modification in regulation of osimertinib resistance and provides potential therapeutic targets for novel approaches to overcome the tolerance of osimertinib and other EGFR tyrosine kinase inhibitors. Implications: This study offers more evidences for the involvement of METTL14-mediated N6-methyladenosine modification in regulation of osimertinib resistance and provides potential therapeutic targets for novel approaches to overcome the tolerance of osimertinib and other EGFR tyrosine kinase inhibitors., (©2024 American Association for Cancer Research.)

Non-small cell lung cancer (NSCLC) patients without targetable driver mutation have limited treatment options. In this study, we aimed to explore a new therapeutic strategy by using established nine patient-derived xenograft (PDX) and two-dimensional (2D) /3D culture models with specific genetic alternations. The gene mutations and copy number aberrations were detected by next-generation sequencing and confirmed using polymerase chain reaction (PCR) followed by DNA sequencing, and genomic DNA quantitative PCR. Protein expression was evaluated by immunohistochemistry. Drug sensitivities of PDX/2D/3D models were evaluated by in vivo and in vitro antitumor assays. RNA interference was performed to silence gene expression. Our study found that 44.4 % (4/9) of cases had CDKN2A homozygous deletion (homdel), while 33.3 % (3/9) had CDKN2B homdel. Additionally, 22.2 % (2/9) had amplification (amp) in wildtype CDK4, 44.4 % (4/9) in CDK6, and 44.4 % (4/9) in EGFR. Among the cases, 77.8 % (7/9) lacked CDKN2A, and 33.3 % (3/9) had high CDK4, CDK6, and EGFR had high protein expression. Moreover, 33.3 % (3/9) had KRAS mutations, and 66.7 % (6/9) had TP53 mutations. Antitumor activity of osimertinib plus palbociclib was assessed in four PDX/2D/3D models, two of which had simultaneous EGFR amp and CDKN2A/2B homdel. The data showed that NSCLC with EGFR amp and CDKN2A/2B homdel were sensitive to combined drugs. Additional oncogenic KRAS mutation reduced the drug's antitumor effect. EGFR amp is responsible for osimertinib sensitivity. Osimertinib plus palbociclib effectively treat NSCLC with wildtype EGFR and CDK6 amp and CDKN2A/2B homdel in the absence of oncogenic KRAS mutation., Competing Interests: Declaration of competing interest The authors declare no conflict-of-interest., (Copyright © 2024. Published by Elsevier Inc.)

Spreading depolarization (SD) describes the near-complete depolarization of central nervous system (CNS) neural cells as a consequence of chemical, electrical, or metabolic perturbations. It is well established as the central mechanism underlying insect coma and various mammalian neurological dysfunctions. Despite significant progress in our understanding, the question remains: which cation channel, if any, generates SD in the CNS? Previously, we speculated that the sodium-potassium ATPase (NKA) might function as a large-conductance ion channel to initiate SD in insects, potentially mediated by a palytoxin (PLTX)-like endogenous activator. In the current study, we evaluate the effectiveness and properties of PLTX as an SD initiator in Locusta migratoria . Whereas bath-applied PLTX failed to ignite SD, direct injection into the neuropil triggered SD in 57% of the preparations. Notably, PLTX-induced SD onset was significantly more rapid compared with ouabain (OUA) injection and azide controls, though their electrophysiological features remained similar. Furthermore, PLTX-induced SD was recoverable and resulted in a greater frequency of repetitive SD events compared with ouabain. Surprisingly, prior PLTX treatment disrupted the onset and recovery of subsequent SD evoked by other means. PLTX injection could attenuate the amplitude and even completely inhibit the onset of azide-induced SD at higher doses. These results show that PLTX can trigger repetitive and reversible SD-like events in locusts and simultaneously interfere with anoxic SD occurrence. We suggest that the well-documented NKA pump conversion into an open nonselective cationic channel is a plausible mechanism of SD activation in the locust CNS, warranting additional investigations. NEW & NOTEWORTHY Spreading depolarization (SD) is a critical mechanism underlying central nervous system (CNS) shutdown and injury under stress, yet the initiating ion channel remains unknown. Here, we used the marine poison palytoxin (PLTX), which converts the sodium-potassium ATPase (NKA) into an open channel, to initiate SD in intact locust CNS. We show for the first time that PLTX-induced SD is rapid and recoverable in vivo, providing support that NKA conversion to a channel may be the SD-initiating mechanism.

Generally, oligolysine has poor antibacterial effect and almost no antibacterial activity. Herein, low cost and easily available oligolysines were chosen to prepare injectable antibacterial hydrogel (PVAL-gel) for wound healing. The hydrogel network was formed by cross-linking vanillin acrylate-N, N-dimethylacrylamide copolymer P(VA-co-DMA), oligolysine and adipate dihydrazide through Schiff base bond. The obtained hydrogel PVAL-gel exhibited not only excellent self-healing capability and injectability, but also the efficient contact antibacterial ability and good inhibitory effects on E.coli and S.aureus. In vitro, 99.9 % of pathogenic bacteria was killed within 160 min. Furthermore, the injectable PVAL-gel could rapidly eradicate bacteria in infected wounds and notably enhance the healing of full-thickness skin wounds. Therefore, PVAL-gel is expected to be used as a high-end dressing for the treatment of infected skin wounds, which can promote wound healing., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Third-generation EGFR tyrosine kinase inhibitor (TKIs) have revolutionized the treatment landscape for patients with non-small cell lung cancer (NSCLC) harboring EGFR activating mutations, with improved long-term outcomes compared to first-generation TKIs. Nevertheless, disease progression inevitably occurs, limiting osimertinib long-term efficacy. Indeed, the molecular biology underlying acquired resistance to first-line osimertinib is multifaceted and includes the emergence of on-target and off-target alterations. EGFR-C797S mutation represents the most frequent mechanism of on-target resistance and hinders drug binding to the target site. EGFR-independent resistance includes the activation of alternative signaling pathways, such as MET amplification and HER2 mutations, and histological transformation. In this setting, chemotherapy is the current therapeutic option, with modest clinical outcomes. Therefore, the development of novel therapeutic strategies to overcome resistance to osimertinib is a major challenge. In this setting, fourth-generation TKIs are emerging as an interesting therapeutic option to overcome on-target resistance. Preclinical drug development has led to the discovery of thiazole-amid inhibitors, which activity is mediated by the allosteric inhibition of EGFR, resulting in high specificity towards mutant-EGFR. Early phase 1/2 clinical trials are ongoing to elucidate their activity also in the clinical setting. Aim of this review is to provide a state-of-the-art analysis on preclinical development of fourth-generation EGFR-TKIs and promising preliminary clinical data., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Introduction: EGFR tyrosine kinase inhibitor (EGFR-TKI)-sensitizing and -resistance mutations may be detected in plasma through circulating tumor DNA (ctDNA). Circulating tumor DNA level changes reflect alterations in tumor burden and could be a dynamic indicator of treatment effect. This analysis aimed to determine whether longitudinal EGFR-mutation ctDNA testing could detect progressive disease (PD) before radiologic detection., Methods: This was a retrospective, exploratory ctDNA analysis in two phase 3 trials (FLAURA, NCT02296125; AURA3, NCT02151981). Patients had treatment-naïve (FLAURA) or EGFR-TKI pre-treated (AURA3) advanced NSCLC with EGFR mutations and on-study PD (RECIST [Response Evaluation Criteria in Solid Tumors]), with a baseline ctDNA result and EGFR-mutation ctDNA monitoring beyond Cycle 3 Day 1. Patients received osimertinib versus comparator EGFR-TKIs (FLAURA) or chemotherapy (AURA3). Outcomes included time from ctDNA PD to RECIST PD and the first subsequent treatment (FLAURA only)., Results: Circulating tumor DNA PD preceded or co-occurred with RECIST-defined PD in 93 out of 146 patients (64%) in FLAURA and 82 out of 146 patients (56%) in AURA3. Median time from ctDNA PD to RECIST-defined PD (mo) was 3.4 and 2.6 in the osimertinib and comparator EGFR-TKI arms (FLAURA) and 2.8 and 1.5 in the osimertinib and chemotherapy arms (AURA3). In FLAURA, the median time from ctDNA PD to the first subsequent treatment (mo) was 6.0 and 4.7 in the osimertinib (n = 51) and comparator EGFR-TKI arms (n = 70)., Conclusions: Among patients with EGFR mutation-positive advanced NSCLC receiving EGFR-TKI or chemotherapy with ctDNA data and RECIST-defined PD, ctDNA PD preceded/co-occurred with RECIST-defined PD in approximately 60% of cases. Longitudinal ctDNA monitoring may detect PD before radiologic PD., Competing Interests: Disclosure Dr. Gray reports participation on the board of directors for IASLC; consulting or advisory fees from AbbVie, AstraZeneca, Blueprint Medicines, Daiichi-Sankyo, EMD Serono, Gilead Sciences, Inc., IDEOlogy Health, Janssen Scientific Affairs, LLC, Jazz Pharmaceuticals, Loxo Oncology Inc., Merck & Co., Novartis, OncoCyte, Spectrum ODAC, Takeda, and Triptych Health Partners; research grants from AstraZeneca, Boehringer Ingelheim, Bristol Myers Squibb, Genentech, G1 Therapeutics, Ludwig Institute of Cancer Research, Merck & Co., Novartis, and Pfizer; employment with Moffitt Cancer Center; holding a leadership role as SWOG Lung Committee Chair and ASCO Education Committee Ex-Chair; and payments/honoraria from AbbVie, AstraZeneca, Blueprint Medicines, Daiichi-Sankyo, EMD Serono, Gilead Sciences, Inc., IDEOlogy Health, Janssen Scientific Affairs, LLC, Jazz Pharmaceuticals, Loxo Oncology Inc., Merck & Co., Novartis, OncoCyte, Spectrum ODAC, Takeda, and Triptych Health Partners. Dr. Markovets reports employment with AstraZeneca. Dr. Reungwetwattana reports honoraria from Amgen, AstraZeneca, Bristol Myers Squibb, Merck Sharp & Dohme, Novartis, Roche, Takeda, Yuhan, and Zuellig; and research grants from AstraZeneca, Novartis, Roche, and Yuhan. Dr. Majem reports honoraria from Amgen, AstraZeneca, BeiGene, Bristol Myers Squibb, Helsinn Therapeutics, Immedica, Janssen, Merck Sharp & Dohme, Novartis, Pfizer, Pierre Fabre, Roche, Sanofi, and Takeda; research grants from AstraZeneca, Bristol Myers Squibb, and Roche; and travel support from AstraZeneca, Eli Lilly, Merck Sharp & Dohme, Pfizer and Roche. Dr. Cho reports an advisory role for Bridge Biotherapeutics, Inc., Cyrus Therapeutics, Inc., Guardant Health, Joseah BIO, and KANAPH Therapeutic, Inc.; participation on boards of directors for Gencurix, Inc. and Interpark Bio Convergence Corp.; consulting fees from AstraZeneca, Blueprint Medicines, Boehringer Ingelheim, Bristol Myers Squibb, Eli Lilly, Janssen, Medpacto, Merck Sharp & Dohme, Novartis, Ono Pharma Co., Ltd., Pfizer, Roche, Takeda, and Yuhan; employment with Yonsei University College of Medicine; being a founder of DAAN Biotherapeutics; grants or funding from AbbVie, AstraZeneca, Bayer, Blueprint Medicines, Champions Oncology, Dizal Pharmaceutical Co., Ltd., Dong-A ST, Eli Lilly, GI Innovation, Interpark Bio Convergence Corp., Janssen, Medpacto, Merck Sharp & Dohme, MOGAM Institute, Novartis, Ono Pharma Co., Ltd., and Yuhan; royalties from Champions Oncology; and stock or stock options with Bridge Biotherapeutics, Inc., Cyrus Therapeutics, Inc., Gencurix, Inc., KANAPH Therapeutic, Inc., Interpark Bio Convergence Corp., and TheraCanVac, Inc. Dr. Chewaskulyong reports honoraria and research grants from AstraZeneca. Dr. John reports consulting fees from Amgen, AstraZeneca, Boehringer Ingelheim, Bristol Myers Squibb, Gilead Sciences, Inc., Ignyta, Merck Sharp & Dohme, Merck KGaA, Novartis, Pfizer, PharmaMar, Roche/Genentech, and Specialised Therapeutics; honoraria from AstraZeneca/MedImmune, Bristol Myers Squibb, Merck Sharp & Dohme, and Roche/Genentech; and travel support from AstraZeneca, Boehringer Ingelheim, Bristol Myers Squibb, Merck Sharp & Dohme, and Roche. Dr. Sebastian reports participation on an advisory board for Amgen; consulting fees from Amgen, AstraZeneca, Boehringer lngelheim, Bristol Myers Squibb, GSK, Johnson & Johnson, Eli Lilly, Merck Sharp & Dohme, Merck Serono, Novartis, Roche, and Takeda; grants or contracts from AstraZeneca; honoraria from Amgen, AstraZeneca, BioNTech, Boehringer lngelheim, Bristol Myers Squibb, CureVac, Daiichi-Sankyo, GSK, Johnson & Johnson, Eli Lilly, Merck Sharp & Dohme, Merck Serono, Novartis, Pfizer, Roche, and Takeda; and travel support from Bristol Myers Squibb and Pfizer. Drs Todd, Rukazenkov, Barrett, and Chmielecki report employment and holding stock or stock options with AstraZeneca. Dr. Ramalingam reports personal fees from Amgen, AstraZeneca, Bristol Myers Squibb, Genentech, GSK, Merck, Takeda, and Tesaro and other non-financial support from AstraZeneca. Dr. Hartmaier reports being an inventor on patent US11066709B2, and employment and holding stock or stock options with AstraZeneca. The remaining authors declare no conflict of interest., (Copyright © 2024 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.)

Background: Osimertinib shows higher effectiveness than first-generation epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) in the initial treatment of EGFR-mutated non-small cell lung cancer. However, its superiority in terms of overall survival in the Asian population, especially Japanese patients, remains uncertain., Objective: To evaluate the survival benefit of osimertinib over other EGFR-TKIs in Japanese patients, using real-world data. METHODS : As part of the Tokushukai REAl-world Data project, a retrospective multi-institutional study across 46 hospitals in Japan was conducted to evaluate the overall survival of patients with advanced EGFR-mutated non-small cell lung cancer using propensity score matching. The study involved patients receiving osimertinib as the first-line treatment (1L-Osi), those initially treated with other EGFR-TKIs (1L-non-Osi), and those receiving osimertinib after initial EGFR-TKI treatment (2L/later-Osi) between April 2010 and December 2022 and followed up until April 2023., Results: Among 1062 Japanese patients with EGFR-mutated non-small cell lung cancer, 416 (39.2%) received 1L-Osi, while 646 (60.8%) received 1L-non-Osi, including 139 (13.1%) who received 2L/later-Osi. Within these groups, 416 (39.2%), 293 (27.6%), and 75 (7.1%) patients received first-line EGFR-TKI treatment post-osimertinib approval as a later-line treatment in Japan (March 2016). After propensity score matching, the overall survival of the 1L-Osi group was comparable to that of the 1L-non-Osi group in the post-March 2016 subset (n = 283, 42.0 vs 42.4 months). Similar trends were observed in the Del19 and L858R subgroups. The median overall survival of the 2L/later-Osi group was notably long: 60.2 months post-March 2016 (n = 75). A subgroup analysis based on initial EGFR-TKI treatment in the 1L-non-Osi and 2L/later-Osi groups revealed no significant differences among the gefitinib, erlotinib, and afatinib groups., Conclusions: Based on real-world data, osimertinib did not show a significant improvement in overall survival compared to other EGFR-TKIs as a first-line treatment for EGFR-mutated advanced non-small cell lung cancer in the Japanese (Asian) population., Clinical Trial Registration: This study was registered at the University Hospital Medical Information Network Clinical Trials Registry on 9 March, 2023 (identification UMIN000050552)., Competing Interests: Declarations Funding No external funding was used in the preparation of this article. Conflict of interest Makoto Hibino received research funding from and engaged in collaborative research with Shionogi & Co., Ltd. outside the submitted work. Tomoya Fukui received speakers’ bureau fees/honoraria from AstraZeneca, Boehringer Ingelheim, Chugai Pharm, Eli Lilly, Nippon Kayaku, Novartis Pharma, Ono Pharm, Pfizer, and Taiho Pharm outside the submitted work. Imamura received speakers’ bureau fees/honoraria from Bayer, Bristol-Myers Squibb, Daiichi-Sankyo, Pfizer, and Ono Pharm outside the submitted work. Minami received speakers’ bureau fees/honoraria from Daiichi-Sankyo and Ono Pharm; research funding from Astelas-Amgen Biopharma, Bayer, Bristol-Myers Squibb, Chugai, Daiichi-Sankyo, Incite, Novartis, Ono Pharm, Pfizer, and Rakuten Medical; and scholarship donations from Bayer, Chugai, Daiichi-Sankyo, Eisai, Kyowa-Kirin, Lilly, Ono Pharmaceutical, Pfizer, Taiho Pharma, and Takeda outside the submitted work. These organizations had no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; and in the decision to submit the article for publication. Rai Shimoyama, Ryuta Fukai, Akihiko Iwase, Yukihiro Tamura, Yusuke Chihara, Takafumi Okabe, Uryu Kiyoaki, Tadahisa Okuda, and Masataka Taguri have no conflicts of interest that are directly relevant to the content of this article. Ethics approval The study was conducted following the principles of the Declaration of Helsinki and was reviewed and approved by the Ethics Committee of the Tokushukai Group on 2 March, 2023 (approval number: TGE 01427-008). This study was registered at the University Hospital Medical Information Network Clinical Trials Registry on 9 March, 2023 (identification UMIN000050552). Consent to participate The requirement for obtaining informed consent from patients was waived because this was a retrospective analysis of anonymized patient data. Patients were allowed to opt out of the research use of their data, and related information is publicly available on the Tokushukai Medical Group website. Consent for publication Not applicable. Availability of data and material The data supporting the findings of this study are available from the corresponding author on reasonable request. Code availability Not applicable. Authors’ contributions Conceptualization, data curation, formal analysis, investigation, methodology, resources, writing (original draft): MH. Resources, writing (review and editing): KU. Resources, writing (review and editing): RS. Resources, writing (review and editing): TF. Resources, writing (review and editing): RF. Resources, writing (review and editing): AI. Resources, writing (review and editing): YT. Resources, writing (review and editing): YC. Resources, writing (review and editing): MF. Formal analysis: TO. Formal analysis: MT. Writing (review and editing): YI. Writing (review and editing): HM., (© 2024. The Author(s).)

Background: The PHILA study suggests that pyrotinib, trastuzumab, and docetaxel significantly improved progression-free survival (PFS) compared with placebo, trastuzumab, and docetaxel in patients with untreated HER2-positive metastatic breast cancer. In this study, we aimed to investigate the synergistic mechanisms of pyrotinib plus trastuzumab and provide further insights for the PHILA trial., Methods: The in vitro activity of combination treatments was assessed through cell biological and biochemical experiments. The in vivo efficacy was evaluated in cell-derived xenografts, a TUBO tumour model, and one clinical case. Next-generation sequencing was performed on circulating tumour DNA (ctDNA) from patients in the PHILA trial., Findings: The combination of pyrotinib and trastuzumab more effectively inhibited cell growth than pyrotinib or trastuzumab alone in models of HER2-dependent breast cancer. It potentiated membrane HER2 ubiquitination and downregulation, which resulted in a comprehensive blockade of the HER2 signalling pathway. The pyrotinib-altered membrane HER2 levels had no significant effect on trastuzumab-mediated antibody-dependent cell-mediated cytotoxicity (ADCC). We further validated the synergistic mechanisms in TUBO tumours and one clinical case, rather than models of HCC1954 cells harbouring the PIK3CA H1047R mutation. Similarly, in our centre cohort of the PHILA study, patients with genetic alterations in the HER2 signalling cascade had significantly shorter median PFS than individuals with the wild-type pathway., Interpretation: Our findings underscore the robust synergy between pyrotinib and trastuzumab in overcoming HER2 dependency and provide a rationale for pyrotinib, trastuzumab, and docetaxel as one of the optimal choices for patients with untreated HER2-positive metastatic breast cancer, who are dependent on the HER2 signalling cascade., Funding: This work was supported by the National Key Research and Development Program of China (2021YFF1201300), the National Natural Science Foundation of China (82172875), the CAMS Innovation Fund for Medical Sciences (CIFMS) (2022-I2M-2-001), and the Joint Innovative Fund of Beijing Natural Science Foundation and Changping District (L234004)., Competing Interests: Declaration of interests The authors declare that they have no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Epidermal growth factor receptor (EGFR) kinase has been implicated in the uncontrolled cell growth associated with non-small cell lung cancer (NSCLC). This has prompted the development of 3 generations of EGFR inhibitors over the last 2 decades due to the rapid development of drug resistance issues caused by clinical mutations, including T790M, L858R and the double mutant T790M & L858R. In this work we report the design, preparation and biological assessment of new irreversible 2,4-diaminopyrimidine-based inhibitors of EGFR kinase. Twenty new compounds have been prepared and evaluated which incorporate a range of electrophilic moieties. These include acrylamide, 2-chloroacetamide and (2E)-3-phenylprop-2-enamide, to allow reaction with residue Cys797. In addition, more polar groups have been incorporated to provide a better balance of physical properties than clinical candidate Rociletinib. Inhibitory activities against EGFR wildtype (WT) and EGFR T790M & L858R have been evaluated along with cytotoxicity against EGFR-overexpressing (A549, A431) and normal cell lines (HepG2). Selectivity against JAK3 kinase as well as physicochemical properties determination (logD 7.4 and phosphate buffer solubility) have been used to profile the compounds. We have identified 20, 21 and 23 as potent mutant EGFR inhibitors (≤20 nM), with comparable or better selectivity over WT EGFR, and lower activity at JAK3, than Osimertinib or Rociletinib. Compounds 21 displayed the best combination of EGFR mutant activity, JAK3 selectivity, cellular activity and physicochemical properties. Finally, kinetic studies on 21 were performed, confirming a covalent mechanism of action at EGFR., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Matthew Paul Gleeson reports financial support was provided by King Mongkut’s Institute of Technology Ladkrabang. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The most common type of non-small cell lung cancer is lung adenocarcinoma (LUAD), which is characterized by high morbidity and poor survival. Up-regulation of SASS6 expression can lead to the progression of various malignant tumors. However, there are no relevant studies on the role of SASS6 in LUAD. SASS6 was highly expressed in most tumors, reflecting a good diagnostic value, and its overexpression in LUAD indicated discouraging overall prognosis. Functional enrichment analysis suggested that SASS6 was associated with cell cycle in LUAD. In addition, patients with high SASS6 expression had worse immune infiltration, but higher TMB and immune checkpoint, and higher sensitivity to multiple targeted drugs such as osimertinib. Cell experiments confirmed that knockdown of SASS6 could inhibit the viability of tumor cells.SASS6 has important value in the diagnosis of cancer. In particular, SASS6 is a crucial factor in the progression of LUAD, and has important clinical value, especially in the diagnosis, prognosis and treatment., (© 2024. The Author(s).)

Overcoming resistance to epidermal growth factor receptor tyrosine kinase inhibitors, including osimertinib, is urgent to improve lung cancer treatment outcomes. Extracellular vesicle (EV)-derived microRNAs (EV-miRNAs) play important roles in drug resistance and serve as promising biomarkers. In this study, we aimed to identify EV-miRNAs associated with osimertinib resistance and investigate their clinical relevance. The release of excess EVs was confirmed in the osimertinib-resistant lung adenocarcinoma cell line PC9OR. The exposure of PC9OR-derived EVs and EV-miRNAs to PC9 cells increased cell viability after osimertinib treatment. Microarray analysis revealed that miR-130a-3p was upregulated in EVs derived from PC9OR cells and another osimertinib-resistant cell line (H1975OR). Transfection with miR-130a-3p attenuated osimertinib-induced cytotoxicity and apoptosis in both PC9 and H1975 cells, whereas osimertinib resistance in PC9OR cells was reversed after miR-130a-3p inhibition. Bioinformatics analysis revealed that runt-related transcription factor 3 is a target gene of miR-130a-3p, and it induced osimertinib resistance in PC9 cells. Patients with lower baseline serum miR-130a-3p concentrations had longer progression-free survival. miR-130a-3p is a potential therapeutic target and a predictive biomarker of osimertinib resistance in adenocarcinomas., (© 2024. The Author(s).)

The current study aimed to develop a reusable antibacterial coating that can be employed for efficient bacterial killing. We synthesized a water-soluble methacrylamide-based copolymer consisting of cationic and hydrophobic groups and coated it onto a glass surface through the formation of semi-interpenetrating polymer networks (semi-IPN) of aminopropyl triethoxysilane and glutaraldehyde. The coated surface was exposed to Gram-negative and Gram-positive bacteria, where the surface exhibited rapid bacterial killing ability within 5-15 min. The substrates displayed a minimal loss of antibacterial activity even after two water rinse cycles. The coatings were able to kill both the bacterial strains even after 5 weeks, suggesting excellent longevity. The surfaces were stable after repeated wiping cycles with 70% IPA using Kim wipes and 5 min sonication in DI water as no bactericidal activity was lost. Thus, a sustainable antibacterial copolymer coating was developed, and it is stable and reusable against bacterial contamination and could be employed as a long-term antibacterial coating.

Despite the widespread recognition of the global concern regarding the onset of cardiovascular diseases in a significant number of patients following cancer treatment, definitive strategies for prevention and treatment remain elusive. In this study, we established systems to evaluate the influence of anti-cancer drugs on the quality control of mitochondria, pivotal for energy metabolism, using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Osimertinib, an epidermal growth factor receptor tyrosine kinase inhibitor used for treatment in lung cancer, reportedly increases the risk of cardiovascular disease. However, its underlying mechanism is largely unknown. Here, we found that the treatment of hiPSC-CMs with osimertinib and doxorubicin, but not trastuzumab and cisplatin, revealed a concentration-dependent impairment of respiratory function accompanied by mitochondrial fission. We previously reported the significant role of sulfur metabolism in maintaining mitochondrial quality in the heart. Co-treatment with various inorganic sulfur donors (Na 2 S, Na 2 S 2 , Na 2 S 3 ) alongside anti-cancer drugs demonstrated that Na 2 S attenuated the cardiotoxicity of osimertinib but not doxorubicin. Osimertinib decreased intracellular reduced sulfur levels, while Na 2 S treatment suppressed the sulfur leakage, suggesting its potential in mitigating osimertinib-induced cardiotoxicity. These results imply the prospect of inorganic sulfides, such as Na 2 S, as a seed for precision pharmacotherapy to alleviate osimertinib's cardiotoxic effects., Competing Interests: Declaration of competing interest The authors have no conflict of interest., (Copyright © 2024 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Osimertinib, a tyrosine kinase inhibitor targeting mutant EGFR, has received approval for initial treatment in patients with Non-Small Cell Lung Cancer (NSCLC). While effective in both first- and second-line treatments, patients eventually develop acquired resistance. Metabolic reprogramming represents a strategy through which cancer cells may resist and adapt to the selective pressure exerted by the drug. In the current study, we investigated the metabolic adaptations associated with osimertinib-resistance in NSCLC cells under low glucose culture conditions. We demonstrated that, unlike osimertinib-sensitive cells, osimertinib-resistant cells were able to survive under low glucose conditions by increasing the rate of glucose and glutamine uptake and by shifting towards mitochondrial metabolism. Inhibiting glucose/pyruvate contribution to mitochondrial respiration, glutamine deamination to glutamate, and oxidative phosphorylation decreased the proliferation and survival abilities of osimertinib-resistant cells to glucose starvation. Our findings underscore the remarkable adaptability of osimertinib-resistant NSCLC cells in a low glucose environment and highlight the pivotal role of mitochondrial metabolism in mediating this adaptation. Targeting the metabolic adaptive responses triggered by glucose shortage emerges as a promising strategy, effectively inhibiting cell proliferation and promoting cell death in osimertinib-resistant cells., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Inflammatory bowel disease (IBD) encompasses chronic disorders that cause severe inflammation in the digestive tract. This study evaluates (E)-3-(3,4-dichlorophenyl)-N-(2,6-dioxopiperidin-3-yl) acrylamide (named SKT40), a derivative of dioxopiperidinamide, as a potential novel treatment for IBD. The pharmacological activity of SKT40 indicated positive interactions using network pharmacology and molecular docking in silico. In vivo, adult and larval zebrafish were tested to evaluate the effectiveness of SKT40 at different concentrations (7.5 μM, 10 μM, 15 μM) in preventing dextran sulfate sodium (DSS)-induced intestinal inflammation. The administration of SKT40 resulted in positive effects by reducing reactive oxygen species (ROS), lipid peroxidation, and cell apoptosis in zebrafish larvae. SKT40 demonstrated a significant reduction in intestinal damage in adult zebrafish by increasing antioxidant enzymes that combat the causes of IBD, such as superoxide dismutase (SOD), catalase (CAT), glutathione-S-transferase (GST), and glutathione peroxidase (GPx). It also reduces cellular damage and inflammation, as indicated by decreased levels of lactate dehydrogenase (LDH) and malondialdehyde (MDA). Gene expression analysis identified downregulation in gene expression of inflammatory mediators such as TNF-α, IL-1β, COX-2, and IL-6. Histopathological analysis showed tissue repair from DSS-induced damage and indicated reduced hyperplasia of goblet cells. These findings suggest that SKT40 effectively treats intestinal damage, highlighting its potential as a promising candidate for IBD therapy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

In the development of covalent inhibitors, acrylamides warhead is one of the most popular classes of covalent warheads. In recent years, researchers have made different structural modifications to acrylamides warheads, resulting in the creation of fluorinated acrylamide warheads and cyano acrylamide warheads. These new warheads exhibit superior selectivity, intracellular accumulation, and pharmacokinetic properties. Additionally, although ketoamide warheads have been applied in the design of covalent inhibitors for viral proteins, it has not received sufficient attention. Combined with the studies in kinase inhibitors and antiviral drugs, this review presents the structural features and the progression of acrylamides warheads, offering a perspective on future research and development in this field., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: We investigated the unknown mechanisms of osimertinib-resistant EGFR-mutant lung cancer., Methods: An osimertinib-resistant cell line (PC-9/OsmR2) was established through continuous exposure to osimertinib using an EGFR exon 19 deletion (19Del) lung adenocarcinoma cell line (PC-9). EGFR 19Del (M1), L858R/T790M/C797S (M6), and L858R/C797S (M8) expression vectors were introduced into Ba/F3 cells. A second osimertinib-resistant line (M1/OsmR) was established through continuous exposure to osimertinib using M1 cells., Results: SLC1A3 had the highest mRNA expression level in PC-9/OsmR2 compared to PC-9 cells by microarray analysis and SLC1A3 was increased by flow cytometry. In PC-9/OsmR2 cells, osimertinib sensitivity was significantly increased in combination with siSLC1A3. Because SLC1A3 functions in glutamic acid transport, osimertinib with a glutaminase inhibitor (CB-839) or an SLC1A3 inhibitor (TFB-TBOA) increased the sensitivity. Also, CB-839 plus TFB-TBOA without osimertinib resulted in greater susceptibility than did CB-839 or TFB-TBOA plus osimertinib. Comprehensive metabolome analysis showed that the M1/OsmR cells had significantly more glutamine and glutamic acid than M1 cells. CB-839 plus osimertinib exerted a synergistic effect on M6 cells and an additive effect on M8 cells., Conclusion: Targeting glutaminase and glutamic acid may overcome the osimertinib-resistant EGFR-mutant lung cancer., Competing Interests: Declaration of competing interest Dr. Takigawa has received grants and personal fees from Eli Lilly Japan, AstraZeneca, Daiichi-Sankyo Pharmaceutical, Chugai Pharmaceutical, Taiho Pharmaceutical, Pfizer Inc. Japan, Boehringer-Ingelheim Japan, and Ono Pharmaceutical; grants from Kyowa Hakko Kirin, Nippon Kayaku Co. Ltd., and Takeda Pharmaceutical Co. Ltd.; and personal fees from MSD and Bristol-Myers Squibb Company Japan outside the submitted work. Dr. Ochi has received personal fees from Eli Lilly Japan K.K., Taiho Pharmaceutical Co. ltd., Chugai Pharmaceutical, Pfizer Japan Inc., Ono pharmaceutical, MSD, Bristol-Myers Squibb, Boehringer Ingelheim, and Nippon Kayaku outside the submitted work. Dr. Fukazawa has received personal fees from Boehringer-Ingelheim Japan outside the submitted work. Dr. Kiura has received grants and personal fees from Taiho Pharmaceutical Co., Ltd., Nippon Boehringer Ingelheim Co., Ltd., Ono Pharmaceutical, Chugai Pharmaceutical, Nippon Kayaku Co., Ltd., Teijin Pharma Ltd., Shionogi Pharma Co., Ltd., Novartis Pharma, Takeda Pharmaceutical Co., Ltd., Kyorin Pharmaceutical Co., Ltd., Merck Biopharma Co., Ltd., MSD, and Pfizer Japan Inc. and personal fees from Eli Lilly Japan, AstraZeneca, Daiichi-Sankyo Pharmaceutical, Bristol-Myers Squibb, and Nipro Pharma Co. outside the submitted work. The other authors have no conflicts of interest to declare., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background/aim: Low selectivity and high frequency of side-effects are the major problems of currently used chemotherapeutics. Among natural compounds, the polyprenylated acylphloroglucinol, guttiferone E, isolated from Brazilian red propolis, has attracted attention due to its marked anticancer properties and was evaluated here for its role against osimertinib-resistant H1975 cells (with double mutations of epidermal growth factor receptor: EGFR L858R/T790M)., Materials and Methods: Guttiferone E was obtained from red propolis using established extraction procedures. Guttiferone E was tested using the H1975 cell line in in vitro (2D and 3D) cell cultures and in vivo in BALB/c athymic nude mice. Live/dead assay was also performed to support the results. Tumor tissues obtained from in vivo studies were used for western blotting. Guttiferone E reduced H1975 cell viability in a concentration-dependent manner. The IC 50 values in 2D and 3D cell lines were 2.56±0.12 μM and 11.25±0.34 μM. Furthermore, at 10 mg/kg intraperitoneally, guttiferone E significantly reduced the tumor volume in tumor xenografts when used alone and in combination with carboplatin. Guttiferone E and carboplatin displayed synergistic inhibition of H1975 cells and animal tumors. Co-treatment of guttiferone E with carboplatin induced more prominent apoptosis than treatment with either drug alone. Guttiferone E treatment induced cleavage of poly-ADP ribose polymerase and induced apoptosis by significantly reducing levels of mammalian target of rapamycin, sirtuin 1, sirtuin 7, superoxide dismutase, programmed death-ligand 1, and programmed cell death 1 in tumor tissues., Conclusion: Our results show guttiferone E to be a promising, novel and potent antitumor drug candidate for osimertinib-resistant lung cancer with EGFR L858R/T790M mutations., (Copyright © 2024 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

The serine/threonine kinase PAK4 plays a crucial role in regulating cell proliferation, survival, migration, and invasion. Overexpression of PAK4 correlates with poor prognosis in some cancers. KPT-9274, a PAK4 inhibitor, significantly reduces the growth of triple-negative breast cancer cells and mammary tumors in mouse models, and it also inhibits the growth of several other types of cancer cells. Interestingly, although it was first identified as a PAK4 inhibitor, KPT-9274 was also found to inhibit the enzyme NAMPT (nicotinamide phosphoribosyltransferase), which is crucial for NAD (nicotinamide adenine dinucleotide) synthesis and vital for cellular energy and growth. These results made us question whether growth inhibition in response to KPT-9274 was due to PAK4 inhibition, NAMPT inhibition, or both. To address this, we tested several other PAK4 inhibitors that also inhibit cell growth, to determine whether they also inhibit NAMPT activity. Our findings confirm that multiple PAK4 inhibitors also inhibit NAMPT activity. This was assessed both in cell-free assays and in a breast cancer cell line. Molecular docking studies were also used to help us better understand the mechanism by which PAK4 inhibitors block PAK4 and NAMPT activity, and we identified specific residues on the PAK4 inhibitors that interact with NAMPT and PAK4. Our results suggest that PAK4 inhibitors may have a more complex mechanism of action than previously understood, necessitating further exploration of how they influence cancer cell growth.

Multiresponsive hydrogels are valuable as biomaterials due to their ability to respond to multiple biologically relevant stimuli, i.e., temperature, pH, or reactive oxygen species (ROS), which can be present simultaneously in the body. In this work, we synthesize triple-responsive hydrogels through UV light photopolymerization of selected monomer compositions that encompass thermoresponsive N -isopropylacrylamide (NIPAM), pH-responsive methacrylic acid (MAA), and a tailor-made ROS-responsive diacrylate thioether monomer (EG 3 SA). As a result, smart P[NIPAM x - co -MAA y - co -(EG 3 SA) z ] hydrogels capable of being manufactured by digital light processing (DLP) 4D printing are obtained. The thermo-, pH-, and ROS-response of the hydrogels are studied by swelling tests and rheological measurements at different temperatures (25 and 37 °C), pHs (3, 5, 7.4, and 11), and in the absence or presence of ROS (H 2 O 2 ). The hydrogels are employed as matrixes for the encapsulation of ketoprofen (KET), an anti-inflammatory drug that shows a tunable release, depending on the hydrogel composition and stimuli applied. The cytotoxicity properties of the hydrogels are tested in vitro with mouse embryonic fibroblasts (NIH 3T3) and RAW 264.7 murine macrophage (RAW) cells. Finally, the anti-inflammatory properties are assessed, and the results exhibit a ≈70% nitric oxide reduction up to base values of pro-inflammatory RAW cells, which highlights the anti-inflammatory capacity of P[NIPAM 80 - co -MAA 15 - co -(EG 3 SA) 5 ] hydrogels, per se , without being necessary to encapsulate an anti-inflammatory drug within their network. It opens the route for the fabrication of customizable 4D printable scaffolds for the effective treatment of inflammatory pathologies.

Activating EGFR (epidermal growth factor receptor) mutations can be inhibited by specific tyrosine kinase inhibitors (TKIs), which have changed the landscape of lung cancer therapy. However, due to secondary mutations and bypass receptors, such as AXL (AXL receptor tyrosine kinase), drug resistance eventually emerges in most patients treated with the first-, second-, or third-generation TKIs (e.g., osimertinib). To inhibit AXL and resistance to osimertinib, we compare two anti-AXL drugs, an antibody (mAb654) and a TKI (bemcentinib). While no pair of osimertinib and an anti-AXL drug is able to prevent relapses, triplets combining osimertinib, cetuximab (an anti-EGFR antibody), and either anti-AXL drug are initially effective. However, longer monitoring uncovers superiority of the mAb654-containing triplet, possibly due to induction of receptor endocytosis, activation of immune mechanisms, or disabling intrinsic mutators. Hence, we constructed a bispecific antibody that engages both AXL and EGFR. When combined with osimertinib, the bispecific antibody consistently inhibits tumor relapses, which warrants clinical trials., Competing Interests: Declaration of interests We, the authors, plan to file a patent application for the bispecific antibody (EGFR/AXL), namely Bis1, which has been developed in this study., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)