Methods in molecular biology (Clifton, N.J.)Cewen Chen, Qiaochu Fu, Chao Gao, Shinya Tanaka, Masamichi Imajo
Colorectal cancer (CRC) is a major malignancy with significant global implications for human health. The development of a reliable and pathologically relevant orthotopic CRC model is essential for advancing our understanding of its molecular mechanisms and for developing more effective therapeutic interventions. However, the construction of such models is fraught with challenges primarily because of the technical complexities involved in the transplantation of CRC cells into the intestinal epithelium. In this chapter, we describe a recently developed method for generating an orthotopic CRC model within the mouse cecal epithelium. This model enables tumor development and progression within a natural tissue microenvironment and facilitates complex interactions between tumor cells and surrounding normal cells, thereby replicating the intratumor heterogeneity of CRC. All procedures in this method are visualized under stereomicroscopic observation, leading to the efficient engraftment of CRC cells and subsequent tumor development. The establishment of this reliable orthotopic CRC model, which mimics tumor development in a more natural microenvironment, offers new opportunities to explore the molecular mechanisms underlying CRC and to assess novel anticancer therapies in pathologically relevant contexts.
Colorectal cancer (CRC) remains a leading cause of cancer-related morbidity and mortality worldwide. Recent advancements in cancer research emphasize the importance of personalized medicine, necessitating robust preclinical models that closely mimic the tumor microenvironment. Colorectal cancer organoids have emerged as a pivotal tool in this domain, providing a platform that preserves the cellular heterogeneity and allows high-throughput drug testing. However, organoids do not recapitulate the complexity of the tumor ecosystem, its architecture, and stromal interactions characteristic of native tumors. Here, we describe a model to maintain CRC tissue explants in culture. These explants enable the assessment of therapeutic responses ex vivo. Furthermore, they may offer insights into tumor-immune interactions and facilitate the evaluation of novel immunotherapeutic agents. The continued refinement of CRC tissue explant models promises to enhance our understanding of tumor biology and improve the predictive accuracy of preclinical studies, ultimately contributing to more effective and personalized treatment strategies for colorectal cancer patients.
Reviews of physiology, biochemistry and pharmacologyUndurti N Das
Polyunsaturated fatty acids (PUFAs), especially gamma-linolenic acid (GLA), arachidonic acid (AA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), can block HMG-CoA reductase activity and thus decrease cholesterol synthesis/formation. Cholesterol has antioxidant actions and thus reduces the formation of ROS (reactive oxygen species). In contrast, PUFAs augment the generation of ROS. PUFAs undergo peroxidation to form lipid peroxides that are toxic to tumor cells but not normal cells. GPX4 (glutathione peroxidase) reduces the formation of lipid peroxides and thus enhances tumor cell resistance to the tumoricidal actions of radiation, anticancer drugs, and immune checkpoint inhibitors (ICIs). Tumor cells have relatively high amounts of GPX4 and hence are resistant to lipid peroxide-induced ferroptosis/apoptosis. In contrast, normal cells can upregulate their GPX4 activity/content upon exposure to toxic lipid peroxides and hence are resistant to ferroptosis or apoptosis when supplemented with various PUFAs or exposed to radiation, anticancer drugs, and ICIs. Thus, PUFAs (especially GLA, AA, and DHA) are selectively toxic to tumors but not normal cells. Interferon-γ (IFN-γ) is secreted by CD8+ T cells, and AA induces ferroptosis of tumor cells in an ACSL4 (the protein encoded by this gene is an isozyme of the long-chain fatty-acid-coenzyme A ligase family)-dependent lipid peroxidation process. IL-6, TNF-α, and IFN-γ activate PLA2 to release AA from membrane lipids, which is then peroxidized to trigger apoptosis or ferroptosis of cancer cells. CD8+ T cells/macrophages/TILs/NK cells and other immunocytes downregulate the expression of SLC3A2 and SLC7A11, two subunits of the glutamate-cystine antiporter system x c - , and impair the uptake of cystine by tumor cells, because of which accumulation of toxic lipid peroxides occurs in the tumor cells, leading to their apoptosis/ferroptosis/necrosis. In C6 glioma cells, the inhibition of the cystine/glutamate (XC) antiporter and glutamate-cysteine ligase (GCL) results in inhibition of glutathione biosynthesis that leads to ferroptosis of cancer cells because of accumulation of toxic lipid peroxides. Thus, the lipid peroxidation process is at the center of tumor cell apoptosis/ferroptosis.
Methods in molecular biology (Clifton, N.J.)Benjamin L Kidder
Deep learning has transformed medical image analysis, but progress in cancer and stem cell applications is often constrained by limited access to large, diverse, well-annotated imaging datasets. This bottleneck is especially acute for studies of tumor heterogeneity and cancer stem cell (CSC) biology, where rare phenotypes and dynamic cell-state transitions-frequently linked to stemness-associated transcriptional programs (e.g., OCT4, SOX2, NANOG)-benefit from high-quality imaging across many samples and conditions. At the same time, regulatory and practical barriers (patient privacy, acquisition cost, and uneven institutional data sharing) restrict dataset scale and reuse. Diffusion models offer a practical route to synthetic data expansion by generating high-fidelity synthetic images that retain salient radiologic and pathologic features. In this chapter, we present an end-to-end protocol for adapting latent diffusion (Stable Diffusion) to oncology imaging using DreamBooth fine-tuning with small numbers of representative images, coupled with text-to-image and image-to-image workflows to generate controlled variations across modalities and disease presentations (e.g., brain tumor MRI, breast cancer mammography/CESM). We also describe quantitative and qualitative evaluation strategies, including Fréchet Inception Distance (FID) benchmarking and expert review considerations, to assess realism and diversity. These methods enable cancer and stem cell biologists to augment training data for segmentation and classification, build shareable educational resources, and prototype analyses for rare tumors or stemness-enriched subtypes while potentially reducing reliance on direct sharing of patient images.
Methods in molecular biology (Clifton, N.J.)Nadia Nasir, Maryalice Stetler-Stevenson, Hao-Wei Wang
Flow cytometry is the cornerstone for establishing the diagnosis of chronic lymphocytic leukemia (CLL), owing to its characteristic and well-defined immunophenotype that enables accurate distinction from other leukemias and lymphomas. Beyond diagnosis, flow cytometry provides essential prognostic information and allows sensitive detection of minimal residual disease (MRD), a strong predictor of clinical outcome. CLL MRD assessment is increasingly used to guide risk stratification, therapeutic decision-making, and treatment duration in the era of targeted therapies and immunotherapies. This chapter reviews best practices for specimen collection, processing, staining, and data analysis and summarizes the principles of flow cytometric MRD assessment in CLL.
Methods in molecular biology (Clifton, N.J.)Samna Sagadevan, Paloma Ordóñez-Morán
Organoids and fibroblasts derived from patient tissue serve as physiologically meaningful in vitro models to investigate tissue biology, diseases, and treatment responses. In this study, we present a robust protocol for the simultaneous isolation and long-term culture of patient-derived organoids (PDOs) and fibroblasts from both healthy and colorectal cancer samples. The workflow uses enzymatic dissociation followed by efficient separation into epithelial and stromal cell fractions. Organoids are embedded in Matrigel or Matrigel-Collagen I mixtures to support three-dimensional growth, whereas fibroblasts are maintained on conventional two-dimensional culture dishes. This dual-culture approach facilitates a broad range of downstream applications.
Methods in molecular biology (Clifton, N.J.)Benjamin L Kidder
Cancer cell identity is governed by coordinated transcriptional programs that are frequently rewired during tumorigenesis. Systematic identification of cancer type-specific gene regulatory networks provides a framework for understanding oncogenic state transitions and for prioritizing candidate therapeutic targets. Here, we present a reproducible network-based workflow for reconstructing and analyzing transcriptional regulatory programs across human cancer types using publicly available expression datasets. We describe procedures for curating and preprocessing microarray data from the Gene Expression Omnibus, implementing random forest classification, and reconstructing gene regulatory networks using the CellNet platform. Detailed guidance is provided for evaluating classifier performance, quantifying network influence scores, integrating transcription factor, target interaction resources, and performing functional enrichment analyses. In addition, we outline approaches for comparing cancer-specific networks with corresponding normal tissue profiles to identify candidate drivers of malignant cell identity and potential prognostic biomarkers. Together, these protocols provide investigators with a scalable computational framework for defining cancer type-specific transcriptional states and for systematically interrogating regulatory mechanisms underlying tumor heterogeneity.
International braz j urol : official journal of the Brazilian Society of UrologyNaiwen Chen, Lichen Chen, Jinming Cai, Mingyue Tan, Dongliang Xu
INTRODUCTION: Radical cystectomy (RC) is the standard therapy for muscle-invasive bladder cancer (MIBC) and refractory high-risk non-muscle-invasive bladder cancer (1-3). Single-port robot-assisted radical cystectomy (RARC) offers notable minimally invasive advantages (4-6), whereas conventional transperitoneal approaches are associated with intestinal and gastrointestinal complications (7). This study evaluated an optimized extraperitoneal single-port RARC with orthotopic neobladder reconstruction for improved surgical safety and clinical outcomes. MATERIALS AND METHODS: A 57-year-old male presented with three months of intermittent painless gross hematuria. Pelvic CT revealed a bladder mass. Preoperative biopsy confirmed high-grade urothelial carcinoma with muscularis propria invasion, and the patient received gemcitabine-cisplatin neoadjuvant chemotherapy before radical surgery. Preoperative MRI identified a 3.2×2.9×2.6 cm bladder lesion without extravesical invasion or lymphadenopathy. During the procedure, the patient was placed in a supine position with buttocks elevated. A 5-cm infraumbilical single incision was made for da Vinci Xi-assisted extraperitoneal RARC, and a single-port multichannel device was deployed through the incision. Robotic instruments including a 30° endoscope, monopolar scissors, bipolar forceps, and a robotic stapler were arranged in a chopstick configuration. Via the extraperitoneal approach, we mobilized the bilateral ureters, bilateral vas deferens, and umbilical artery, dissected the bladder lateral ligaments with vascular ligation, and established a sufficient extraperitoneal working space. Bladder and prostate dissection was performed along the perivesical avascular plane, with careful protection and precise hemostasis of the dorsal venous complex. Standard pelvic lymphadenectomy was concurrently conducted during radical resection of the bladder and prostate (8). A segment of ileum was then harvested to construct an orthotopic neobladder (9), followed by anastomosis with the bilateral ureters and urethral stump. RESULTS: Operative time was 400 minutes with 200 mL blood loss and no transfusion. No perioperative complications occurred, and the patient was discharged on postoperative day 6. Pathological diagnosis was pT2aN0M0 with negative surgical margins and negative lymph nodes. The 12-month follow-up showed no tumor recurrence, normal renal function, complete daytime continence, and mild nocturnal incontinence requiring one nightly pad. CONCLUSIONS: Extraperitoneal single-port RARC with orthotopic neobladder reconstruction is a feasible minimally invasive procedure for MIBC. The optimized infraumbilical single-incision technique preserves peritoneal integrity and avoids intestinal mobilization, effectively reducing abdominal complications. It achieves reliable oncological results and improves patients' postoperative continence and quality of life.
Methods in molecular biology (Clifton, N.J.)Dana Delgado, Jolene L Cardinali, Joseph A DiGiuseppe
Immunophenotyping by flow cytometry is an important component in the diagnostic evaluation of patients with acute lymphoblastic leukemia. This technique further permits the detection of minimal residual disease after therapy, a robust prognostic factor that may guide individualized treatment. We describe here laboratory methods for both the initial characterization of lymphoblasts at diagnosis and the detection of rare leukemic lymphoblasts after treatment. In addition to antibody combinations suitable for diagnosis and detection of minimal residual disease, we describe procedures for peripheral blood and bone marrow sample preparation, procedures for labeling of cell-surface and intracellular proteins with fluorochrome-conjugated antibodies, and approaches to analysis of immunophenotypic data, including those obtained in patients following CD19-targeted therapies.
Aberrant three-dimensional genome organization is a hallmark of cancer, often driving oncogene activation through mechanisms such as enhancer hijacking. High-throughput chromosome conformation capture (Hi-C) maps these interactions on a genome-wide scale. Unlike earlier dilution-based methods, in situ Hi-C performs proximity ligation within intact nuclei, minimizing random ligation noise and enabling fine-scale structure detection. This chapter describes an optimized in situ Hi-C protocol tailored for cancer cell lines using MboI digestion and biotin-mediated pull-down to generate high-complexity libraries. We further outline a computational workflow that extends beyond standard topological mapping of compartments and topologically associating domains to identify cancer-specific aberrations. Specifically, we focus on detecting chromosomal rearrangements (structural variants) and characterizing the distinct circular topology of extrachromosomal DNA. This integrated experimental and analytical framework provides the necessary tools to dissect the spatial dysregulation underlying tumor evolution.
Methods in molecular biology (Clifton, N.J.)Pallavi Kanwar Galera, Raul Braylan
Immunophenotyping by multiparameter flow cytometry is a rapid and efficient technique to simultaneously assess and correlate multiple individual cell properties like size and internal complexity along with antigen expression in a population of cells. This method is utilized for rapid characterization of the blasts and classification of acute myeloid leukemia (AML) in both the peripheral blood (PB) and bone marrow (BM). This technique is not only useful in the initial diagnosis but also in monitoring and determining the prognosis of the disease through minimal residual disease (MRD) testing. This chapter provides an overview of procedures for specimen processing, staining, and immunophenotyping of AML and describes the principles of data analysis for AML classification and MRD testing.
Methods in molecular biology (Clifton, N.J.)Prashant R Tembhare, Nilesh Deshpande, Sitaram Ghogale
Measurable residual disease (MRD) serves as a critical biomarker of prognosis, treatment efficacy, and clinical outcome. It captures the presence of residual tumor cells below the detection threshold of conventional microscopy. Multiparametric flow cytometry (MFC) offers a rapid, cost-efficient, and widely applicable platform for MRD detection through leukemia-associated immunophenotypes (LAIPs) and deviation-from-normal (DfN) antigen maturation patterns. This technique underpins a wide range of clinical applications, including risk stratification, therapeutic decision-making, and post-transplant surveillance. This chapter outlines essential technical parameters for achieving high-sensitivity MRD detection across hematologic malignancies such as B-ALL, T-ALL, AML, MM, and CLL. Key topics covered include material required, reagent preparation, antibody panel design, sample processing and acquisition, gating strategies, and illustrative examples. Special emphasis is given to adaptations necessary for MRD monitoring following CD19-targeted therapies in B-ALL. By integrating rigorous methodology with evolving innovations, MFC continues to advance as a precise and expedient tool for MRD assessment, contributing significantly to personalized treatment strategies.
International braz j urol : official journal of the Brazilian Society of UrologyMarcos Tobias-Machado, Ricardo C Brianson, John Eder Gamarra Bravo, Alcedir Raiser, Eliney Faria
INTRODUCTION: Horseshoe kidney is an uncommon congenital fusion anomaly that can make renal tumor surgery especially challenging because of altered rotation, limited mobility, variable vascular supply, and an unpredictable collecting system (1-7). This video presents a robot-assisted partial nephrectomy for a high-complexity renal tumor in this setting. CASE PRESENTATION: A 33-year-old man, with ECOG 0 and no relevant comorbidities, was diagnosed with a 7.5-cm solid renal mass in the central posterior portion of the left moiety of a horseshoe kidney. The lesion had a RENAL score of 10p. Contrast-enhanced computed tomography and three-dimensional reconstruction were used to understand the relationship between the tumor, aberrant vessels, renal hilum, and collecting system, supporting the decision to attempt nephron-sparing surgery (5, 8). Surgical technique and results: The procedure was performed through a transperitoneal robotic approach with the patient in right lateral decubitus using the Da Vinci Si platform. Port placement followed a standard renal robotic configuration, with a paramedian supraumbilical camera port, three robotic working ports along a craniocaudal lateral axis, a caudal fourth-arm port, and two medial assistant ports for suction, exposure, and support during renorrhaphy. After exposure of the horseshoe kidney and left hilar dissection, two arterial branches and one renal vein were identified. Tumor excision was performed under vascular control, with 20 minutes of warm ischemia and no collecting system opening, followed by two-layer absorbable renorrhaphy with adjunctive hemostatic agents. The operative time was 150 minutes. No transfusion, conversion, drain placement, or relevant immediate complication occurred. The urinary catheter was removed after 24 hours, and the patient was discharged 72 hours after surgery. Pathology showed clear cell renal cell carcinoma, Fuhrman grade 3, pT2N0M0, with negative surgical margins. During 12 months of oncologic follow-up, renal function remained stable and semiannual imaging showed no evidence of recurrence. Contemporary video reports have also emphasized the feasibility of advanced robotic renal surgery and complex partial nephrectomy strategies in selected patients (9, 10). CONCLUSION: In a carefully selected patient, robot-assisted partial nephrectomy supported by three-dimensional planning was feasible for a complex renal tumor in a horseshoe kidney, with negative surgical margins, preserved renal function, and no recurrence during 12 months of follow-up.
Cell plasticity is a major driver of colorectal cancer (CRC), as tumors hijack intestinal stem cell regenerative programs to create a dynamic, bidirectional axis between complementary stem-like states. Cancer stem-like cells (CSCs) display hallmark stem cell properties, including self-renewal, multipotency, controlled differentiation, and quiescence, and are now recognized as key orchestrators of CRC biology. They can sustain tumor initiation and long-term growth, seed metastases, enable periods of dormancy, and promote resistance to chemotherapy. As a result, therapeutically dismantling CSC populations and their supportive niches offers a compelling strategy to improve treatment responses and ultimately patient survival.
The Immuno-Oncology Biological Research (IOBR) package is an R-based analysis tool for exploring the tumor microenvironment (TME) and its influence on anti-tumor immunity. Built for high-throughput data-spanning both transcriptomic and genomic profiles-IOBR integrates six analytical modules, including transcriptomic data preprocessing, TME profiling, TME pattern identification, ligand-receptor interaction analysis, genome-TME interaction assessment, and visualization. In this chapter, we walk through a multi-omics workflow using example datasets, illustrating data preparation, distribution analyses, result interpretation, and graphical output. IOBR is open source and is available at https://github.com/IOBR/IOBR and a detailed GitBook ( https://iobr.github.io/book/ ) offers a complete manual and analysis guide for each function.
Methods in molecular biology (Clifton, N.J.)Benjamin L Kidder
Teratoma formation is the gold standard assay for evaluating the developmental pluripotency of human and mouse embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). Following subcutaneous injection into immunodeficient mice, pluripotent stem cells spontaneously differentiate into derivatives representing all three embryonic germ layers-ectoderm, mesoderm, and endoderm. Beyond serving as a functional assay for pluripotency, teratomas provide a unique three-dimensional model system for studying early human development and lineage specification in vivo. This chapter describes comprehensive protocols for teratoma formation in immunodeficient mice, tissue processing for multiple downstream genomic applications, and multi-omics profiling approaches. We detail methods for embryonic stem cell culture, teratoma generation via subcutaneous injection, tissue dissection and processing for chromatin immunoprecipitation followed by sequencing (ChIP-Seq), RNA sequencing (RNA-Seq), single-cell multiome profiling combining chromatin accessibility (ATAC-Seq) and gene expression (scRNA-Seq), and histological analysis using hematoxylin and eosin (H&E) staining. Additionally, we provide bioinformatics workflows for analyzing the resulting genomic datasets to characterize the epigenetic and transcriptional landscapes of teratoma-derived tissues. These methods enable comprehensive molecular characterization of developmental processes and provide valuable resources for stem cell biologists studying pluripotency, differentiation, and early embryonic development.
Methods in molecular biology (Clifton, N.J.)Benjamin L Kidder
The rapid growth of biomedical literature has created an urgent need for computational tools that enable researchers to systematically analyze publication trends, identify emerging research themes, and map the evolution of scientific fields. PubMed Atlas is a command-line and web-enabled workflow for topic-driven bibliometrics and trend intelligence using PubMed E-utilities. The pipeline executes PubMed queries, retrieves matching PMIDs, downloads full metadata records in batches, parses structured information (title, abstract, authors/affiliations, MeSH terms, publication types, grants, keywords, DOI), and stores normalized data in a local SQLite database for rapid querying and visualization. A Streamlit dashboard provides interactive exploration of publication trends, journal distributions, MeSH term summaries, geographic distributions, and recent article browsing with direct PubMed links. This protocol describes the installation, configuration, and operation of PubMed Atlas for cancer stem cell and stem cell transcriptional network research, and other fields, enabling investigators to conduct reproducible bibliometric analyses and identify knowledge gaps in rapidly evolving fields.
Pakistan journal of pharmaceutical sciencesYunpeng Hu, Chen Li, Yiyao Cui
BACKGROUND: Breast cancer (BC) remains a significant global health threat with complex pathogenesis often linked to chemotherapy resistance and inflammatory microenvironments. Histidine triad nucleotide-binding protein 2 (HINT2) is a mitochondrial protein involved in metabolism and immunity, but its specific role in BC and potential as a therapeutic target require further elucidation. OBJECTIVES: This study aimed to identify novel therapeutic targets for BC and evaluate the anti-tumor efficacy and molecular mechanisms of Albiflorin (Al), a natural monoterpene glycoside. METHODS: Proteomic profiling was performed on BC xenografts in nude mice to identify differentially expressed proteins. In-vivo and in-vitro models (MDA-MB-231 cells) were established to assess the effects of Al (20 mg/kg or 20 µM) on tumor growth, apoptosis and cytokine production. Molecular docking and co-immunoprecipitation (Co-IP) were used to investigate the interactions among HINT2, NLRP3 and Al. RESULTS: HINT2 was significantly upregulated in BC tissues and identified as a primary pro-tumorigenic factor. Co-IP confirmed an endogenous interaction between HINT2 and NLRP3. Al exhibited strong binding affinity to HINT2 (-7.36 kcal/mol) and significantly suppressed tumor volume and weight. Furthermore, Al treatment downregulated the HINT2/NLRP3 axis, reduced inflammatory cytokines (IL-1β, IL-6, TNF-α) and promoted apoptosis by modulating Caspase-3, Bax and Bcl-2 levels. CONCLUSION: Albiflorin suppresses BC progression by inhibiting the HINT2/NLRP3 signaling pathway, suggesting HINT2 is a viable biomarker and Al a promising therapeutic agent for BC treatment.
Pakistan journal of pharmaceutical sciencesOrcun Can, Abdullah Sakin
BACKGROUND: Esophageal adenocarcinoma is associated with poor survival despite advances in systemic therapy. Readily available serum biomarkers may improve risk stratification; however, their combined role in biomarker-guided treatment decisions remains insufficiently investigated. OBJECTIVES: To evaluate the prognostic significance of combined baseline carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9) levels and explore their utility for biomarker-based risk stratification in patients with esophageal adenocarcinoma receiving systemic chemotherapy. METHODS: This retrospective real-world cohort study included 38 patients with histologically confirmed esophageal adenocarcinoma who received systemic chemotherapy at a tertiary referral center between January 2016 and December 2021. Patients were classified into three exploratory biomarker-defined risk groups according to baseline serum CEA and CA19-9 levels. Overall survival was evaluated using Kaplan-Meier analysis and Cox proportional hazards regression, while treatment outcomes were explored across biomarker-defined groups and chemotherapy regimens. RESULTS: Elevated baseline CEA and CA19-9 levels were associated with shorter overall survival. Patients with simultaneous elevation of both biomarkers demonstrated the poorest survival outcomes (multivariable hazard ratio = 2.22). Differences in survival were observed across chemotherapy regimen groups within biomarker-defined categories; however, these comparisons were exploratory and may have been influenced by treatment-selection bias and unmeasured clinical factors. Patients with distal or gastroesophageal junction adenocarcinoma showed more favorable outcomes than those with cervical or thoracic tuxzmors. CONCLUSION: Combined assessment of baseline CEA and CA19-9 may provide a practical and inexpensive approach for biological risk stratification in esophageal adenocarcinoma. However, these findings do not establish that the biomarkers can guide chemotherapy selection. Prospective multicenter studies with independent validation are required to determine their prognostic and predictive value before biomarker-guided treatment selection can be recommended in routine clinical practice.
Pakistan journal of pharmaceutical sciencesZaishuang Ju, Yutong Wu, Lili Tian, Dachuan Shen, Ruoyu Wang
BACKGROUND: Patients with lung adenocarcinoma (LUAD) receiving radiotherapy represent an important but underexplored clinical subgroup. These patients often undergo concomitant pharmacologic treatments, yet the prognostic impact and underlying determinants of such combined regimens remain poorly understood. OBJECTIVE: This retrospective observational study aimed to develop and validate a radiotherapy-specific machine learning prognostic model for LUAD and to compare survival across concomitant pharmacologic regimens. METHODS: In this retrospective observational study, using genomic and clinical data from TCGA, a radiotherapy-specific prognostic model for LUAD was developed and validated through ten machine learning algorithms. Survival analyses were conducted across distinct concomitant pharmacologic strategies, followed by functional enrichment to elucidate molecular mechanisms underlying differential outcomes. RESULTS: Demonstrating robust prognostic abilities, the model efficiently sorted patients into high- and low-risk categories. Both treatment type and risk score independently predicted overall survival, with significant interaction effects. Low-risk patients receiving targeted or combination therapy-mainly erlotinib, gefitinib, or bevacizumab-exhibited substantially improved survival compared with those receiving conventional chemotherapy. Enrichment of "Exogenous peptide presentation," "MHC class II assembly," "Peptide-MHC II assembly," and "Symbiotic interaction" pathways indicated immune modulation and host-tumor crosstalk as key mediators of treatment efficacy. CONCLUSION: This study establishes a radiotherapy-specific prognostic model for lung adenocarcinoma, demonstrating distinct molecular and therapeutic heterogeneity and highlighting the superior survival benefit of targeted combination therapy in low-risk patients.
Pakistan journal of pharmaceutical sciencesKai Liang, Xiangyu Zhang, Yin Yang
BACKGROUND: Gastric cancer (GC) remains a leading cause of cancer-related mortality worldwide and effective therapeutic targets remain limited. OBJECTIVES: This study aimed to investigate the expression and functional role of miR-326 in gastric cancer, identify its direct target gene and elucidate the underlying molecular mechanisms involving the PI3K/Akt signaling pathway and the regulation of inflammatory cytokines. METHODS: miR-326 expression was detected by qRT-PCR in 56 paired GC tissues and adjacent normal tissues, as well as in GC cell lines (SGC-7901, MKN-45, AGS) and normal GES-1 cells. CCK-8 assays, flow cytometry, Transwell migration/invasion assays and ELISA were performed to assess cell proliferation, apoptosis, migration/invasion and cytokine secretion. Dual-luciferase reporter assay and Western blotting were used to validate the target gene and pathway activation. RESULTS: miR-326 was significantly upregulated in GC tissues and cell lines. ETS1 was identified as a direct target of miR-326. Overexpression of miR-326 reduced ETS1 expression, activated the PI3K/Akt pathway (increased p-PI3K and p-AKT), promoted cell proliferation, migration and invasion, inhibited apoptosis and induced an imbalance of inflammatory cytokines (increased IL-6 and TNF-α; decreased IL-10 and IL-17). Co-overexpression of ETS1 or treatment with the PI3K inhibitor LY294002 reversed these effects. CONCLUSIONS: miR-326 promotes malignant phenotypes in gastric cancer cells by targeting ETS1, activating the PI3K/Akt signaling pathway, and inducing dysregulation of inflammatory cytokines. The miR-326/ETS1/PI3K/Akt axis may serve as a potential diagnostic biomarker and therapeutic target for gastric cancer.
Pakistan journal of pharmaceutical sciencesOrcun Can
BACKGROUND: The biology of the estrogen receptor-positive (ER+) and human epidermal growth factor receptor 2-negative (HER2-) breast cancers is heterogeneous even when they are categorized by their risk via genomics. Transcriptomic PGR expression reflects endocrine pathway activity and may provide complementary biological information within established GENE70-derived genomic-risk categories. Whether this molecular marker improves the biological interpretation of genomic-risk stratification beyond conventional clinicopathological assessment remains uncertain. OBJECTIVES: The aim of this study was to determine whether transcriptomic PGR expression provides complementary biological and prognostic information within reconstructed GENE70-derived genomic-risk categories and refines the characterization of endocrine-related tumour biology in ER-positive/HER2-negative breast cancer. METHODS: This study analysed publicly available transcriptomic and clinical data from three cohorts: METABRIC (discovery cohort), GSE96058/SCAN-B cohort (validation cohort) and TCGA-BRCA cohort (molecular validation cohort). The GENE70-derived genomic-risk score was reconstructed for each cohort using matched genes. Cox regression, Kaplan-Meier analysis and subgroup comparisons were used to assess relationships between PGR expression, clinicopathologic variables, molecular features and survival outcomes. RESULTS: Across the three independent cohorts, low transcriptomic PGR expression was consistently associated with higher GENE70-derived genomic risk, increased MKI67 expression, reduced ESR1 expression and enrichment of the Luminal B subtype. Survival findings differed between cohorts. In the discovery METABRIC cohort, transcriptomic PGR expression showed heterogeneous associations with survival, particularly within GENE70-derived high-risk subgroups, whereas the external GSE96058/SCAN-B validation cohort demonstrated consistent associations between low PGR expression and poorer overall survival in both the overall ER-positive/HER2-negative population and GENE70-derived high-risk subgroups. CONCLUSION: These findings suggest that transcriptomic PGR provides complementary biological and prognostic information within GENE70-derived genomic-risk categories. However, because treatment response was not evaluated in the present study, the findings should not be interpreted as evidence of predictive or pharmacogenomic utility and prospective studies incorporating treatment-response analyses are required before such applications can be established.
Journal of medical economicsWilliam V Padula, Benjamin G Cohen, Paul Steel, Sreevalsa Appukkuttan, Bashir Kalayeh, Matthew Rettig
AIMS: To evaluate the cost-effectiveness of darolutamide plus androgen deprivation therapy (DARO+ADT) versus enzalutamide plus ADT (ENZA+ADT) and apalutamide plus ADT (APA+ADT) as first-line androgen receptor pathway inhibitor (ARPI) double therapies for metastatic castration-sensitive prostate cancer (mCSPC) from a US healthcare sector perspective. METHODS: A partitioned survival model with three health states (progression-free mCSPC, progressed mCRPC, and death) projected lifetime costs and outcomes over a 30-year horizon with monthly cycles. DARO+ADT efficacy was derived from the ARANOTE trial; comparative efficacy for ENZA+ADT and APA+ADT was estimated via network meta-analysis-derived hazard ratios. Costs (drug acquisition, adverse event management, subsequent therapy, end-of-life care) and utilities were derived from trial data, published literature, and US cost sources. Costs and outcomes were discounted at 3% annually. Deterministic, probabilistic, and two-way sensitivity analyses assessed parameter uncertainty at a willingness-to-pay threshold of $150,000/QALY. RESULTS: DARO+ADT generated 4.18 QALYs versus 4.03 (ENZA+ADT) and 3.99 (APA+ADT), driven by longer treatment persistence and lower adverse-event-related disutility. Versus ENZA+ADT, DARO+ADT yielded 0.15 incremental QALYs at $12,564 incremental cost (ICER: $85,108/QALY; net monetary benefit: $9,579). Versus APA+ADT, DARO+ADT was dominant (0.19 additional QALYs, $4,293 lower cost; net monetary benefit: $33,153). Two-way sensitivity analyses varying off-treatment utilities and post-discontinuation therapy costs showed that DARO+ADT remained cost-effective versus ENZA+ADT and dominant versus APA+ADT, respectively, across the parameter ranges tested. Probabilistic sensitivity analysis showed DARO+ADT was cost-effective in 51% and 52% of simulations versus ENZA+ADT and APA+ADT, respectively, which demonstrates uncertainty around the incremental findings. LIMITATIONS: The analysis relies on indirect comparisons and extrapolated modeling, factors that may limit the generalizability of the findings. CONCLUSIONS: DARO+ADT was cost-effective versus ENZA+ADT and dominant versus APA+ADT, driven by improved tolerability and treatment persistence, supporting its value as a first-line mCSPC option.
International journal of hyperthermia : the official journal of European Society for Hyperthermic Oncology, North American Hyperthermia GroupJohannes Crezee, C Paola Tello Valverde, M Willemijn Kolff, H J G Desirée van den Bongard, Ben J Slotman, Akke Bakker, H Petra Kok
BACKGROUND: Thermoradiotherapy at 40-43 °C is an effective treatment for locoregional recurrent (LRR) breast cancer patients. During hyperthermia temperature monitoring is often limited to skin temperatures, but their predictive value for achieving locoregional tumor control (LRC) remains unclear for tumor depths ≥1cm. This study compared predictive value of skin versus invasive intratarget temperatures. METHODS: We reported earlier the thermal dose-effect relationship in 112 patients with LRR breast cancer treated in 2010-2017 with postoperative re-irradiation (8 × 4 Gy/23 × 2 Gy) and 4-5 weekly hyperthermia sessions with 434 MHz microwave applicators. Bolus cooling enabled semi-superficial target depths (1-4 cm), thermocouple thermometry monitored both skin and intratarget temperatures. The present study analyzes relationships between LRC and average skin and intratarget median temperature T50 and best session with highest median thermal dose CEM43T50 (cumulative equivalent minutes at 43 °C). Pearson correlation coefficient analysis evaluated correlation between intratarget and skin temperature parameters. RESULTS: Average median skin and invasive temperatures were T50 = 40.6 °C (Interquartile Range IQR 40.3-40.8) and T50 = 40.7 °C (IQR 40.0-41.3), respectively, without association between skin and intratarget temperatures. Median CEM43T50 for skin and intratarget were 2.8 (IQR 2.0-4.3) and 7.2 min (IQR 3.4-15.9) in the best hyperthermia session, respectively. Twenty-four patients developed infield recurrences; median time-to-recurrence was 3.4 years (IQR 2.7-4.6 years). Univariate and multivariate analysis showed significant association of intratarget temperature parameters with LRC (p = 0.015; p = 0.021, respectively), but not for skin temperature parameters and LRC (p = 0.131; p = 0.215, respectively). CONCLUSION: Temperature monitoring using both skin and invasive intratarget measurements is key for ensuring good quality superficial hyperthermia delivery for tumor depths ≥1cm.
Human vaccines & immunotherapeuticsRong Jiang, May Z Gao, Vincent M D'Anniballe, Matthew Lu, Tammara L Watts, Gregory D Zimet, Darien J Weatherspoon, Erika L Thompson, Nosayaba Osazuwa-Peters
Human papillomavirus (HPV) infection contributes to rising incidence of HPV-related oropharyngeal cancer (OPC). After the U.S. expanded HPV vaccination to adults aged 27-45 y in 2018, COVID-19 caused preventive visits and routine immunizations to plunge. We analyzed 2019 and 2022 NHIS data among adults aged 27-45 (2019 n = 8,745; 2022 n = 7,303). The primary outcome was ever HPV vaccination (≥1 dose), regardless of age at first vaccination. We estimated age-adjusted, survey-weighted coverage and used multivariable logistic regression to estimate adjusted odds of vaccination by sociodemographic characteristics; secondary 2022 analyses assessed initiation between ages 27 and 45. Ever HPV vaccine uptake increased from 15.1% in 2019 to 20.7% in 2022 (difference 5.6 percentage points; 95% CI, 4.03-7.07). In both years, lower odds were observed with increasing age, men, non-Hispanic Asian adults, individuals with lower educational attainment, and those without a usual place for care. By 2022, lower income-to-poverty ratios and lack of insurance were associated with reduced odds, whereas LGB+ and unmarried adults had higher odds, and adults in the Northeast had higher odds than those in the South. Only 1.43% (95% CI, 1.07-1.80) initiated vaccination between ages 27 and 45 in 2022. Initiation was more common among females, LGB+ and non-Hispanic Asian adults, and unmarried adults, and less common among those with lower education and no usual place for care. The increase in lifetime HPV vaccine coverage likely reflects childhood and adolescent vaccination rather than initiation at ages 27-45, which remained uncommon, and persistent inequities highlight missed opportunities for equitable OPC prevention.
Gynecological endocrinology : the official journal of the International Society of Gynecological EndocrinologyXiangyan Ruan, Juan Du, Muqing Gu, Fengyu Jin, Jing Jin, Yanglu Li, Jiaojiao Cheng, Yanqiu Li, Weimin Kong, Qi Zhang, Zecheng Wang, Mingzhen Zhang, Anming Liu,…
OBJECTIVE: This article reports on the world's first live birth after ovarian tissue cryopreservation and transplantation (OTCT) in a patient with colon cancer. METHODS: A 29-year-old unmarried nulliparous patient underwent ovarian tissue cryopreservation before chemotherapy. After chemotherapy-induced premature ovarian insufficiency, she received the first transplantation of six thawed ovarian cortex strips. Despite normal ovarian function being maintained for over 2 years, she requested and underwent a second transplantation with another six strips to maximize her chances of pregnancy. RESULTS: After the first transplantation, she resumed spontaneous menstruation with restored ovarian function, which was remained functional for 28 months until the second transplantation. Following the second transplantation, she conceived spontaneously without assisted reproductive technology two months after grafting. Routine prenatal examinations showed normal fetal development without abnormalities. She delivered a healthy baby girl weighing 2785 g with Apgar scores of 8/9 at 1 and 5 min. CONCLUSIONS: This case demonstrates the feasibility and safety of OTCT for fertility and ovarian function preservation in patients with colon cancer.
Annals of medicinePeixin Wang, Mengyu Zhang, Yucheng Lai, Tao Song, Yan Shen, Haiying Chen, Xiaoyan Chen
BACKGROUND: PAX1/JAM3 methylation are promising biomarkers for high-grade cervical intraepithelial neoplasia (CIN) detection, but menopausal status may affect diagnostic accuracy. This retrospective study compared their clinical performance between premenopausal and postmenopausal women undergoing colposcopy. METHODS: 705 women undergoing colposcopy for abnormal screening results and/or clinical symptoms were stratified by menopausal status, including 192 postmenopausal and 513 premenopausal women. Subjects were further classified into ≤CIN1 and CIN2+ groups according to histopathology. Diagnostic and triage utility of PAX1/JAM3 methylation for identifying CIN2+ lesions were compared between premenopausal and postmenopausal women. RESULTS: PAX1/JAM3 methylation demonstrated superior diagnostic performance for CIN2+ detection in postmenopausal women compared with premenopausal women, either alone or in combination. Sensitivities of PAX1m and JAM3m in postmenopausal women (90.00%, 95% CI: 76.34-97.21%; 82.50%, 95% CI: 67.22-92.66%) were higher than those in premenopausal women (61.98%, 95% CI: 52.71-70.65%; 60.33%, 95% CI: 51.04-69.11%) (all p < 0.05). The combination improved sensitivity to 72.73% in premenopausal women and maintained 92.50% in postmenopausal women. Besides, PAX1/JAM3 methylation outperformed cytology and HPV16/18 testing in both groups (p < 0.001). In HPV16/18-positive or cytology triage, methylation increased positive predictive values (PPVs) for CIN2+/CIN3+ and reduced colposcopy referrals . However, CIN2+ missed rates in premenopausal women (27.54% for HPV16/18-positive triage and 23.85% for cytology triage) were higher than those in postmenopausal counterparts. CONCLUSIONS: The diagnostic performance of PAX1/JAM3 methylation for high-grade cervical lesions differs distinctly between premenopausal and postmenopausal women undergoing colposcopy. Menopausal status should be considered when applying methylation biomarkers, particularly as a valuable auxiliary tool to improve diagnostic accuracy in elderly women.
Annals of medicineXinyun Tu, Youni Zhang, Li Wang, Yiyi Shan, Lihong Xie, Huafeng Shou, Yan Liang
BACKGROUND: Ovarian, cervical and endometrial cancers have been the leading cause of morbidity and mortality in women, often resulting from late diagnosis, resistance to therapies and off-target toxicity of conventional therapies. DISCUSSION: The review discusses recent progress in the development of dual-targeted nanocarrier systems in gynaecologic oncology, relying on peer-reviewed publications in scientific databases published within the past decade. Applications of these systems are also discussed in terms of the potential to improve the selectivity and therapeutic efficiency of tumour therapy. Major available evidence is based on preclinical in vitro and in vivo models. Clinical translation is in its early-phase trials, suggesting clinical validation in gynaecologic oncology remains in its infancy. The most important challenges in translation are scalable manufacturing, long-term safety, biological complexity and regulatory challenges. CONCLUSION: This review describes the prevailing situation and constraints of dual-targeted nanotechnology, focusing on possible applications in the development of more accurate and individualized therapies for gynaecologic cancers.
BACKGROUND: Evidence regarding repeated remimazolam administration remains limited. We hypothesized that remimazolam would be associated with fewer adverse events than propofol while maintaining stable sedative requirements across three repeated sedation sessions. METHODS: Patients with cervical cancer undergoing three fractions of interstitial brachytherapy were randomized to propofol or remimazolam. The primary outcome was the proportion of sedation episodes with at least one prespecified intraoperative adverse event across three sessions. Secondary outcomes included sedative dose, hemodynamics, and sleep quality assessed using the Pittsburgh Sleep Quality Index (PSQI) and Athens Insomnia Scale (AIS). RESULTS: A total of 100 participants were randomized, of whom 91 completed all three prespecified sedation sessions and were included in the primary per-protocol analysis (46 in the propofol group and 45 in the remimazolam group). Across 138 propofol and 135 remimazolam episodes, overall adverse events were less frequent with remimazolam (17.78% vs. 59.42%; RR, 0.30; 95% CI, 0.18-0.49; p < 0.001). Respiratory depression (3.70% vs. 26.09%; RR, 0.14; 95% CI, 0.04-0.53; p = 0.004) and hypotension (8.15% vs. 41.30%; RR, 0.20; 95% CI, 0.09-0.42; p < 0.001) were also lower with remimazolam. Remimazolam requirements remained stable across sessions (third vs. first: mean difference, 0.19 mg; 95% CI, -0.78 to 1.15; p = 0.846). No significant between-group differences were observed in PSQI or AIS scores. CONCLUSION: Across three consecutive sedation sessions for interstitial brachytherapy, remimazolam was associated with fewer respiratory and hemodynamic adverse events than propofol, while sedative requirements remained stable across the three sessions. TRIAL REGISTRATION: Chinese Clinical Trial Registry (identifier: ChiCTR2500108294).
Human vaccines & immunotherapeuticsMuhammad Inam, Muhammad Jamshed, Abuzar Osman
Nanotechnology has transformed vaccine development by engineering nanovaccines that offer precise control over antigen performance, immune modulation and targeted delivery. These platforms influence physicochemical properties size, charge, and ligand expression to direct antigen trafficking, co-deliver adjuvants and antigen-presenting cells, stimulating potent and robust adaptive immune responses. This review discusses the development of nanovaccines for prophylactic vaccines which induce antibody and memory B-cell responses and therapeutic cancer vaccines, which overcome tolerance and immunosuppression to activate CD8+ T cells with particular emphasis on lipid nanoparticles (LNPs), polymeric NPs, inorganic-core NPs, virus-like particles (VLP) and mechanisms by which each platform exploits physicochemical properties to enhance the antigen encapsulation, adjuvant incorporation, lymphatic transferring, quality and quantity of adaptive immune responses. We reviewed the clinical and preclinical developments of these nanovaccine platforms in cancer and infectious disease immunotherapy, including major successes and new opportunities that will shape the future of vaccines for prevention and treatment.
OncoimmunologyGuido Kroemer, Xiaolian Deng, Baldur Sveinbjørnsson, Oliver Kepp
In a recent paper published in Nature, Yuan et al. described an acid-responsive membranolytic peptide that temporally separates lysosomes from later plasma membrane rupture, thereby converting oncolysis into immunogenic cell death. This elegant design permits systemic administration, elicits antigen-specific immunity, and sensitizes tumors to programmed death ligand 1 blockade with remarkably limited toxicity.
OncoimmunologyJuan Zafra-Martin, Juan Luis Onieva, Herminda Jimenez-Rodriguez, Beatriz Martinez, Laura Figueroa-Ortiz, Alicia Roman, Rafael Ordoñez, Elisabeth Pérez-Ruiz, An…
Stereotactic ablative radiotherapy (SABR) may extend the clinical benefit of immune checkpoint inhibitors (ICI) in patients with oligoprogressive disease. However, individual benefit varies, and validated biomarkers to guide patient selection are lacking. We hypothesize that cell-free DNA (cfDNA) and the neutrophil-to-lymphocyte ratio (NLR) may be associated with outcomes after SABR in this setting. This prospective observational study included patients with oligoprogression under ICI therapy who received concomitant SABR to all oligoprogressive lesions (cohort A). Cohort B was a parallel biomarker cohort of oligometastatic patients receiving only SABR. Blood samples were collected before SABR (T1), after the first (T2) and last (T3) fractions, and two months after SABR (T4). The objective response rate (ORR) was evaluated by iRECIST in all lesions. Prespecified cfDNA and NLR time points were explored in relation to modified progression-free and overall survival (OS). We assessed 104 patients. With a median follow-up of 12 months, ORR was 59% in cohort A and 65% in cohort B. In cohort A, cfDNA <0.37 ng/µL at T3 was associated with improved 1-year OS (91% vs 62%, p = 0.031), while NLR <1.8 at T1 was associated with improved 1-year OS (100% vs 70%, p = 0.01). In cohort B, a >3% increase in cfDNA from T2 to T4 and NLR <4 at T4 were associated with improved OS. These findings support further evaluation of cfDNA and NLR as accessible biomarkers for patient stratification in oligoprogressive disease treated with SABR while maintaining an ICI.
Hematology (Amsterdam, Netherlands)Sinian Li, Mingyue Xiong, Zhonggui Gan, Haishan Lu, Yiheng Zhao, Hong Huang, Tingzhuang Yi
BACKGROUND: Hodgkin lymphoma (HL) survivors face an elevated long-term risk of developing second primary malignancies (SPMs), yet accurate prediction tools remain scarce. This study aimed to develop and internally validate a machine learning-based prediction model for SPM risk in HL survivors using a competing risk framework. METHODS: We analysed 57,429 patients diagnosed with HL between 2006 and 2020 from the Surveillance, Epidemiology, and End Results (SEER) database. Patients were randomly divided into training (n = 40,201, 70%) and internal validation (n = 17,228, 30%) cohorts. A random survival forest (RSF) model was developed to predict SPM occurrence, with death without SPM treated as a competing event. Model performance was compared against the traditional Fine-Grey subdistribution hazard regression. Discrimination was assessed using time-dependent area under the receiver operating characteristic curve (AUC), and calibration was evaluated through calibration plots. Clinical utility was determined via decision curve analysis (DCA). SHapley Additive exPlanations (SHAP) analysis was employed to interpret the model. RESULTS: During follow-up, 4,954 patients (8.6%) developed SPMs, while 9,750 (17.0%) died without SPM. The RSF model demonstrated superior discriminative ability compared to Fine-Grey regression across all time points: AUC at 3 years (0.721 vs. 0.549), 5 years (0.761 vs. 0.555), and 10 years (0.804 vs. 0.553). Calibration plots indicated good agreement between predicted and observed risks. DCA confirmed the RSF model provided greater net clinical benefit across a wide range of threshold probabilities. SHAP analysis identified age as the most influential predictor, followed by disease stage, radiotherapy status, race, sex, and chemotherapy. CONCLUSIONS: The RSF-based machine learning model accurately predicts SPM risk in HL survivors within a competing risk framework, substantially outperforming traditional regression approaches. This interpretable prediction tool may facilitate personalised surveillance strategies and risk-adapted follow-up for HL survivors.
BACKGROUND: Germinal centre B-cell-like diffuse large B-cell lymphoma (GCB-DLBCL) generally has a better prognosis than activated B-cell-like DLBCL, yet a clinically significant subset of patients still develops aggressive disease, suggesting marked malignant cellular heterogeneity within this subtype. This study aimed to characterise malignant cell states in GCB-DLBCL and assess their prognostic relevance. METHODS: Single-cell transcriptomic data were integrated with inferred copy-number variation (CNV) analysis to characterise malignant B-cell states. Regulatory programmes, functional features, and genetic associations were investigated using regulon analysis, functional profiling, and scPAGWAS. Single-cell-derived signatures were projected onto three independent bulk transcriptomic cohorts by deconvolution to evaluate clinical relevance. Integrative analyses were then performed to identify candidate molecular features associated with the M-C2 state. RESULTS: Marked malignant-cell heterogeneity was identified. A high-CNV compartment displayed dark-zone-like transcriptional programmes, enrichment of proliferation- and survival-associated features, and reduced antigen-presentation capacity. Subclustering resolved three malignant states, among which M-C2 showed the highest inferred CNV burden and a distinct germinal-centre regulatory programme marked by enhanced BCL6, TCF3, and PAX5 activity. M-C2 was enriched for DLBCL susceptibility-associated signals, and its higher abundance was consistently associated with inferior overall survival across independent bulk cohorts. Integrative prioritisation identified an eight-gene M-C2-associated panel comprising IRF4, AKT1S1, PIEZO1, POLR1A, EML6, TTN, FCRL5, and IGHG2. An exploratory prognostic model integrating molecular and clinical features further supported the potential predictive value of this programme. CONCLUSIONS: This study identifies a high-CNV, dark-zone-enriched malignant cell state within GCB-DLBCL, represented by the M-C2 subpopulation, associated with impaired antigen presentation and adverse prognosis. This framework improves understanding of malignant heterogeneity and warrants further validation for risk stratification.
OncoimmunologyMehmet Haluk Yucel, Maral Martin Mildanoglu, Ebru Engin Delipoyraz, Hakan Ozcelik, Erdem Sunger, Sena Fidan, Cihat Terzioglu, Tansel Cakir, Harun Muglu, Burcin…
Growing evidence suggests that circadian timing influences the efficacy of immune checkpoint inhibitors across multiple malignancies. However, evidence in extensive-stage small cell lung cancer (ES-SCLC) remains limited to a single East Asian cohort, and independent validation in other populations is lacking. We conducted a retrospective single-center cohort study of 107 patients with ES-SCLC treated with first-line platinum-etoposide plus atezolizumab or durvalumab. The time of day of administration (ToDA) was defined as the median start time of the first four immunotherapy infusions, and patients were categorized according to the cohort median ToDA. The primary endpoints were progression-free survival (PFS) and overall survival (OS). The cohort median ToDA was 11:54. Patients treated before the median ToDA had significantly longer median PFS (9.07 vs. 7.23 months, p = 0.002) and OS (21.17 vs. 11.43 months, p = 0.002) than those treated later. In multivariable Cox regression analysis, later administration remained independently associated with shorter PFS (adjusted HR 1.63, 95% CI 1.03-2.58; p = 0.038) and OS (adjusted HR 1.65, 95% CI 1.00-2.71; p = 0.049), whereas objective response rates, treatment-related toxicity, and immune-related adverse events were comparable between groups. Morning administration of first-line chemoimmunotherapy was associated with significantly longer survival without increased toxicity in patients with ES-SCLC. These findings support growing evidence that circadian timing may influence immune checkpoint inhibitor efficacy and warrant prospective evaluation of treatment timing as a readily modifiable strategy to optimize immunotherapy delivery.
Cancer biology & therapyLi Zhou, Jin Ke, Shanlan Zhang, Sufang Xiao, Hui Zhang, Shan Liu, Li Liu, Mingan Liu, Zuohua Lu, Songguo Zheng
BACKGROUND: Triple-negative breast cancer (TNBC) is a highly aggressive breast cancer with high brain metastatic (BM) potential. Tumor-derived exosomes are implicated as key modulators during the formation of the pre-metastatic niche (PMN). However, the regulation of TNBC-BM-derived exosomes on brain PMN remains enigmatic. METHODS: The morphology and uptake of exosomes were identified using transmission electron microscopy and live-cell imaging, respectively. Differentially secreted miRNAs and mRNA were identified using high-throughput sequencing. The permeability of the blood‒brain barrier (BBB) and tight junction integrity were examined using immunofluorescence staining. Cell viability was examined using the CCK-8 assay. The apoptosis and intracellular ROS were investigated using fluorescence staining. The expression of mRNAs was examined using qPCR. RESULTS: miR-103b was significantly increased in exosomes derived from TNBC-BM cells. miR-103b compromised the tight junctions of HUVECs and increased BBB permeability in mice. Additionally, miR-103b promoted apoptosis and increased intracellular ROS in HUVECs and U251 cells, two cell lines commonly used as surrogates for brain microvascular endothelial cells and astrocytes, respectively. miR-103b-regulated differentially expressed genes modulated tight junction, apoptosis, oxidative stress, inflammation, and metabolism. Target analysis identified 216 targets of miR-103b, among which hub targets were associated with biological processes involved in maintaining the BBB and the glucose metabolism signaling pathway. CONCLUSION: Collectively, our findings for the first time reveal that exosomal miR-103b from TNBC-BM cells potentially regulates the two major cellular constituents of the brain PMN, highlighting its role in the formation of the brain PMN during TNBC-BM and its potential as a promising therapeutic target.
Contemporary models of tumor progression highlight the key role of the tumor microenvironment (TME) and the extracellular matrix (ECM) in regulating angiogenesis, metastasis, and immune resistance. Among ECM components, oncofetal fibronectin (FN) isoforms containing the EDA and EDB domains are of particular interest because they are highly expressed in solid tumors but absent from normal adult tissues. This review article synthesizes current data on the pathophysiological functions of FN variants in oncogenesis. Particular attention was paid to their role in pathological vascular remodeling, the induction of epithelial-mesenchymal transition (EMT), the formation of pre-metastatic niches, and the generation of an immunosuppressive environment. The study also critically assesses the clinical significance of FN as a biomarker and a potential target for new therapies in breast cancer, ovarian cancer and glioblastoma. The prospects for the use of innovative stromal strategies - including antibody-drug conjugates (ADCs) and immunocytokines - to overcome resistance to standard immunotherapy are also highlighted.
Biochimica et biophysica acta. Molecular basis of diseaseYongqing Zhao, Jian Yang, Weixiong Zhu, Guoqing Zhang, Shaozhen Rui, Teng Ma, Zhao Guo, Wence Zhou
BACKGROUND: The global burden of metabolic-associated hepatocellular carcinoma (HCC) is increasing, with obesity emerging as a key causal factor. However, the molecular mechanisms linking lipid metabolic dysregulation to HCC progression and therapeutic vulnerability remain unclear. METHODS: We analyzed Global Burden of Disease 2021 data to assess liver cancer burden attributable to metabolic risks from 1990 to 2021. Mendelian randomization was used to evaluate causal associations between metabolic traits and liver cancer risk. TCGA, GTEx, and GEO datasets were integrated to identify lipid stress-responsive regulators. Clinical relevance was assessed using public datasets and tissue microarray immunohistochemistry. Functional validation was performed in HCC cells and a high-fat diet-fed syngeneic mouse tumor model. RESULTS: Liver cancer deaths and DALYs attributable to metabolic risks increased markedly from 1990 to 2021. Mendelian randomization showed that obesity-related traits, including BMI, waist circumference, and body fat percentage, were causally associated with liver cancer risk, whereas glycemic traits were not. Bioinformatics screening identified GPAT3 as a lipid metabolism regulator upregulated in HCC, induced by palmitic acid, associated with poor prognosis, and enriched in patients with higher BMI. Tissue microarray analysis confirmed increased GPAT3 protein expression in HCC and its association with higher BMI and GPX4 expression. GPAT3 depletion sensitized HCC cells to palmitic acid-induced ferroptosis, whereas Fer-1 rescue and GPAT3 overexpression supported its protective role. In vivo, FSG67 enhanced sorafenib-associated antitumor effects and increased tumor lipid peroxidation. CONCLUSIONS: GPAT3 protects HCC cells from lipid stress-induced ferroptosis and represents a potential metabolic vulnerability in obesity-associated HCC.
OBJECTIVES: This study aimed to investigate Investigate the expression level, prognostic value, and biological function of miR-877-5p in MM. METHODS: A total of 103 MM patients and 98 non-tumor controls were included in the study. miR-877-5p expression level was verified by RT-qPCR, and its prognostic value was evaluated through Kaplan-Meier and the Cox regression model. MM cell proliferation and apoptosis were assessed with the CCK‑8 assay and flow cytometry. The potential target genes of miR-877-5p were predicted by bioinformatics analysis and verified by dual luciferase assay. Rescue assays confirmed that MAPK8 mediates the regulatory function of miR-877-5p. RESULTS: miR-877-5p was reduced in MM, and its low level was associated with a poorer overall survival of patients, being an independent prognosticator of poor survival in MM. Function experiments confirmed that overexpression of miR-877-5p could inhibit the proliferation of MM cells and promote apoptosis. Mechanistically, MAPK8 was a direct downstream target of miR-877-5p. MAPK8 was upregulated in MM and negatively correlated with miR-877-5p levels. Rescue assays were conducted to verify that MAPK8 overexpression reversed the effect of miR-877-5p on MM cell proliferation and apoptosis. DISCUSSION: These findings indicate that miR-877-5p exerts tumor-suppressive functions in MM, likely through negative regulation of MAPK8. The miR-877-5p/MAPK8 axis may represent a novel regulatory pathway involved in MM progression. CONCLUSIONS: miR-877-5p acts as a tumor suppressor in MM and may serve as a potential biomarker for prognosis assessment. Its regulatory role in MM progression is likely mediated through targeting and inhibiting MAPK8.
Pancreatic ductal adenocarcinoma (PDAC) remains one of the most lethal solid tumors due to its dense desmoplastic stroma, poor vascular perfusion, and the limited intratumoral delivery of chemotherapeutic agents such as oxaliplatin (OXA). Extracellular vesicles (EVs) offer a biocompatible platform for drug delivery, yet their intrinsic lack of tumor specificity constrains therapeutic efficacy. Ephrin type-A receptor 2 (EphA2), which is highly expressed and efficiently internalized in PDAC, represents an attractive molecular target for guiding EVs into tumor cells. In this study, we engineered HEK293T-derived EVs to display membrane-anchored anti-EphA2 Fab fragments and encapsulate OXA (EVs-EphA2/OXA) as a targeted delivery system for PDAC therapy. Stable producer cells expressed the engineered Fab on the plasma membrane and released vesicles that retained canonical EV markers and robust antigen-binding capability. EVs-EphA2 demonstrated selective uptake into EphA2-positive AsPC-1 and BxPC-3 cells and induced significantly greater cytotoxicity than free OXA or untargeted EVs/OXA in vitro. In xenograft models, EVs-EphA2/OXA achieved the most pronounced tumor suppression, accompanied by increased γH2AX-associated DNA damage, enhanced TUNEL-positive apoptosis, and preferential tumor accumulation in biodistribution imaging. These findings demonstrate that EphA2-targeted EVs can substantially improve intratumoral delivery of OXA and amplify antitumor efficacy, supporting EVs-EphA2/OXA as a promising platform for receptor-guided chemotherapy in PDAC.