Patient-derived organoids (PDOs) have emerged as physiologically relevant cancer models that preserve key genetic, histological, and functional features of the tumors from which they are derived. In parallel, CRISPR-based perturbation technologies have transformed functional genomics by enabling scalable interrogation of gene function. Their integration provides a powerful framework for identifying cancer dependencies, modeling oncogenic evolution, and investigating mechanisms of drug response and resistance in patient-relevant settings. This review examines how CRISPR knockout, CRISPR interference/activation, and precision editing approaches have been applied in PDO systems to uncover context-specific vulnerabilities, reconstruct mutational trajectories, and study tumor heterogeneity. We further compare pooled and arrayed screening formats and discuss what is uniquely enabled by performing CRISPR screens in organoids rather than conventional 2D models. Particular emphasis is placed on the technical and analytical constraints of organoid-based screening, including variable editing efficiency, clonal bottlenecks, biological heterogeneity, and limited scalability. We argue that the major value of organoid-based CRISPR screening lies in its ability to identify functionally actionable cancer vulnerabilities in a patient-contextualized model, while also introducing methodological challenges that must be addressed for robust clinical translation.

Circulating tumor DNA (ctDNA) monitoring may detect resistance to KRAS^G12C inhibitors earlier than conventional imaging. However, in the specific context of KRAS^G12C inhibitors, the temporal relationship between molecular, radiologic, and clinical progression remains poorly characterized. A 66-year-old woman with KRAS^G12C/STK11/KEAP1-mutant lung adenocarcinoma received second-line adagrasib after progression on chemo-immunotherapy. Serial ctDNA monitoring by droplet digital PCR and targeted next-generation sequencing was performed. Baseline KRAS^G12C ctDNA (6205.3 copies/mL) declined to 3.2 copies/mL after 7 weeks, indicating initial response. However, ctDNA levels rose to 21.1 copies/mL at month 4 and 315.2 copies/mL at month 6. Concurrent NGS revealed three emergent KRAS mutations (G12D, G13D, Q61H), confirming polyclonal resistance. Notably, CT and MRI imaging remained stable by RECIST criteria throughout this period. Significant clinical deterioration, requiring prolonged hospitalization, occurred approximately 8 weeks after initial ctDNA rise. The patient died 2 months after detection of resistance mutations. In this case, NGS ctDNA detected three acquired KRAS resistance mutations in addition to the original G12C, during a prolonged period of radiographic stability, illustrating polyclonal evolution under adagrasib pressure. These findings suggest that longitudinal liquid biopsy monitoring may provide early detection of resistance in KRAS^G12C-mutant NSCLC.

To investigate the expression profile of IL-6 in patients with acute promyelocytic leukemia (APL) and its role in modulating the ability of hPMSCs to regulate the differentiation of NB4 cells.

Peripheral blood samples were obtained from newly diagnosed APL patients and healthy controls. IL-6 levels were measured using flow cytometry-based (FCM) multiplex immunoassays. A co-culture system of hPMSCs and NB4 cells was established. The effects of hPMSCs on ATRA-induced differentiation, proliferation, and phagocytic activity of NB4 cells were evaluated by Wright-Giemsa staining, FCM, CCK-8 assays, and phagocytosis assays. Western blot analysis was performed to investigate the signaling mechanisms by which IL-6 regulates CD73 expression in hPMSCs. To determine whether the regulatory effects of hPMSCs are mediated by CD73, interventions using a CD73 inhibitor and CD73-knockdown hPMSCs were conducted.

IL-6 levels were significantly elevated in the peripheral blood of APL patients and showed a positive correlation with both risk stratification and the severity of retinoic acid syndrome (RAS). hPMSCs enhanced ATRA-induced differentiation of NB4 cells while suppressing their proliferation and phagocytic function. Moreover, inhibition or knockdown of CD73 markedly attenuated the pro-differentiation effect of hPMSCs on NB4 cells. NB4 cells exhibited high surface expression of adenosine receptors. IL-6 was found to upregulate CD73 expression in hPMSCs through activation of the PI3K/AKT signaling pathway.

IL-6 enhances the pro-differentiation capacity of hPMSCs toward NB4 cells by upregulating CD73 expression, offering a potential new strategy for the clinical treatment of APL.

BACKGROUND Thoracic SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 4 (SMARCA4)-deficient undifferentiated tumor (SMARCA4-UT) is a rare and highly aggressive thoracic malignancy. It predominantly affects male smokers and typically arises in the mediastinum, where rapid tumor growth and early metastasis contribute to a poor prognosis. Recent evidence suggests that this tumor exhibits biological heterogeneity and variable responses to therapy, underscoring the need for further clinical characterization. CASE REPORT A 67-year-old man presented with dyspnea and superior vena cava syndrome. Computed tomography revealed a 7-cm mass in the anterior and superior mediastinum with suspected lymph node and bone metastases. Histological examination demonstrated a sheet-like proliferation of large atypical cells with rhabdoid features and necrosis. Immunohistochemical analysis showed loss of SMARCA4 (Brahma-related gene 1 [BRG1]) expression, positivity for cluster of differentiation 34 (CD34) and sex-determining region Y-box 2 (SOX2), weak epithelial membrane antigen expression, preserved integrase interactor 1 (INI1) expression, and negativity for other epithelial markers, fulfilling the diagnostic criteria for SMARCA4-UT. Despite palliative radiotherapy and combination immunotherapy with nivolumab and ipilimumab, the tumor rapidly progressed. The patient developed grade 4 drug-induced pneumonitis; transient stabilization was achieved, but his condition deteriorated. He died 6 months after disease onset. Autopsy revealed widespread metastases with minimal therapeutic effect, highlighting the aggressive clinical course and treatment resistance. CONCLUSIONS SMARCA4-UT is a highly aggressive tumor requiring comprehensive immunohistochemical evaluation for accurate diagnosis. This case highlights the limited efficacy of immune checkpoint inhibitors in a PD-L1-negative setting and underscores the need for more effective therapeutic strategies.

The tumour microenvironment (TME) of relapsed/refractory (R/R) diffuse large B-cell lymphoma (DLBCL) patients is associated with resistance of DLBCL cells to CD19 CAR-T cells. How to improve TME in DLBCL and improve the efficacy of CAR-T cell therapy remains to be further explored. We observed the sensitivity of HBL-1/U2932 cells pretreated with BTK inhibitors (BTKi) to CAR-T cells with flow cytometry (FCM). The effect of pretreatment of BTKi on the polarisation state of alternative activated M2 macrophages was observed with FCM, real-time PCR and Western blot method. The effect of Notch1 agonist on expressions of Arg-1 protein, iNOS protein, Notch1 protein and RBP-J protein in alternative activated M2 macrophages was observed by Western blot method. Then the expression consistency of Notch-1 and RBP-J in activated M2 macrophages was observed by siRNA transfection of Notch-1. The cytotoxicity of CD19 CAR-T cells on HBL-1/U2932 cells pretreated with ibrutinib/orelabrutinib was higher than that of HBL-1/U2932 cells unpretreated with ibrutinib/orelabrutinib. Cytotoxicity of CAR-T cells to HBL-1 cells in coculture system with alternative activated M2 macrophages was very low. This drug resistance could be reversed by replacing the M2 macrophages (M2 macrophages after 48 h pretreatment with BTKi) in coculture system. Pretreatment with BTKi could down-regulate the expression of CD206 and IL-10 in activated M2 macrophages. And pretreatment with BTKi down-regulated the expression of Arg-1 and upregulated the expression of iNOS in activated M2 macrophages. The upregulation polarisation of M2 macrophages by Notch1 agonist could be reversed by BTKi. Expression of RBP-J protein decreased in alternative activated M2 macrophages by siRNA silencing Notch 1. Pretreatment with BTKi could down-regulate the polarisation of M2 macrophages and reverse the resistance of DLBCL cells which were cocultured with alternative activated M2 macrophages to CAR-T cells. This effect might be achieved by downregulating the Notch-RBP-J pathway.

WHO grade 2 meningiomas exhibit highly heterogeneous clinical courses. While the Ki67 proliferation index is a standard biomarker, its prognostic utility remains limited by methodological inconsistency and potential time-dependent dynamics. We evaluated an automated, artifact-adjusted Ki67 assessment and its integration with molecular risk profiling. 98 WHO grade 2 meningiomas (WHO 2021) were analyzed using an automated QuPath-based pipeline with HistoART for artifact exclusion. Molecular risk was defined by methylation and copy number profiling to calculate the integrated molecular-morphologic risk score by Maas et al. We employed extended Cox models to account for proportional hazards violations. Automated Ki67 values were significantly lower than routine pathological estimates (median 2.91% vs. 10%; p < 0.001) and correlated modestly with integrated risk scores (ρ = 0.26, p = 0.009). We identified a biphasic risk pattern: within the first 38 postoperative months, an automated Ki67 > 3.62% was a strong independent predictor for local recurrence (HR 5.06, p < 0.001) and progression-free survival (HR 4.15, p = 0.002), remaining significant alongside subtotal resection and the integrated risk group. Beyond 38 months, prognostic impact attenuated. Ki67 and the integrated molecular risk score contributed independently in multivariable models, suggesting complementary biological dimensions. Automated, artifact-adjusted Ki67 quantification provides time-dependent, independent prognostic information in WHO grade 2 meningioma, complementary to molecular risk stratification. It may serve as a cost-effective surveillance marker-both as an adjunct to molecular profiling and as a standalone tool where molecular testing is unavailable.

Complete hydatidiform mole with a coexisting live fetus in a twin pregnancy is exceptionally rare and carries major maternal and fetal risks. We describe two such pregnancies to illustrate management options. The first followed frozen-thawed single blastocyst transfer and presented at 13 weeks with a live fetus, a separate molar mass, an intramural fibroid, markedly elevated serum beta-human chorionic gonadotropin, and biochemical hyperthyroidism. Multidisciplinary assessment led to hysterotomy with removal of the fetus, molar tissue, and fibroids, followed by rapid recovery and no gestational trophoblastic neoplasia. The second was a spontaneously conceived twin pregnancy with a live 9-week fetus and multicystic molar tissue that was evacuated by suction curettage. An initial decline in beta-human chorionic gonadotropin was followed by a low-level plateau that resolved after a single low-dose methotrexate cycle. These cases highlight the need for individualized, multidisciplinary care and close biochemical follow-up.

HPV18-positive cervical large cell neuroendocrine carcinoma (LCNEC) is exceptionally rare. We report a 40-years-old woman with FIGO 2018 stage IVB LCNEC who progressed rapidly despite paclitaxel-carboplatin-bevacizumab plus pembrolizumab, followed by concurrent chemoradiotherapy and later cytotoxic regimens. Biopsy showed chromogranin A and synaptophysin positivity, diffuse p16, and Ki-67%~80%. Comprehensive profiling revealed very low tumor mutational burden (TMB, 1.21/Mb), microsatellite stability, no homologous recombination deficiency, and minimal PD-L1 (~1%), consistent with a poorly immunogenic tumor microenvironment (TME). This profile may explain the limited effect of immune checkpoint inhibition in this histology. The case suggests that HPV or p16 positivity alone did not predict clinical benefit from pembrolizumab-based therapy in advanced cervical LCNEC, and supports careful biological assessment before treatment selection. These observations provide practical guidance for clinicians and support evaluation of alternative or investigational strategies for this aggressive tumor.

Colorectal cancer (CRC) is a leading cause of cancer-related morbidity and mortality worldwide; thus, understanding its molecular mechanisms is critical for developing novel therapeutic targets. Long non-coding RNAs (lncRNAs) play crucial isoform-specific roles in cancer. While oncogenic lncRNA LINC00673 is known to be involved in multiple malignancies and possesses five distinct transcript variants, the functional role of LINC00673-V4-a highly expressed transcript variant in CRC-remains largely unexplored. This study investigated how LINC00673-V4 drives CRC proliferation by modulating the Hippo-Yes-associated protein (Hippo-YAP) signaling pathway.

Survival analysis was performed using the GSE39582 dataset to assess the prognostic value of LINC00673 in patients with CRC. The expression of LINC00673 transcript variants in CRC cell lines was detected by quantitative PCR. Cell Counting Kit-8 and 5-ethynyl-2'-deoxyuridine (EdU) incorporation assays were used to assess cell proliferation upon LINC00673-V4 knockdown or overexpression. Western blotting and immunofluorescence staining were utilized to analyze the expression of Hippo-YAP target genes. To confirm the interaction between LINC00673-V4 and fused in sarcoma (FUS), RNA immunoprecipitation (RIP) was performed, while co-IP was used to investigate potential protein-protein interactions.

High LINC00673 expression was observed in CRC cell lines and was associated with a poor prognosis in patients with CRC. Among the five LINC00673 transcript variants, LINC00673-V3 and LINC00673-V4 were the predominantly expressed isoforms; however, only LINC00673-V4 significantly promoted CRC cell proliferation. LINC00673-V4 inhibited YAP phosphorylation at Ser127, promoted YAP nuclear translocation, and upregulated the expression of Hippo-YAP target genes (connective tissue growth factor, cysteine-rich angiogenic inducer 61, survivin). RIP assays confirmed an association between LINC00673-V4 and FUS, while co-immunoprecipitation (co-IP) assays revealed that FUS interacted with both YAP and large tumor suppressor 1 (LATS1), suggesting FUS may facilitate LATS1-mediated YAP phosphorylation. FUS was found to promote LATS1-mediated YAP Ser127 phosphorylation, induce YAP cytoplasmic retention, and down-regulate Hippo-YAP target gene expression. Rescue experiments showed that LINC00673-V4 reversed the FUS-induced suppression of Hippo-YAP target gene expression.

Our study identified LINC00673-V4 as an isoform-specific oncogenic lncRNA in CRC. By associating with FUS, LINC00673-V4 sequesters FUS away from the LATS1/YAP complex; thereby inhibiting FUS-mediated YAP phosphorylation and enhancing Hippo-YAP target gene expression. These findings expand the landscape of lncRNA isoform-specific regulation in cancer, highlighting LINC00673-V4 as a potential prognostic biomarker and therapeutic target for CRC.

Gut microbiota dysbiosis is a crucial driver of the initiation and progression of colorectal cancer (CRC), where functional gut microbes and their metabolites play key roles in the microecological regulation of CRC. Currently, the association between Lachnospiraceae and CRC progression, as well as the underlying mechanisms, remains incompletely understood and warrants further investigation.

Bioinformatics analysis was performed to explore the co-pathway association between gut microbiota and metabolites in CRC patient samples. In vivo animal models were established to assess the regulatory effects of Lachnospiraceae on CRC tumorigenesis and gut microbiota homeostasis. The anti-CRC activities of Lachnospiraceae and its metabolite malate were investigated using in vitro experiments that measured cell viability, proliferation, apoptosis, and colony formation. Western blotting was performed to detect the expression levels of key proteins in the Wingless/Integrated (Wnt)/β-catenin signaling pathway.

Bioinformatics analysis revealed that malate was significantly downregulated in CRC patients, accompanied by gut microbiota dysbiosis driven predominantly by short-chain fatty acid (SCFA)-related Firmicutes such as Lachnospiraceae and Ruminococcaceae. In vivo, Lachnospiraceae restored gut microbiota homeostasis, reduced tumor number, and decreased tumor load. In vitro, Lachnospiraceae suppressed colorectal tumorigenesis and increased colonic D-malate levels.

Lachnospiraceae bacterium biologics abstracts accession-2278 (BAA-2278) is associated with anti-CRC effects in preclinical models, potentially mediated through regulation of gut microbiota homeostasis and inhibition of Wnt/β-catenin signaling via its metabolite malate.