3D engineered scaffold for large-scale Vigil immunotherapy production.

Previously, we reported successful cellular expansion of a murine colorectal carcinoma cell line (CT-26) using a three-dimensional (3D) engineered extracellular matrix (EECM) fibrillar scaffold structure. CCL-247 were grown over a limited time period of 8 days on 3D EECM or tissue culture polystyrene (TCPS). Cells were then assayed for growth, electroporation efficiency and Vigil manufacturing release criteria. Using EECM scaffolds, we report an expansion of CCL-247 (HCT116), a colorectal carcinoma cell line, from a starting concentration of 2.45 × 105 cells to 1.9 × 106 cells per scaffold. Following expansion, 3D EECM-derived cells were assessed based on clinical release criteria of the Vigil manufacturing process utilized for Phase IIb trial operation with the FDA. 3D EECM-derived cells passed all Vigil manufacturing release criteria including cytokine expression. Here, we demonstrate successful Vigil product manufacture achieving the specifications necessary for the clinical trial product release of Vigil treatment. Our results confirm that 3D EECM can be utilized for the expansion of human cancer cell CCL-247, justifying further clinical development involving human tissue sample manufacturing including core needle biopsy and minimal ascites samples.

Spatial distribution pattern of immune cells is associated with patient prognosis in colorectal cancer.

BACKGROUND: The spatial context of tumor-infiltrating immune cells (TIICs) is important in predicting colorectal cancer (CRC) patients' clinical outcomes. However, the prognostic value of the TIIC spatial distribution is unknown. Thus, we aimed to investigate the association between TIICs in situ and patient prognosis in a large CRC sample. METHODS: We implemented multiplex immunohistochemistry staining technology in 190 CRC samples to quantify 14 TIIC subgroups in situ. To delineate the spatial relationship of TIICs to tumor cells, tissue slides were segmented into tumor cell and microenvironment compartments based on image recognition technology, and the distance between immune and tumor cells was calculated by implementing the computational pipeline phenoptr. RESULTS: MPO+ neutrophils and CD68+IDO1+ tumor-associated macrophages (TAMs) were enriched in the epithelial compartment, and myeloid lineage cells were located nearest to tumor cells. Except for CD68+CD163+ TAMs, other cells were all positively associated with favorable prognosis. The prognostic predictive power of TIICs was highly related to their distance to tumor cells. Unsupervised clustering analysis divided colorectal cancer into three subtypes with distinct prognostic outcomes, and correlation analysis revealed the synergy among B cells, CD68+IDO1+TAMs, and T lineage cells in producing an effective immune response. CONCLUSIONS: Our study suggests that the integration of spatial localization with TIIC abundance is important for comprehensive prognostic assessment.

Boron Neutron Capture Therapy-Derived Extracellular Vesicles via DNA Accumulation Boost Antitumor Dendritic Cell Vaccine Efficacy.

Radiated tumor cell-derived extracellular vesicles (RT-EVs) encapsulate abundant DNA fragments from irradiated tumor cells, in addition to acting as integrators of multiple tumor antigens. Accumulating evidence indicates these DNA fragments from damaged cells are involved in downstream immune responses, but most of them are degraded in cells before incorporation into derived RT-EVs, thus the low abundance of DNA fragments limits immune responses of RT-EVs. Here, this study found that different radiations affected fates of DNA fragments in RT-EVs. Boron neutron capture therapy (BNCT) induced DNA accumulation in RT-EVs (BEVs) by causing more DNA breaks and DNA oxidation resisting nuclease degradation. This is attributed to the high-linear energy transfer (LET) properties of alpha particles from the neutron capture reaction of 10B. When being internalized by dendritic cells (DCs), BEVs activated the DNA sensing pathway, resulting in functional enhancements including antigen presentation, migration capacity, and cytokine secretion. After vaccination of the BEVs-educated DCs (BEV@BMDCs), the effector T cells significantly expanded and infiltrated into tumors, suggesting robust anti-tumor immune activation. BEV@BMDCs not only effectively inhibited the primary tumor growth and metastasis formation but also elicited long-term immune memory. In conclusion, a successful DC vaccine is provided as a promising candidate for tumor vaccine.

Circulating tumor cells with increasing aneuploidy predict inferior prognosis and therapeutic resistance in small cell lung cancer.

AIMS: Treatment resistance commonly emerges in small cell lung cancer (SCLC), necessitating the development of novel and effective biomarkers to dynamically assess therapeutic efficacy. This study aims to evaluate the clinical utility of aneuploid circulating tumor cells (CTCs) for risk stratification and treatment response monitoring. METHODS: A total of 126 SCLC patients (two cohorts) from two independent cancer centers were recruited as the study subjects. Blood samples were collected from these patients and aneuploid CTCs were detected. Aneuploid CTC count (ACC) and aneuploid CTC score (ACS), were used to predict progression-free survival (PFS) and overall survival (OS). The performance of the ACC and the ACS was evaluated by calculating the area under the receiver operating characteristic (ROC) curve (AUC). RESULTS: Compared to ACC, ACS exhibited superior predictive power for PFS and OS in these 126 patients. Moreover, both univariate and multivariate analyses revealed that ACS was an independent prognostic factor. Dynamic ACS changes reflected treatment response, which is more precise than ACC changes. ACS can be used to assess chemotherapy resistance and is more sensitive than radiological examination (with a median lead time of 2.8 months; P < 0.001). When patients had high ACS levels (> 1.115) at baseline, the combination of immunotherapy and chemotherapy resulted in longer PFS (median PFS, 7.7 months; P = 0.007) and OS (median OS, 16.3 months; P = 0.033) than chemotherapy alone (median PFS, 4.9 months; median OS, 13.6 months). CONCLUSIONS: ACS could be used as a biomarker for risk stratification, treatment response monitoring, and individualized therapeutic intervention in SCLC patients.

Tumor Dormancy and Reactivation: The Role of Heat Shock Proteins.

Tumors are a heterogeneous group of cell masses originating in various organs or tissues. The cellular composition of the tumor cell mass interacts in an intricate manner, influenced by humoral, genetic, molecular, and tumor microenvironment cues that dictate tumor growth or suppression. As a result, tumors undergo a period of a dormant state before their clinically discernible stage, which surpasses the clinical dormancy threshold. Moreover, as a genetically imprinted strategy, early-seeder cells, a distinct population of tumor cells, break off to dock nearby or extravasate into blood vessels to secondary tissues, where they form disseminated solitary dormant tumor cells with reversible capacity. Among the various mechanisms underlying the dormant tumor mass and dormant tumor cell formation, heat shock proteins (HSPs) might play one of the most important roles in how the dormancy program plays out. It is known that numerous aberrant cellular processes, such as malignant transformation, cancer cell stemness, tumor invasion, metastasis, angiogenesis, and signaling pathway maintenance, are influenced by the HSPs. An accumulating body of knowledge suggests that HSPs may be involved in the angiogenic switch, immune editing, and extracellular matrix (ECM) remodeling cascades, crucial genetically imprinted strategies important to the tumor dormancy initiation and dormancy maintenance program. In this review, we highlight the biological events that orchestrate the dormancy state and the body of work that has been conducted on the dynamics of HSPs in a tumor mass, as well as tumor cell dormancy and reactivation. Additionally, we propose a conceptual framework that could possibly underlie dormant tumor reactivation in metastatic relapse.

Advances in the role of circulating tumor cell heterogeneity in metastatic small cell lung cancer.

Small cell lung cancer (SCLC), a highly aggressive malignancy, is rapidly at an extensive stage once diagnosed and is one of the leading causes of death from malignancy. In the past decade, the treatment of SCLC has largely remained unchanged, and chemotherapy remains the cornerstone of SCLC treatment. The therapeutic value of adding immune checkpoint inhibitors to chemotherapy for SCLC is low, and only a few SCLC patients have shown a response to immune checkpoint inhibitors. Circulating tumor cells (CTCs) are tumor cells shed from solid tumor masses into the peripheral circulation and are key to tumor metastasis. Single-cell sequencing has revealed that the genetic profiles of individual CTCs are highly heterogeneous and contribute to the poor outcome and prognosis of SCLC patients. Theoretically, phenotypic analysis of CTCs may be able to predict the diagnostic significance of new potential targets for metastatic tumors. In this paper, we will discuss in depth the heterogeneity of CTCs in SCLC and the value of CTCs for the diagnosis and prognosis of SCLC and as relevant tumor markers in metastatic SCLC.

Establishment and characterization of a novel multidrug-resistant pancreatic ductal adenocarcinoma cell line, PDAC-X1.

Drug resistance remains a significant challenge in the treatment of pancreatic cancer. The development of drug-resistant cell lines is crucial to understanding the underlying mechanisms of resistance and developing novel drugs to improve clinical outcomes. Here, a novel pancreatic cancer cell line, PDAC-X1, derived from Chinese patients has been established. PDAC-X1 was characterized by the immune phenotype, biology, genetics, molecular characteristics, and tumorigenicity. In vitro analysis revealed that PDAC-X1 cells exhibited epithelial morphology and cell markers (CK7 and CK19), expressed cancer-associated markers (E-cadherin, Vimentin, Ki-67, CEA, CA19-9), and produced pancreatic cancer-like organs in suspension culture. In vivo analysis showed that PDAC-X1 cells maintained tumorigenicity with a 100% tumor formation rate. This cell line exhibited a complex karyotype, dominated by subtriploid karyotypes. In addition, PDAC-X1 cells exhibited intrinsic multidrug resistance to multiple drugs, including gemcitabine, paclitaxel, 5-fluorouracil, and oxaliplatin. In conclusion, the PDAC-X1 cell line has been established and characterized, representing a useful and valuable preclinical model to study the underlying mechanisms of drug resistance and develop novel drug therapeutics to improve patient outcomes.

13C Tracer Analysis and Metabolomics in Dormant Cancer Cells.

Over the last two decades, major advances in the field of tumor dormancy have been made. Yet, it is not completely understood how dormant disseminated tumor cells survive and transition to a proliferative state to generate a metastatic lesion. On the other hand, metabolic rewiring has been shown to influence metastasis development through the modulation of both intracellular signaling and the crosstalk between metastatic cells and their microenvironment. Thus, studying the metabolic features of dormant disseminated tumor cells has gained importance in understanding the dormancy process. Here, we describe a method to perform metabolomics and 13C tracer analysis in 3D cultures of dormant breast cancer cells.

Radiomics and visual analysis for predicting success of transplantation of heterotopic glioblastoma in mice with MRI.

BACKGROUND: Quantifying tumor growth and treatment response noninvasively poses a challenge to all experimental tumor models. The aim of our study was, to assess the value of quantitative and visual examination and radiomic feature analysis of high-resolution MR images of heterotopic glioblastoma xenografts in mice to determine tumor cell proliferation (TCP). METHODS: Human glioblastoma cells were injected subcutaneously into both flanks of immunodeficient mice and followed up on a 3 T MR scanner. Volumes and signal intensities were calculated. Visual assessment of the internal tumor structure was based on a scoring system. Radiomic feature analysis was performed using MaZda software. The results were correlated with histopathology and immunochemistry. RESULTS: 21 tumors in 14 animals were analyzed. The volumes of xenografts with high TCP (H-TCP) increased, whereas those with low TCP (L-TCP) or no TCP (N-TCP) continued to decrease over time (p < 0.05). A low intensity rim (rim sign) on unenhanced T1-weighted images provided the highest diagnostic accuracy at visual analysis for assessing H-TCP (p < 0.05). Applying radiomic feature analysis, wavelet transform parameters were best for distinguishing between H-TCP and L-TCP / N-TCP (p < 0.05). CONCLUSION: Visual and radiomic feature analysis of the internal structure of heterotopically implanted glioblastomas provide reproducible and quantifiable results to predict the success of transplantation.

Circulating tumor cell phenotype detection based on epithelial-mesenchymal transition markers combined with clinicopathological risk has potential to better predict recurrence in stage III breast cancer treated with neoadjuvant chemotherapy: a pilot study.

BACKGROUND: The potential value of detecting epithelial-mesenchymal transition (EMT) CTCs in early breast cancer, especially during the neoadjuvant therapy period, requires further investigation. We analyzed dynamic CTC phenotype status, to improve recurrence risk stratification for patients with stage III breast cancers. METHODS: We enrolled 45 patients with stage III breast cancers from 2 clinical trials undergoing neoadjuvant chemotherapy and utilized the CanPatrol CTC enrichment technique pre- and post-chemotherapy to identify CTC phenotypes, including epithelial CTCs, biphenotypic epithelial/mesenchymal CTCs, and mesenchymal CTCs, in peripheral blood samples. Kaplan-Meier analyses were conducted to explore the prognostic value of dynamic change of CTC count and the proportion of CTCs with different phenotypes. Then, redefine the risk stratification based on CTC status and clinicopathological risk in combination. RESULTS: Increased proportion of M + CTCs was a high-risk CTC status that was associated with decreased DFS (HR, 3.584; 95% CI, 1.057-12.15). In a combined analysis with clinicopathological risk, patients with high-risk tumors had an elevated risk of recurrence compared to patients with low-risk tumors (HR, 4.482; 95% CI, 1.246-16.12). The recurrence risk could be effectively stratified by newly defined risk stratification criteria, with 5-year DFS of 100.0%, 77.3%, and 50.0%, respectively, for low-risk, mid-risk, and high-risk patients (P = 0.0077). Finally, in the ROC analysis, the redefined risk stratification demonstrated higher predictive significance with an AUC of 0.7727, compared to CTC status alone (AUC of 0.6751) or clinicopathological risk alone (AUC of 0.6858). CONCLUSION: The proportion of M + CTCs increased after neoadjuvant chemotherapy indicating a higher risk of tumor recurrence. Combining CTC status with clinicopathological risk has potential to redefine the risk stratification of stage III breast cancers and provide improved predictions of relapse.

Hydrogel activation of Mincle receptors for tumor cell processing: A novel approach in cancer immunotherapy.

An obstacle in current tumor immunotherapies lies in the challenge of achieving sustained and tumor-targeting T cell immunity, impeded by the limited antigen processing and cross-presentation of tumor antigens. Here, we propose a hydrogel-based multicellular immune factory within the body that autonomously converts tumor cells into an antitumor vaccine. Within the body, the scaffold, formed by a calcium-containing chitosan hydrogel complex (ChitoCa) entraps tumor cells and attracts immune cells to establish a durable and multicellular microenvironment. Within this context, tumor cells are completely eliminated by antigen-presenting cells (APCs) and processed for cross-antigen presentation. The regulatory mechanism relies on the Mincle receptor, a cell-phagocytosis-inducing C-type lectin receptor specifically activated on ChitoCa-recruited APCs, which serves as a recognition synapse, facilitating a tenfold increase in tumor cell engulfment and subsequent elimination. The ChitoCa-induced tumor cell processing further promotes the cross-presentation of tumor antigens to prime protective CD8+ T cell responses. Therefore, the ChitoCa treatment establishes an immune niche within the tumor microenvironment, resulting in effective tumor regression either used alone or in combination with other immunotherapies. This hydrogel-induced immune factory establishes a functional organ-like multicellular colony for tumor-specific immunotherapy, paving the way for innovative strategies in cancer treatment.

Two-pronged anti-cancer nanovaccines enpowered by exogenous/endogenous tumor-associated antigens.

Cancer vaccines based on single-source (exogenous or endogenous) tumor-associated antigens (TAAs) are often challenged by the insufficient T cell response and the immunosuppressive tumor microenvironment (TME). Herein, a dual TAAs-boosted nanovaccine based on cancer cell (4T1) membrane-cloaked, CO-immobilized Prussian blue nanoparticles (4T1-PB-CO NPs) is developed and coupled with anti-interleukin (IL)-10 therapy to maximize the efficacy of antitumor immunotherapy. 4T1 cell membrane not only endows NPs with tumor targeting ability, but also serves as exogenous TAAs to trigger CD4+ T cell response and M1-phenotype polarization of tumor-associated macrophages. Under near-infrared light irradiation, 4T1-PB-CO NPs release CO to induce immunogenic cell death (ICD) of tumor cells, thus generating endogenous TAAs to activate CD8+ T cell response. Meanwhile, ICD triggers release of damage-associated molecular patterns, which can promote DC maturation to amplify the antitumor T cell response. When combined with anti-IL-10 that reverses the immunosuppressive TME, 4T1-PB-CO NPs efficiently suppress the primary tumors and produce an abscopal effect to inhibit distant tumors in a breast tumor-bearing mouse model. Such a two-pronged cancer vaccine represents a promising paradigm for robust antitumor immunotherapy.

Ω-Shaped fiber optic LSPR coated with hybridized nanolayers for tumor cell sensing and photothermal treatment.

Circulating tumor cell (CTC) in the blood is the main cause of cancer metastasis for death in cancer patients. It is extremely important for cancer diagnosis at an early stage and treatment to simultaneously detect and kill the CTCs. In this work, a new hybridized nanolayer, namely gold nanoparticle/gold nanorods@ Polydopamine (AuNPs/AuNRs@PDA), was coated on the Ω-shaped fiber optics (Ω-FO) for localized surface plasmonic resonance (LSPR) to perform tumor cell sensing and photothermal treatment (PTT). The PDA nanolayer was formed on a bare fiber optic through the self-polymerization of dopamine under mild conditions. The AuNRs and AuNPs were absorbed on the surface of the PDA nanolayer to form a hybridized nanolayer. The hybridized nanolayer-modified Ω-FO LSPR exhibited a high refractive index sensitivity (RIS) of 37.59 (a.u/RIU) and photothermal conversion efficiency. After being modified with the recognition element of aptamer, the Ω-FO LSPR was used to develop a sensitive and specifical tumor cell sensing. Under the irradiation of near-infrared light (NIR) laser, the Ω-FO LSPR can kill the captured tumor cells with the apoptotic/necrotic rate of 62.6 % and low side-effect for the nontarget cells. The FO LSPR sensor realized the dual functions of CTC sensing and PTT, which provided a new idea for the early diagnosis and treatment of cancer.

Spontaneous-stimulated Raman co-localization dual-modal analysis approach for efficient identification of tumor cells.

The study of highly heterogeneous tumor cells, especially acute myeloid leukemia (AML) cells, usually relies on invasive analytical methods such as morphology, immunology, cytogenetics, and molecular biology classification, which are complex and time-consuming to perform. Mortality is high if patients are not diagnosed in a timely manner, so rapid label-free analysis of gene expression and metabolites within single-cell substructures is extremely important for clinical diagnosis and treatment. As a label-free and non-destructive vibrational detection technique, spontaneous Raman scattering provides molecular information across the full spectrum of the cell but lacks rapid imaging localization capabilities. In contrast, stimulated Raman scattering (SRS) provides a high-speed, high-resolution imaging view that can offer real-time subcellular localization assistance for spontaneous Raman spectroscopic detection. In this paper, we combined multi-color SRS microscopy with spontaneous Raman to develop a co-localized Raman imaging and spectral detection system (CRIS) for high-speed chemical imaging and quantitative spectral analysis of subcellular structures. Combined with multivariate statistical analysis methods, CRIS efficiently differentiated AML from normal leukocytes with an accuracy of 98.1 % and revealed the differences in the composition of nuclei and cytoplasm of AML relative to normal leukocytes. Compared to conventional Raman spectroscopy blind sampling without imaging localization, CRIS increased the efficiency of single-cell detection by at least three times. In addition, using the same approach for further identification of AML subtypes M2 and M3, we demonstrated that intracytoplasmic differential expression of proteins is a marker for their rapid and accurate classifying. CRIS analysis methods are expected to pave the way for clinical translation of rapid tumor cell identification.

Therapeutic Apheresis Using a ß2-Microglobulin Removal Column Reduces Circulating Tumor Cell Count.

An elevated serum ß2-microglobulin (ß2M) level is indicative of impaired glomerular filtration and prerenal diseases, such as malignant tumors, autoimmune disorders, and liver diseases. An elevated serum ß2M level has been shown to promote metastasis via the induction of epithelial-mesenchymal transition (EMT) in cancer cells. However, the therapeutic potential of targeting ß2M remains unclear. Here, we aimed to investigate the efficacy of Filtor, a small polymethyl methacrylate fiber-based ß2M removal column, in reducing the ß2M level and suppressing cancer cell-induced EMT and metastasis. We assessed the effects of Filtor on the changes in metastasis based on the number of circulating tumor cells (CTCs), which reflects the post-EMT cancer cell population. We performed therapeutic apheresis using Filtor on a male patient with sinonasal neuroendocrine carcinoma, a female patient with a history of colorectal cancer, and another female patient with a history of pancreatic ductal adenocarcinoma. Significantly low serum ß2M levels and CTC counts were observed immediately and 4 weeks after treatment compared with those in the pretreatment phase. Moreover, the CTC count immediately after therapeutic intervention was markedly reduced, likely because Filtor had trapped CTCs directly. These findings suggest that therapeutic apheresis with Filtor can prevent cancer metastasis and recurrence by directly removing CTCs.

Poptosis or Peptide-Induced Transmembrane Pore Formation: A Novel Way to Kill Cancer Cells without Affecting Normal Cells.

Recent advances in cancer treatment like personalized chemotherapy and immunotherapy are aimed at tumors that meet certain specifications. In this review, we describe a new approach to general cancer treatment, termed peptide-induced poptosis, in which specific peptides, e.g., PNC-27 and its shorter analogue, PNC-28, that contain the segment of the p53 transactivating 12-26 domain that bind to HDM-2 in its 1-109 domain, bind to HDM-2 in the membranes of cancer cells, resulting in transmembrane pore formation and the rapid extrusion of cancer cell contents, i.e., tumor cell necrosis. These peptides cause tumor cell necrosis of a wide variety of solid tissue and hematopoietic tumors but have no effect on the viability and growth of normal cells since they express at most low levels of membrane-bound HDM-2. They have been found to successfully treat a highly metastatic pancreatic tumor as well as stem-cell-enriched human acute myelogenous leukemias in nude mice, with no evidence of off-target effects. These peptides also are cytotoxic to chemotherapy-resistant cancers and to primary tumors. We performed high-resolution scanning immuno-electron microscopy and visualized the pores in cancer cells induced by PNC-27. This peptide forms 1:1 complexes with HDM-2 in a temperature-independent step, followed by dimerization of these complexes to form transmembrane channels in a highly temperature-dependent step parallel to the mode of action of other membranolytic but less specific agents like streptolysin. These peptides therefore may be effective as general anti-cancer agents.

The Design for a Phase II, Randomized, Multicenter Study of CtDNA-Guided Treatment With Furmonertinib Combined Therapy or Furmonertinib Alone for Untreated Advanced EGFR Mutant Non-small-cell Lung Cancer Patients: The FOCUS-C Study.

BACKGROUND: Third-generation epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) have significant antitumor activity to advanced non-small-cell lung cancer (NSCLC) patients with classic EGFR mutations. However, EGFR-TKI monotherapy shows poor efficacy in patients whose circulating tumor cell DNA (ctDNA) of EGFR mutations cannot be rapidly cleared. MATERIALS AND METHODS: As a third-generation TKI, furmonertinib has shown superior antitumor activity and minor toxicity. The FOCUS-C study is a prospective, multicenter, randomized controlled trial (NCT05334277) to explore the efficacy and safety of furmonertinib plus pemetrexed-platinum doublet chemotherapy with or without bevacizumab versus furmonertinib monotherapy in untreated advanced EGFR mutant NSCLC patients without EGFR clearance after the induction therapy of furmonertinib. Patients with EGFR clearance will still receive furmonertinib as Arm A. Patients without ctDNA clearance will be randomized in a 2:2:1 ratio as Arm B1 (furmonertinib), Arm B2 (furmonertinib combined with carboplatin and pemetrexed for 4 cycles, and then furmonertinib and pemetrexed as maintenance therapy) and Arm B3 (Arm B2 regimen plus bevacizumab). The primary endpoint is progression-free survival (PFS) in Arm B2/B1. Secondary endpoints include PFS in Arm B3/B1, PFS in Arm A/B1, PFS in Arm B3/B2, objective response and disease control rate, overall survival and safety in all Arms. Exploratory endpoints are focused on the efficacy based on plasma NGS at different timepoints. CONCLUSION: This study will evaluate the efficacy and tolerability of furmonertinib plus carboplatin and pemetrexed with or without bevacizumab verses furmonertinib alone in untreated patients with advanced EGFR mutant NSCLC without EGFR clearance.

A ratiometric SERS strategy for the prediction of cancer cell proportion and guidance of glioma surgical resection.

Rapid and accurate identification of tumor boundaries is critical for the cure of glioma, but it is difficult due to the invasive nature of glioma cells. This paper aimed to explore a rapid diagnostic strategy based on a label-free surface-enhanced Raman scattering (SERS) technique for the quantitative detection of glioma cell proportion intraoperatively. With silver nanoparticles as substrate, an in-depth SERS analysis was performed on simulated clinical samples containing normal brain tissue and different concentrations of patient-derived glioma cells. The results revealed two universal characteristic peaks of 655 and 717 cm-1, which strongly correlated with glioma cell proportion regardless of individual differences. Based on the intensity ratio of the two peaks, a ratiometric SERS strategy for the quantification of glioma cells was established by employing an artificial neuron network model and a polynomial regression model. Such a strategy accurately estimated the proportion of glioma cells in simulated clinical samples (R2 = 0.98) and frozen samples (R2 = 0.85). More importantly, it accurately facilitated the delineation of tumor margins in freshly obtained samples. Taken together, this SERS-based method ensured a rapid and more detailed identification of tumor margins during surgical resection, which could be beneficial for intraoperative decision-making and pathological evaluation.

Adaptive resistance of tumor cells to anti-vascular endothelial growth factor therapy: A reversible phenomenon.

Vascular endothelial growth factor (VEGF) inhibition is an essential targeted strategy for malignant tumors, but its efficacy is severely constrained by drug resistance. The traditional view holds that the target of VEGF inhibition is endothelial cells, and thus compensatory angiogenesis is considered the main mechanism of drug resistance. In this study, we found that tumor cells themselves could develop acquired resistance to VEGF therapy, indicating an independent resistance mechanism apart from angiogenesis. Notably, this acquired resistance was temporary, disappearing completely four days after discontinuing exposure to the drug in vitro. Our findings suggest that tumor cells may also be targets of VEGF inhibition, and their response to treatment should not be overlooked in contributing to drug resistance.

Tethering of hexokinase 2 to mitochondria promotes resistance of liver cancer cells to natural killer cell cytotoxicity.

Hexokinases (HKs) control the first step of glucose catabolism. A switch of expression from liver HK (glucokinase, GCK) to the tumor isoenzyme HK2 is observed in hepatocellular carcinoma progression. Our prior work revealed that HK isoenzyme switch in hepatocytes not only regulates hepatic metabolic functions but also modulates innate immunity and sensitivity to Natural Killer (NK) cell cytotoxicity. This study investigates the impact of HK2 expression and its mitochondrial binding on the resistance of human liver cancer cells to NK-cell-induced cytolysis. We have shown that HK2 expression induces resistance to NK cell cytotoxicity in a process requiring mitochondrial binding of HK2. Neither HK2 nor GCK expression affects target cells' ability to activate NK cells. In contrast, mitochondrial binding of HK2 reduces effector caspase 3/7 activity both at baseline and upon NK-cell activation. Furthermore, HK2 tethering to mitochondria enhances their resistance to cytochrome c release triggered by tBID. These findings indicate that HK2 mitochondrial binding in liver cancer cells is an intrinsic resistance factor to cytolysis and an escape mechanism from immune surveillance.