Direct oral anticoagulants versus low-molecular-weight heparin in patients with cancer-associated venous thrombosis: a cost-effectiveness analysis.

Background: Direct oral anticoagulants (DOACs) have demonstrated clinical benefits and better patient adherence over low-molecular-weight heparin (LMWH) in treating patients with cancer-associated venous thrombosis (CAT). We aimed to compare the cost-effectiveness of DOACs against LMWH in patients with CAT from the perspective of the Hong Kong healthcare system. Methods: A Markov state-transition model was performed to estimate the incremental cost-effectiveness ratio (ICER) per quality-adjusted life years (QALYs) for DOACs and LMWH in a hypothetical cohort of 10,000 patients with CAT over a 5-year lifetime horizon. The model was primarily based on the health states of no event, recurrent venous thromboembolism, bleeding, and death. Transition probabilities, relative risks, and utilities were derived from the literature. Resource cost data were obtained from the Hong Kong Hospital Authority. Deterministic and probabilistic sensitivity analyses tested the robustness of the results. Results: Relative to LMWH, DOACs were associated with increased QALYs (1.52 versus 1.50) at a lower medical cost of USD 2,232 versus 8,224 in five years. The cost of LMWH was the main contributor to the outcome. Out of 10,000 simulated cases, DOACs were dominant in 15.8% and cost-effective in 42.1%, at the willingness-to-pay threshold of USD 148,392 per additional QALY. Conclusions: DOACs were associated with greater QALY improvements and lower overall costs compared to LMWH. Accounting for uncertainty, DOACs were between cost-effective and dominant in 57.9% of cases. DOACs are a cost-effective alternative to LMWH in the management of CAT in Hong Kong.

Many-body enhancement of high-harmonic generation in monolayer MoS2.

Many-body effects play an important role in enhancing and modifying optical absorption and other excited-state properties of solids in the perturbative regime, but their role in high harmonic generation (HHG) and other nonlinear response beyond the perturbative regime is not well-understood. We develop here an ab initio many-body method to study nonperturbative HHG based on the real-time propagation of the non-equilibrium Green's function with the GW self energy. We calculate the HHG of monolayer MoS2 and obtain good agreement with experiment, including the reproduction of characteristic patterns of monotonic and nonmonotonic harmonic yield in the parallel and perpendicular responses, respectively. Here, we show that many-body effects are especially important to accurately reproduce the spectral features in the perpendicular response, which reflect a complex interplay of electron-hole interactions (or exciton effects) in tandem with the many-body renormalization and Berry curvature of the independent quasiparticle bandstructure.

Evaluating Virtual Contrast-enhanced MRI (VCE-MRI) in Nasopharyngeal Carcinoma Radiotherapy: A Retrospective Analysis for Primary Gross Tumor Delineation.

PURPOSE: To investigate the potential of virtual contrast-enhanced MRI (VCE-MRI) for gross-tumor-volume (GTV) delineation of nasopharyngeal carcinoma (NPC) using multi-institutional data. METHODS AND MATERIALS: This study retrospectively retrieved T1-weighted (T1w), T2-weighted (T2w) MRI, gadolinium-based contrast-enhanced MRI (CE-MRI) and planning CT of 348 biopsy-proven NPC patients from three oncology centers. A multimodality-guided synergistic neural network (MMgSN-Net) was trained using 288 patients to leverage complementary features in T1w and T2w MRI for VCE-MRI synthesis, which was independently evaluated using 60 patients. Three board-certified radiation oncologists and two medical physicists participated in clinical evaluations in three aspects: image quality assessment of the synthetic VCE-MRI, VCE-MRI in assisting target volume delineation, and effectiveness of VCE-MRI-based contours in treatment planning. The image quality assessment includes distinguishability between VCE-MRI and CE-MRI, clarity of tumor-to-normal tissue interface and veracity of contrast enhancement in tumor invasion risk areas. Primary tumor delineation and treatment planning were manually performed by radiation oncologists and medical physicists, respectively. RESULTS: The mean accuracy to distinguish VCE-MRI from CE-MRI was 31.67%; no significant difference was observed in the clarity of tumor-to-normal tissue interface between VCE-MRI and CE-MRI; for the veracity of contrast enhancement in tumor invasion risk areas, an accuracy of 85.8% was obtained. The image quality assessment results suggest that the image quality of VCE-MRI is highly similar to real CE-MRI. The mean dosimetric difference of planning target volumes were less than 1Gy. CONCLUSIONS: The VCE-MRI is highly promising to replace the use of gadolinium-based CE-MRI in tumor delineation of NPC patients.

Unsupervised Domain Adaptation for Low-Dose CT Reconstruction via Bayesian Uncertainty Alignment.

Low-dose computed tomography (LDCT) image reconstruction techniques can reduce patient radiation exposure while maintaining acceptable imaging quality. Deep learning (DL) is widely used in this problem, but the performance of testing data (also known as target domain) is often degraded in clinical scenarios due to the variations that were not encountered in training data (also known as source domain). Unsupervised domain adaptation (UDA) of LDCT reconstruction has been proposed to solve this problem through distribution alignment. However, existing UDA methods fail to explore the usage of uncertainty quantification, which is crucial for reliable intelligent medical systems in clinical scenarios with unexpected variations. Moreover, existing direct alignment for different patients would lead to content mismatch issues. To address these issues, we propose to leverage a probabilistic reconstruction framework to conduct a joint discrepancy minimization between source and target domains in both the latent and image spaces. In the latent space, we devise a Bayesian uncertainty alignment to reduce the epistemic gap between the two domains. This approach reduces the uncertainty level of target domain data, making it more likely to render well-reconstructed results on target domains. In the image space, we propose a sharpness-aware distribution alignment (SDA) to achieve a match of second-order information, which can ensure that the reconstructed images from the target domain have similar sharpness to normal-dose CT (NDCT) images from the source domain. Experimental results on two simulated datasets and one clinical low-dose imaging dataset show that our proposed method outperforms other methods in quantitative and visualized performance.

Effectiveness and safety of continuous low-molecular-weight heparin versus switching to direct oral anticoagulants in cancer-associated venous thrombosis.

Given the existing uncertainty regarding the effectiveness and safety of switching from low-molecular-weight heparin (LMWH) to direct oral anticoagulants (DOACs) in patients with cancer-associated venous thrombosis (CAT), we conducted a comprehensive population-based cohort study utilizing electronic health database in Hong Kong. A total of 4356 patients with CAT between 2010 and 2022 were included, with 1700 (39.0%) patients switching to DOAC treatment. Compared to continuous LMWH treatment, switching to DOACs was associated with a significantly lower risk of hospitalization due to venous thromboembolism (HR: 0.49 [95% CI = 0.35-0.68]) and all-cause mortality (HR: 0.67 [95% CI = 0.61-0.74]), with no significant difference in major bleeding (HR: 1.04 [95% CI = 0.83-1.31]) within six months. These findings provide reassurance regarding the effectiveness and safety of switching from LMWH to DOACs among patients with CAT, including vulnerable patient groups.

Stereotactic Body Radiotherapy and Liver Transplant for Liver Cancer: A Nonrandomized Controlled Trial.

Importance: Whether stereotactic body radiotherapy (SBRT) as a bridge to liver transplant for hepatocellular carcinoma (HCC) is effective and safe is still unknown. Objective: To investigate the feasibility of SBRT before deceased donor liver transplant (DDLT) for previously untreated unresectable HCC. Design, Setting, and Participants: In this phase 2 nonrandomized controlled trial conducted between June 1, 2015, and October 18, 2019, 32 eligible patients within UCSF (University of California, San Francisco) criteria underwent dual-tracer (18F-fluorodeoxyglucose and 11C-acetate [ACC]) positron emission tomography with computed tomography (PET-CT) and magnetic resonance imaging (MRI) with gadoxetate followed by SBRT of 35 to 50 Gy in 5 fractions, and the same imaging afterward while awaiting DDLT. Statistical analysis was performed on an intention-to-treat basis between October 1 and 31, 2023. Intervention: Patients received SBRT followed by DDLT when matched deceased donor grafts were available. Main Outcomes and Measures: Coprimary end points were progression-free survival (PFS) and objective response rates (ORRs) by the Response Evaluation Criteria in Solid Tumors, version 1.1 (RECIST 1.1), modified RECIST (mRECIST), and PET Response Criteria in Solid Tumors (PERCIST). Secondary end points were local control rate, overall survival (OS), and safety. Results: A total of 32 patients (median age, 59 years [IQR, 54-63 years]; 22 men [68.8%]) with 56 lesions received SBRT. After a median follow-up of 74.6 months (IQR, 40.1-102.9 months), the median PFS was 17.6 months (95% CI, 6.6-28.6 months), and the median OS was 60.5 months (95% CI, 29.7-91.2 months). The 5-year PFS was 39.9% (95% CI, 19.9%-59.9%), and the 5-year OS was 51.3% (95% CI, 31.7%-70.9%). In terms of number of patients, ORRs were 62.5% ([n = 20] 95% CI, 54.2%-68.7%) by RECIST 1.1, 71.9% ([n = 23] 95% CI, 63.7%-79.0%) by mRECIST, and 78.1% ([n = 25] 95% CI, 73.2%-86.7%) by PERCIST. In terms of number of lesions, ORRs were 75.0% ([n = 42] 95% CI, 61.6%-80.8%) by RECIST 1.1, 83.9% ([n = 47] 95% CI, 74.7%-90.6%) by mRECIST, and 87.5% ([n = 49] 95% CI, 81.3%-98.6%) by PERCIST. Twenty patients with 36 lesions received DDLT, of whom 15 patients (75.0%) with 21 lesions (58.3%) exhibited pathologic complete response. Multivariable analyses revealed that pretreatment metabolic tumor volume (MTV) based on ACC (hazard ratio [HR], 1.06 [95% CI, 1.01-1.10]; P = .01) and complete metabolic response (CMR) by PERCIST (HR, 0.31 [95% CI, 0.10-0.96]; P = .04) were associated with PFS, while pretreatment MTV based on ACC (HR, 1.07 [95% CI, 1.03-1.16]; P = .01), total lesion activity based on ACC (HR, 1.01 [95% CI, 1.00-1.02]; P = .02), and CMR by PERCIST (HR, 0.21 [95% CI, 0.07-0.73]; P = .01) were associated with OS. Toxic effects associated with SBRT were reported for 9 patients (28.1%), with 1 grade 3 event. Conclusions and Relevance: This phase 2 nonrandomized controlled trial demonstrated promising survival and safety outcomes of SBRT before DDLT for unresectable HCC. Future randomized clinical trials are warranted.

Identification and Characterization of Metastasis-Initiating Cells in ESCC in a Multi-Timepoint Pulmonary Metastasis Mouse Model.

Metastasis is the biggest obstacle to esophageal squamous cell carcinoma (ESCC) treatment. Single-cell RNA sequencing analyses are applied to investigate lung metastatic ESCC cells isolated from pulmonary metastasis mouse model at multiple timepoints to characterize early metastatic microenvironment. A small population of parental KYSE30 cell line (Cluster S) resembling metastasis-initiating cells (MICs) is identified because they survive and colonize at lung metastatic sites. Differential expression profile comparisons between Cluster S and other subpopulations identified a panel of 7 metastasis-initiating signature genes (MIS), including CD44 and TACSTD2, to represent MICs in ESCC. Functional studies demonstrated MICs (CD44high) exhibited significantly enhanced cell survival (resistances to oxidative stress and apoptosis), migration, invasion, stemness, and in vivo lung metastasis capabilities, while bioinformatics analyses revealed enhanced organ development, stress responses, and neuron development, potentially remodel early metastasis microenvironment. Meanwhile, early metastasizing cells demonstrate quasi-epithelial-mesenchymal phenotype to support both invasion and anchorage. Multiplex immunohistochemistry (mIHC) staining of 4 MISs (CD44, S100A14, RHOD, and TACSTD2) in ESCC clinical samples demonstrated differential MIS expression scores (dMISs) predict lymph node metastasis, overall survival, and risk of carcinothrombosis.

Using the genomic adjusted radiation dose (GARD) to personalize the radiation dose in nasopharyngeal cancer.

BACKGROUND: Locally advanced nasopharyngeal cancer (NPC) patients undergoing radiotherapy are at risk of treatment failure, particularly locoregional recurrence. To optimize the individual radiation dose, we hypothesize that the genomic adjusted radiation dose (GARD) can be used to correlate with locoregional control. METHODS: A total of 92 patients with American Joint Committee on Cancer / International Union Against Cancer stage III to stage IVB recruited in a randomized phase III trial were assessed (NPC-0501) (NCT00379262). Patients were treated with concurrent chemo-radiotherapy plus (neo) adjuvant chemotherapy. The primary endpoint is locoregional failure free rate (LRFFR). RESULTS: Despite the homogenous physical radiation dose prescribed (Median: 70 Gy, range 66-76 Gy), there was a wide range of GARD values (median: 50.7, range 31.1-67.8) in this cohort. In multivariable analysis, a GARD threshold (GARDT) of 45 was independently associated with LRFFR (p = 0.008). By evaluating the physical dose required to achieve the GARDT (RxRSI), three distinct clinical subgroups were identified: (1) radiosensitive tumors that RxRSI at dose < 66 Gy (N = 59, 64.1 %) (b) moderately radiosensitive tumors that RxRSI dose within the current standard of care range (66-74 Gy) (N = 20, 21.7 %), (c) radioresistant tumors that need a significant dose escalation above the current standard of care (>74 Gy) (N = 13, 14.1 %). CONCLUSION: GARD is independently associated with locoregional control in radiotherapy-treated NPC patients from a Phase 3 clinical trial. GARD may be a potential framework to personalize radiotherapy dose for NPC patients.

The Role of Natural Killer Cells in the Tumor Immune Microenvironment of EBV-Associated Nasopharyngeal Carcinoma.

Endemic nasopharyngeal carcinoma (NPC) is closely associated with the Epstein-Barr virus (EBV), which contributes to tumor development and influences the tumor immune microenvironment (TIME) in NPC. Natural killer (NK) cells, as part of the innate immune system, play a crucial role in responding to viral infections and malignant cell transformations. Notably, NK cells possess a unique ability to target tumor cells independent of major histocompatibility complex class I (MHC I) expression. This means that MHC I-deficient tumor cells, which can escape from effective T cell attack, are susceptible to NK-cell-mediated killing. The activation of NK cells is determined by the signals generated through inhibitory and activating receptors expressed on their surface. Understanding the role of NK cells in the complex TIME of EBV+ NPC is of utmost importance. In this review, we provide a comprehensive summary of the current understanding of NK cells in NPC, focusing on their subpopulations, interactions, and cytotoxicity within the TIME. Moreover, we discuss the potential translational therapeutic applications of NK cells in NPC. This review aims to enhance our knowledge of the role of NK cells in NPC and provide valuable insights for future investigations.

Learning Robust Shape Regularization for Generalizable Medical Image Segmentation.

Generalizable medical image segmentation enables models to generalize to unseen target domains under domain shift issues. Recent progress demonstrates that the shape of the segmentation objective, with its high consistency and robustness across domains, can serve as a reliable regularization to aid the model for better cross-domain performance, where existing methods typically seek a shared framework to render segmentation maps and shape prior concurrently. However, due to the inherent texture and style preference of modern deep neural networks, the edge or silhouette of the extracted shape will inevitably be undermined by those domain-specific texture and style interferences of medical images under domain shifts. To address this limitation, we devise a novel framework with a separation between the shape regularization and the segmentation map. Specifically, we first customize a novel whitening transform-based probabilistic shape regularization extractor namely WT-PSE to suppress undesirable domain-specific texture and style interferences, leading to more robust and high-quality shape representations. Second, we deliver a Wasserstein distance-guided knowledge distillation scheme to help the WT-PSE to achieve more flexible shape extraction during the inference phase. Finally, by incorporating domain knowledge of medical images, we propose a novel instance-domain whitening transform method to facilitate a more stable training process with improved performance. Experiments demonstrate the performance of our proposed method on both multi-domain and single-domain generalization.

Dosimetric parameters predict radiation-induced temporal lobe necrosis in nasopharyngeal carcinoma patients: A systematic review and meta-analysis.

This systematic review examines the role of dosimetric parameters in predicting temporal lobe necrosis (TLN) risk in nasopharyngeal carcinoma (NPC) patients treated with three-dimensional conformal RT (3D-CRT), intensity-modulated radiation therapy (IMRT) and volumetric modulated arc therapy (VMAT). TLN is a serious late complication that can adversely affect the quality of life of NPC patients. Understanding the relationship between dosimetric parameters and TLN can guide treatment planning and minimize radiation-related complications. A comprehensive search identified relevant studies published up to July 2023. Studies reporting on dosimetric parameters and TLN in NPC patients undergoing 3D-CRT, IMRT, and VMAT were included. TLN incidence, follow-up duration, and correlation with dosimetric parameters of the temporal lobe were analyzed. The review included 30 studies with median follow-up durations ranging from 28 to 110 months. The crude incidence of TLN varied from 2.3 % to 47.3 % and the average crude incidence of TLN is approximately 14 %. Dmax and D1cc emerged as potential predictors of TLN in 3D-CRT and IMRT-treated NPC patients. Threshold values of >72 Gy for Dmax and >62 Gy for D1cc were associated with increased TLN risk. However, other factors should also be considered, including host characteristics, tumor-specific features and therapeutic factors. In conclusion, this systematic review highlights the significance of dosimetric parameters, particularly Dmax and D1cc, in predicting TLN risk in NPC patients undergoing 3D-CRT, IMRT, and VMAT. The findings provide valuable insights that can help in developing optimal treatment planning strategies and contribute to the development of clinical guidelines in this field.

The trends in lung cancer prevalence, incidence, and survival in Hong Kong over the past two decades (2002-2021): a population-based study.

Background: Over the past decades, significant progress in lung cancer management has been made. However, the trends in prevalence and survival of lung cancer in the Chinese population over the last decade remain unexplored. This study utilised a territory-wide electronic medical database in Hong Kong to provide the most up-to-date and comprehensive analysis of the trends in prevalence, incidence, and survival over the past two decades. Methods: Descriptive epidemiology study using a retrospective cohort of lung cancer patients from the Clinical Data Analysis and Reporting System (CDARS). 10-year limited-duration prevalence, incidence, and relative period survival were calculated between 2002 and 2021. Sub-groups of age, sex, and comorbidity were examined. The annual percent change (APC) and average annual percent change (AAPC) were estimated using joinpoint regression. Findings: This study included 87,259 incident cases between 2002 and 2021. The 10-year limited duration prevalence (per 100,000 persons) of lung cancer increased from 153.4 to 228.7 (AAPC: 3.08%). Crude incidence (per 100,000 persons) increased from 55.0 to 70.3 (APC: 1.23%), while age-standardised incidence decreased from 42.9 to 33.2 (APC: -1.32%). The 1-year and 5-year relative period survivals showed an increasing trend but remained low. Disparity in trends was observed among different sex and age groups. Interpretation: Lung cancer burden has been increasing partly due to population ageing. Although survival showed improvement over the years, it remained low, highlighting the potential need for interventions. Further study exploring the disparity in sex-specific trends is warranted. Funding: The Innovation and Technology Commission, Hong Kong.

Shift in Tissue-Specific Immune Niches and CD137 Expression in Tuberculoma of Pembrolizumab-Treated Nasopharyngeal Carcinoma Patients.

The use of immune checkpoint inhibitors (ICIs) in cancer treatment has shown promise but can also have unintended consequences, such as reactivating latent tuberculosis (TB). To develop treatments that address ICIs-related adverse events, it is essential to understand cellular heterogeneity across healthy and pathological tissues. We performed cross-tissue multiplexed staining analysis on samples from two patients with TB reactivation during pembrolizumab treatment for metastatic nasopharyngeal carcinoma. CD8+ T cells, rather than CD4+ T cells, accumulated preferentially in the tuberculoma and were associated with increased production of IFNγ and expression of CD137. Additionally, CD137 enrichment played a role in the spatial organization of the tuberculoma, with specific interaction limited to spatial proximal cells between IFNγ+ CD137+ CD8+ T cells and IL12+ CD137+ type-1 macrophages. This unique feature was not observed in non-tumoral or tumoral tissues. Our analysis of public transcriptomic datasets supported the notion that this cellular interaction was more prominent in patients with durable ICI responses compared to those with non-ICI-related TB. We suggest that shifts towards CD137-rich immune niches are correlated with both off-target immune-related adverse events and anti-tumor efficacy. Targeting the tumor microenvironment through conditional activation of anti-CD137 signaling in combination with ICIs can modulate the reactivity of T cells and macrophages for localized tumor killing without the potential off-target immune-related risks associated with ICIs alone.

Enhanced mitophagy driven by ADAR1-GLI1 editing supports the self-renewal of cancer stem cells in HCC.

BACKGROUND AND AIMS: Deregulation of adenosine-to-inosine editing by adenosine deaminase acting on RNA 1 (ADAR1) leads to tumor-specific transcriptome diversity with prognostic values for HCC. However, ADAR1 editase-dependent mechanisms governing liver cancer stem cell (LCSC) generation and maintenance have remained elusive. APPROACH AND RESULTS: RNA-seq profiling identified ADAR1-responsive recoding editing events in HCC and showed editing frequency of GLI1 , rather than transcript abundance was clinically relevant. Functional differences in LCSC self-renewal and tumor aggressiveness between wild-type (GLI1 wt ) and edited GLI1 (GLI1 edit ) were elucidated. We showed that overediting of GLI1 induced an arginine-to-glycine (R701G) substitution, augmenting tumor-initiating potential and exhibiting a more aggressive phenotype. GLI1 R701G harbored weak affinity to SUFU, which in turn, promoted its cytoplasmic-to-nuclear translocation to support LCSC self-renewal by increased pluripotency gene expression. Moreover, editing predisposed to stabilize GLI1 by abrogating ß-TrCP-GLI1 interaction. Integrative analysis of single-cell transcriptome further revealed hyperactivated mitophagy in ADAR1-enriched LCSCs. GLI1 editing promoted a metabolic switch to oxidative phosphorylation to control stress and stem-like state through PINK1-Parkin-mediated mitophagy in HCC, thereby conferring exclusive metastatic and sorafenib-resistant capacities. CONCLUSIONS: Our findings demonstrate a novel role of ADAR1 as an active regulator for LCSCs properties through editing GLI1 in the highly heterogeneous HCC.

Model Generalizability Investigation for GFCE-MRI Synthesis in NPC Radiotherapy Using Multi-Institutional Patient-Based Data Normalization.

Recently, deep learning has been demonstrated to be feasible in eliminating the use of gadoliniumbased contrast agents (GBCAs) through synthesizing gadolinium-free contrast-enhanced MRI (GFCE-MRI) from contrast-free MRI sequences, providing the community with an alternative to get rid of GBCAs-associated safety issues in patients. Nevertheless, generalizability assessment of the GFCE-MRI model has been largely challenged by the high inter-institutional heterogeneity of MRI data, on top of the scarcity of multi-institutional data itself. Although various data normalization methods have been adopted to address the heterogeneity issue, it has been limited to single-institutional investigation and there is no standard normalization approach presently. In this study, we aimed at investigating generalizability of GFCE-MRI model using data from seven institutions by manipulating heterogeneity of MRI data under five popular normalization approaches. Three state-of-the-art neural networks were applied to map from T1-weighted and T2-weighted MRI to contrast-enhanced MRI (CE-MRI) for GFCE-MRI synthesis in patients with nasopharyngeal carcinoma. MRI data from three institutions were used separately to generate three uni-institution models and jointly for a tri-institution model. The five normalization methods were applied to normalize the data of each model. MRI data from the remaining four institutions served as external cohorts for model generalizability assessment. Quality of GFCE-MRI was quantitatively evaluated against ground-truth CE-MRI using mean absolute error (MAE) and peak signal-to-noise ratio(PSNR). Results showed that performance of all uni-institution models remarkably dropped on the external cohorts. By contrast, model trained using multi-institutional data with Z-Score normalization yielded the best model generalizability improvement.

ENOX2 inhibition enhances infiltration of effector memory T-cell and mediates response to chemotherapy in immune-quiescent nasopharyngeal carcinoma.

INTRODUCTION: The immunosuppressive tumor microenvironment is a major barrier for chemotherapy. Different chemosensitization approaches to reinstate immunological surveillance for cancers that are immune quiescent at the outset, have thus been devised. Cancer-specific ENOX2 expression is correlated with abnormal cell growth and has been proposed as a cellular target for anti-cancer activity. However, the potential effects of ENOX2 on the interaction between immune system and tumor cells remain elusive. OBJECTIVES: To understand the mechanisms by which tumor-intrinsic ENOX2-mediated alterations in anti-tumor activity of T-cells and response to chemotherapy. METHODS: In situ multiplexed immunohistochemistry with single cell and bulk RNA sequencing data from nasopharyngeal carcinoma (NPC) human tissues were used to define tumor phenotypes. Two NPC cell lines, with distinct ENOX2 expression, were used in a co-culture platform to study tumor-immune interactions between cancer cells/spheroids and T-cells. The effect of cisplatin treatment with ENOX2 inhibition by idronoxil (IDX) were tested in vitro and in vivo. Multi-parametric flow cytometry was used to characterize T-cell infiltrates in an NPC tumor humanized mouse model treated with combined treatment. RESULTS: NPC predominantly displayed an immune-excluded profile. This "cold-phenotype" was shown to exhibit higher ENOX2 expression and was associate with poorer progression-free survival (PFS). The therapeutic combination of IDX with cisplatin was effective in promoting CD8+ effector memory T cell (Tem) differentiation and mobilization. This Tem signature was highly cytotoxic, with Tem-mediated preferential lysis of higher ENOX2-expressing NPC cells. A combination-treated humanized mouse model showing dramatic shrinkage in tumors, were intra-tumoral Tem-enriched. CONCLUSION: Tumor-intrinsic ENOX2 expression is associated with tumor phenotype and PFS in NPC. Targeting ENOX2 with IDX and cisplatin impose qualitative control of T-cell response by preferentially increasing immune cells infiltration, Tem differentiation and tumor suppression. We suggest that ENOX2 inhibition may be a promising therapeutic strategy to enhance the effects of chemotherapy.

Single-Cell Secretion Analysis via Microfluidic Cell Membrane Immunosorbent Assay for Immune Profiling.

Single-cell multiplexed phenotypic analysis expands the biomarkers for diagnosis, heralding a new era of precision medicine. Cell secretions are the primary measures of immune function, but single-cell screening remains challenging. Here, a novel cell membrane-based assay was developed using cholesterol-linked antibodies (CLAbs), integrating immunosorbent assays and droplet microfluidics to develop a flexible high-throughput single-cell secretion assay for multiplexed phenotyping. CLAb-grafted single cells were encapsulated in water-in-oil droplets to capture their own secretions. Subsequently, the cells were extracted from droplets for fluorescence labeling and screening. Multiple secretions and surface proteins were simultaneously measured from single cells by flow cytometry. To validate the approach, THP-1 cells, THP-1-derived M1 macrophages, and dendritic cells were assayed, indicating the differentiation efficiency of THP-1 cells under different chemical stimulations. Moreover, peripheral blood mononuclear cells from healthy donors under various stimuli showed varied active immune cell populations (6.62-47.14%). The peripheral blood mononuclear cells (PBMCs) of nasopharyngeal carcinoma patients were analyzed to identify a higher percentage of actively cytokine-secreted single cells in the basal state (2.82 ± 1.48%), compared with that in the health donors (0.70 ± 0.29%).