Evaluation of nanoparticle albumin-bound paclitaxel loaded macrophages for glioblastoma treatment based on a microfluidic chip.

Introduction: Glioblastoma (GBM) is a primary brain malignancy with a dismal prognosis and remains incurable at present. In this study, macrophages (MΦ) were developed to carry nanoparticle albumin-bound paclitaxel (nab-PTX) to form nab-PTX/MΦ. The aim of this study is to use a GBM-on-a-chip to evaluate the anti-GBM effects of nab-PTX/MΦ. Methods: In this study, we constructed nab-PTX/MΦ by incubating live MΦ with nab-PTX. We developed a microfluidic chip to co-culture GBM cells and human umbilical vein endothelial cells, mimicking the simplified blood-brain barrier and GBM. Using a syringe pump, we perform sustainable perfusion of nutrient media. To evaluate the anti-GBM effects nab-PTX/MΦ, we treated the GBM-on-a-chip model with nab-PTX/MΦ and investigated GBM cell proliferation, migration, and spheroid formation. Results: At the chosen concentration, nab-PTX did not significantly affect the viability, chemotaxis and migration of MΦ. The uptake of nab-PTX by MΦ occurred within 1 h of incubation and almost reached saturation at 6 h. Additionally, nab-PTX/MΦ exhibited the M1 phenotype, which inhibits tumor progression. Following phagocytosis, MΦ were able to release nab-PTX, and the release of nab-PTX by MΦ had nearly reached its limit at 48 h. Compared with control group and blank MΦ group, individual nab-PTX group and nab-PTX/MΦ group could inhibit tumor proliferation, invasion and spheroid formation. Meanwhile, the anti-GBM effect of nab-PTX/MΦ was more significant than nab-PTX. Discussion: Our findings demonstrate that nab-PTX/MΦ has a significant anti-GBM effect compared to individual nab-PTX or MΦ administration, suggesting MΦ as potential drug delivery vectors for GBM therapy. Furthermore, the developed GBM-on-a-chip model provides a potential ex vivo platform for innovative cell-based therapies and tailored therapeutic strategies for GBM.

Robust positive association between serum urate and the risk of chronic obstructive pulmonary disease: hospital-based cohort and Mendelian randomisation study.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) and hyperuricaemia are both characterised by systemic inflammation. Preventing chronic diseases among the population with common metabolic abnormality is an effective strategy. However, the association of hyperuricaemia with the higher incidence and risk of COPD remains controversial. Therefore, replicated researches in populations with distinct characteristics or demographics are compellingly warranted. METHODS: This cohort study adopted a design of ambispective hospital-based cohort. We used propensity score matching (PSM) and inverse probability of treatment weighting (IPTW) to minimise the effects of potential confounding factors. A Cox regression model and restricted cubic spline (RCS) model were applied further to assess the effect of serum urate on the risk of developing COPD. Finally, we conducted a two-sample Mendelian randomisation (MR) analysis to explore evidence of causal association. RESULTS: There is a higher incidence in the population with hyperuricaemia compared with the population with normal serum urate (22.29/1000 person-years vs 8.89/1000 person-years, p=0.009). This result is robust after performing PSM (p=0.013) and IPTW (p<0.001). The Cox model confirms that hyperuricaemia is associated with higher risk of developing COPD (adjusted HR=3.35 and 95% CI=1.61 to 6.96). Moreover, RCS shows that the risk of developing COPD rapidly increases with the concentration of serum urate when it is higher than the reference (420 µmol/L). Finally, in MR analysis, the inverse variance weighted method evidences that a significant causal effect of serum urate on COPD (OR=1.153, 95% CI=1.034 to 1.289) is likely to be true. The finding of MR is robust in the repeated analysis using different methods and sensitivity analysis. CONCLUSIONS: Our study provides convincing evidence suggesting a robust positive association between serum urate and the risk of developing COPD, and indicates that the population with hyperuricaemia is at high risk of COPD in the Chinese population who seek medical advice or treatment in the hospital.

Generation of Spatiotemporal Vortex Pulses by Resonant Diffractive Grating.

Spatiotemporal vortex pulses are wave packets that carry transverse orbital angular momentum, exhibiting exotic structured wave fronts that can twist through space and time. Existing methods to generate these pulses require complex setups like spatial light modulators or computer-optimized structures. Here, we demonstrate a new approach to generate spatiotemporal vortex pulses using just a simple diffractive grating. The key is constructing a phase vortex in frequency-momentum space by leveraging symmetry, resonance, and diffraction. Our approach is applicable to any wave system. We use a liquid surface wave (gravity wave) platform to directly demonstrate and observe the real-time generation and evolution of spatiotemporal vortex pulses. This straightforward technique provides opportunities to explore pulse dynamics and potential applications across different disciplines.

En Bloc Resection for Spinal Cord Hemangioblastomas: Surgical Technique and Clinical Outcomes.

BACKGROUND: Spinal cord hemangioblastomas are rare benign and highly vascular tumors that develop either sporadically or as part of von Hippel-Lindau (VHL) disease. Generally, complete resection without significant neurologic deficit remains considerably challenging due to the risk of massive bleeding. The current study therefore aimed to describe en bloc resection of spinal cord hemangioblastomas according to the typical anatomical structures of peripheral lesions and evaluate the neurofunctional prognosis of this technique. METHODS: A total of 39 spinal cord hemangioblastomas from a series of 19 patients who underwent en bloc resection were retrospectively analyzed. In all cases, clinical and radiologic characteristics, as well as surgical tenets, were retrospectively determined and analyzed. Short- and long-term outcomes were analyzed using the McCormick grade and Odom's criteria. Factors significantly associated with poor neurologic function after en bloc resection were also determined. RESULTS: All 39 spinal cord hemangioblastomas, including 28 intramedullary, 2 intramedullary-extramedullary, and 9 extramedullary lesions, were located dorsally or dorsolaterally (100.0%). The most common lesion location was the thoracic segment (53.8%), with most of the lesions being accompanied by syringomyelia (94.7%). Long-term follow-up (mean: 103 ± 50.4 months) for prognosis determination revealed that 88.2% (15/17) of all cases had stable or improved neurofunctional outcomes according to the McCormick grade and Odom's criteria. Only one case with VHL disease developed recurrence 4 years after surgery. Additionally, statistical analysis showed that VHL disease was an independent prognostic factor associated with deteriorating neurologic function (p = 0.015). CONCLUSIONS: En bloc resection facilitated satisfactory long-term functional outcomes in patients with spinal cord hemangioblastomas. Given that VHL disease was identified as a predictor of poor long-term outcomes, regular long-term follow-up of patients with VHL-associated spinal cord hemangioblastoma seems necessary.

Research on the interaction of astragaloside IV and calycosin in Astragalus membranaceus with HMGB1.

BACKGROUND: High mobility group box 1 protein (HMGB1), a lethal late inflammatory mediator, contributes to the pathogenesis of diverse inflammatory and infectious diseases. Astragaloside IV and calycosin as active ingredients in Astragalus membranaceus, possess potent regulatory ability on HMGB1-induced inflammation, however, the interaction between these two phytochemicals and HMGB1 has not been elucidated yet. METHODS: To further investigate the interaction of astragaloside IV, calycosin with HMGB1 protein, surface plasma resonance (SPR) and a series of spectroscopic methods, including UV spectra, fluorescence spectroscopy, circular dichroism (CD), were used. Molecular docking was also carried out to predict the atomic level's binding modes between two components and HMGB1. RESULTS: Astragaloside IV and calycosin were found to be able to bind HMGB1 directly and affect the secondary structure and environment of the chromogenic amino acids of HMGB1 to different extents. In silico, astragaloside IV and calycosin showed a synergistic effect by binding to the two independent domains B-box and A-box in HMGB1, respectively, where hydrogen and hydrophobicity bonds were regarded as the crucial forces. CONCLUSION: These findings showed that the interaction of astragaloside IV and calycosin with HMGB1 impaired its proinflammatory cytokines function, providing a new perspective for understanding the mechanism of A. membranaceus in treating aseptic and infectious diseases.

Nifuroxazide inhibits the growth of glioblastoma and promotes the infiltration of CD8 T cells to enhance antitumour immunity.

INTRODUCTION: Glioblastoma is a primary intracranial tumour with extremely high disability and fatality rates among adults. Existing diagnosis and treatment methods have not significantly improved the overall poor prognosis of patients. Nifuroxazide, an oral antibiotic, has been reported to act as a tumour suppressor in a variety of tumours and to participate in the process of antitumour immunity. However, whether it can inhibit the growth of glioma is still unclear. METHODS: We explored the potential mechanism of nifuroxazide inhibiting the growth of glioblastoma cells through in vitro and in vivo experiments. RESULTS: nifuroxazide can inhibit the proliferation of glioblastoma cells, promote G2 phase arrest, induce apoptosis, and inhibit epithelial-mesenchymal transition through the MAP3K1/JAK2/STAT3 pathway. Similarly, clinical sample analysis confirmed that MAP3K1 combined with STAT3 can affect the prognostic characteristics of patients with glioma. In addition, nifuroxazide can drive the M1 polarization of microglioma cells, inhibit the expression of CTLA4 and PD-L1 in tumour cells, and promote the infiltration of CD8 T cells to exert antitumour effects. Combination treatment with PD-L1 inhibitors can significantly prolong the survival time of mice. CONCLUSION: we found that nifuroxazide can inhibit the growth of glioblastoma and enhance antitumour immunity. Thus, nifuroxazide is an effective drug for the treatment of glioblastoma and has great potential for clinical application.

RRM2 Mediates the Anti-Tumor Effect of the Natural Product Pectolinarigenin on Glioblastoma Through Promoting CDK1 Protein Degradation by Increasing Autophagic Flux.

The natural product pectolinarigenin exerts anti-inflammatory activity and anti-tumor effects, and exhibits different biological functions, particularly in autophagy and cell cycle regulation. However, the antineoplastic effect of pectolinarigenin on glioblastoma (GBM) remains unclear. In the present study, we found that pectolinarigenin inhibits glioblastoma proliferation, increases autophagic flux, and induces cell cycle arrest by inhibiting ribonucleotide reductase subunit M2 (RRM2), which can be reversed by RRM2 overexpression plasmid. Additionally, pectolinarigenin promoted RRM2 protein degradation via autolysosome-dependent pathway by increasing autophagic flow. RRM2 knockdown promoted the degradation of CDK1 protein through autolysosome-dependent pathway by increasing autophagic flow, thereby inhibiting the proliferation of glioblastoma by inducing G2/M phase cell cycle arrest. Clinical data analysis revealed that RRM2 expression in glioma patients was inversely correlated with the overall survival. Collectively, pectolinarigenin promoted the degradation of CDK1 protein dependent on autolysosomal pathway through increasing autophagic flux by inhibiting RRM2, thereby inhibiting the proliferation of glioblastoma cells by inducing G2/M phase cell cycle arrest, and RRM2 may be a potential therapeutic target and a prognosis and predictive biomarker in GBM patients.

Genome Instability-Associated Long Non-Coding RNAs Reveal Biomarkers for Glioma Immunotherapy and Prognosis.

Genome instability is a hallmark of tumors and is involved in proliferation, invasion, migration, and treatment resistance of many tumors. However, the relationship of genome instability with gliomas remains unclear. Here, we constructed genome instability-derived long non-coding RNA (lncRNA)-based gene signatures (GILncSig) using genome instability-related lncRNAs derived from somatic mutations. Multiple platforms were used to confirm that the GILncSig were closely related to patient prognosis and clinical characteristics. We found that GILncSig, the glioma microenvironment, and glioma cell DNA methylation-based stemness index (mDNAsi) interacted with each other to form a complex regulatory network. In summary, this study confirmed that GILncSig was an independent prognostic indicator for patients, distinguished high-risk and low-risk groups, and affected immune-cell infiltration and tumor-cell stemness indicators (mDNAsi) in the tumor microenvironment, resulting in tumor heterogeneity and immunotherapy resistance. GILncSig are expected to provide new molecular targets for the clinical treatment of patients with gliomas.

A novel lncRNA, RPL34-AS1, promotes proliferation and angiogenesis in glioma by regulating VEGFA.

Purpose: Brain gliomas are the most common primary malignant tumors of the central nervous system and one of the leading causes of death in patients with intracranial tumors. The lncRNA RPL34-AS1 is significantly upregulated in glioma tissues. However, the biological function of RPL34-AS1, especially in proliferation in glioma, remains unclear. Methods: The role of RPL34-AS1 in proliferation and angiogenesis in glioma cells was investigated using the LN229, U87, and U251 glioma cell lines. The levels of RPL34-AS1 were detected using real-time quantitative reverse transcription polymerase chain reaction. CCK-8 and colony formation assays were performed to determine the role of RPL34-AS1 in proliferation and survival, and its role in angiogenesis was assessed by an endothelial tube formation assay. Changes in protein levels were assessed by western blotting. Results: RPL34-AS1 was upregulated in glioma tissues and was correlated with tumor grade. RPL34-AS1 expression was also higher in glioma cells than in normal astrocytes. Knockdown of RPL34-AS1 blocked glioma cell proliferation by inhibiting angiogenesis. This effect occurred through decreased ERK/AKT signaling. Conclusions: This study suggests that RPL34-AS1 affects cell proliferation and angiogenesis in glioma and therefore may potentially serve as a valuable diagnostic and prognostic biomarker and therapeutic target in patients with glioma.

The genomic characteristics of different progression patterns in advanced non-small cell lung cancer patients treated with immune checkpoint inhibitors.

BACKGROUND: Fast progression (FP), hyperprogressive disease (HPD), and early death (ED) are the newly reported cancer progression patterns in response to immune checkpoint inhibitor (ICI) treatment. This study aimed to investigate the clinical and genomic characteristics of FP, HPD, and ED following the ICI treatment of advanced non-small cell lung cancer (NSCLC). METHODS: We retrospectively reviewed 117 patients with advanced NSCLC who were treated with ICIs from March 2017 to October 2019. FP was defined as (I) time to treatment failure (TTF) <1.5 months; and (II) ≥50% increase in the sum of the longest diameter (SLD) of target lesions. HPD was defined as (I) TTF <2 months; and (II) ≥50% change in tumor growth rate compared with before ICI initiation. ED was defined as overall survival (OS) <3 months. Tissue samples from 18 FP/HPD/ED patients and 5 partial response (PR) patients were subjected to genomic profiling. Genomic data from 693 tumor mutational burden- and histology-matched lung cancer samples were retrieved from an internal database as a control. RESULTS: FP, HPD, and ED occurred in 7.21%, 9.38%, and 11.97% patients, respectively. The progression-free survival was comparable among the 3 groups. The median overall survival for FP, HPD, and ED were 3.19, 11.2, and 1.84 months, respectively. The genomic landscape revealed 1 EGFR amplification, 1 ALK fusion, 6 KRAS mutations, 1 ERBB2 amplification, 1 MET amplification, and 1 RET fusion among the 18 patients with FP/HPD/ED. Compared with the Control group, ED patients showed higher mutation frequencies for KRAS (P<0.01), CDKN1B (P<0.01), and NTRK1 (P=0.04). Mutations in RAD54L (P=0.018) and MYC (P=0.04) were more common in FP patients; HPD patients showed more frequent RAD54L mutations (P<0.001). CONCLUSIONS: We demonstrated different genomic characteristics across different progression patterns following ICI treatment, which might assist clinicians in the prediction of a patient's response, identifying candidates for more effective ICI therapy.

Liquid biopsy in extranodal NK/T-cell lymphoma: a prospective analysis of cell-free DNA genotyping and monitoring.

Satisfactory tumor material is often hard to obtain for molecular analysis in extranodal natural killer (NK)/T-cell lymphoma (NKTCL) at present. However, the accuracy and utility of circulating cell-free DNA (cfDNA) genotyping have not been adequately assessed in NKTCL. We therefore performed targeted next-generation sequencing on tumor tissues and a series of longitudinal plasma samples prospectively collected from a cohort of high-risk NKTCL patients. Concordance of genotyping results of paired baseline tumor and cfDNA and the predictive value of dynamic cfDNA monitoring were evaluated. At baseline, 59 somatic variants in 31 genes were identified in tumor and/or plasma cfDNA among 19 out of 24 high-risk NKTCL patients (79.2%). Plasma cfDNA had a sensitivity of 72.4% for detection of somatic variants identified in tumor biopsies before treatment. Plasma cfDNA also allowed the identification of mutations that were undetectable in tumor biopsies. These results were also verified in a validation cohort of an additional 23 high-risk NKTCL patients. Furthermore, longitudinal analysis showed that patients with rapid clearance of NKTCL-related mutations from plasma had higher complete remission rates (80.0% vs 0%; P = .004) and more favorable survival (1-year progression-free survival [PFS] rate, 79.0% vs 20.0%; P = .002) compared with those with persisting or emerging mutations in plasma. In addition, low cfDNA concentration before treatment was associated with favorable survival outcome for patients with NKTCL (1-year PFS, 90.0% vs 36.4%; P = .012). In conclusion, cfDNA mirrors tumor biopsy for detection of genetic alterations in NKTCL and noninvasive dynamic plasma cfDNA monitoring might be a promising approach for tracking response and survival outcome for patients with NKTCL.

Interfering with hyaluronic acid metabolism suppresses glioma cell proliferation by regulating autophagy.

The tumor microenvironment plays an important role in tumor progression. Hyaluronic acid (HA), an important component of the extracellular matrix in the tumor microenvironment, abnormally accumulates in a variety of tumors. However, the role of abnormal HA accumulation in glioma remains unclear. The present study indicated that HA, hyaluronic acid synthase 3 (HAS3), and a receptor of HA named CD44 were expressed at high levels in human glioma tissues and negatively correlated with the prognosis of patients with glioma. Silencing HAS3 expression or blocking CD44 inhibited glioma cell proliferation in vitro and in vivo. The underlying mechanism was attributed to the inhibition of autophagy flux and maintaining glioma cell cycle arrest in G1 phase. More importantly, 4-methylumbelliferone (4-MU), a small competitive inhibitor of Uridine diphosphate (UDP) with the ability to penetrate the blood-brain barrier (BBB), also inhibited glioma cell proliferation in vitro and in vivo. Thus, approaches that interfere with HA metabolism by altering the expression of HAS3 and CD44 and the administration of 4-MU potentially represent effective strategies for glioma treatment.

HER2 transmembrane domain mutation: comprehensive characteristics and real-world evidence of treatment response in Chinese lung adenocarcinoma.

BACKGROUND: HER2 transmembrane domain (TMD) mutation has been reported as a rare driver mutation associated with advanced stage disease and a poor prognosis in patients with lung adenocarcinoma (LUAD). We aimed to comprehensively profile the genetic landscape and treatment response information of HER2 TMD-mutant LUAD. METHODS: An in-house database of 7,812 LUAD patients was screened for mutation prevalence. A multi-center cohort of 16 HER2 V659E-mutant patients and an external cohort of 38 HER2-mutant patients from cBioPortal with overall survival (OS) data were analyzed. Eight patients from the in-house cohort were included in the real-world study of treatment response. Molecular docking simulation and binding affinity prediction were performed. RESULTS: In Chinese LUAD, the prevalence of HER2 TMD mutation was 0.18% (14/7,812), and 0.14% (11/7,812) for the HER2 V659E mutation. The most recurrent co-alteration was TP53 mutation (n=4, 25%) and HER2 amplification (n=2, 12.5%). TMD-mutant patients were diagnosed at more advance stages (P<0.001) and had poorer OS (median OS 10.0 vs. 61.6 months, HR =7.9, 95% CI: 1.0-61.0, P<0.001) than non-TMD mutations. The overall response rate of targeted therapy, chemo-based therapy, and immunotherapy was 57.1%, 22.2%, and 0%, respectively. We postulated to challenge the resistance of tyrosine kinase inhibitor (TKI) with another with stronger binding energy to HER2 and supported the conclusion with a successful case. Additionally, we demonstrated a three-month response to the off-label use of pyrotinib in fifth-line therapy. CONCLUSIONS: Comapred with non-TMD mtuations, HER2 TMD mutation is a rare driver mutation with poorer prognosis in LUAD. Targeted therapy is the dominant choice for patients harboring this targetable mutation and longer OS could possibly be achieved through rechallenge with TKI of stronger binding affinity. Response to fifth-line pyrotinib was observed.

HER2 copy number as predictor of disease-free survival in HER2-positive resectable gastric adenocarcinoma.

PURPOSE: The identification of HER2 overexpression in a subset of gastric adenocarcinoma (GA) patients represents a significant step forward in unveiling the molecular complexity of this disease. The predictive and prognostic value of HER2 amplification in advanced HER2 inhibitor-treated GA patients has been investigated. However, its predictive value in resectable patients remains elusive. METHODS: We enrolled 98 treatment-naïve resectable Chinese GA patients with HER2 overexpression assessed using IHC. Capture-based targeted sequencing using a panel consisting of 41 gastrointestinal cancer-related genes was performed on tumor tissues. Furthermore, we also investigated the correlation between HER2 copy number (CN) and survival outcomes. RESULTS: Of the 98 HER2-overexpressed patients, 90 had HER2 CN amplification assessed using next-generation sequencing, achieving 92% concordance. The most commonly seen concurrent mutations were occurring in TP53, EGFR and PIK3CA. We found HER2 CN as a continuous variable was an independent predictor associated with DFS (p = 0.029). Our study revealed HER2 CN-high patients showed a trend of intestinal-type GA predominant (p = 0.075) and older age (p = 0.07). The median HER2 CN was 15.34, which was used to divide the cohort into CN-high and CN-low groups. Patients with high HER2 CN had a significantly shorter DFS than patients with low HER2 CN (p = 0.002). Furthermore, HER2 CN as a categorical variable was also an independent predictor associated with DFS in patients. CONCLUSION: We elucidated the mutation spectrum of HER2-positive resectable Chinese GA patients and the association between HER2 CN and DFS. Our work revealed HER2 CN as an independent risk factor predicted unfavorable prognosis in HER2-positive GA patients and allowed us to further stratify HER2-positive resectable GA patients for disease management.

Refined Stratification Based on Baseline Concomitant Mutations and Longitudinal Circulating Tumor DNA Monitoring in Advanced EGFR-Mutant Lung Adenocarcinoma Under Gefitinib Treatment.

INTRODUCTION: The optimal treatment for EGFR-mutant lung adenocarcinoma (LUAD) remains challenging because of intratumor heterogeneity. We aimed to explore a refined stratification model based on the integrated analysis of circulating tumor DNA (ctDNA) tracking. METHODS: ctDNA was prospectively collected at baseline and at every 8 weeks in patients with advanced treatment-naive EGFR-mutant LUAD under gefitinib treatment enrolled in a phase 2 trial and analyzed using next-generation sequencing of a 168-gene panel. RESULTS: Three subgroups categorized by baseline comutations-EGFR-sensitizing mutations (59, 32.8%), EGFR-sensitizing mutations with tumor suppressor mutations (97, 53.9%), and EGFR-sensitizing mutations with other driver mutations (24, 13.3%)-exhibited distinct progression-free survival (13.2 [11.3-15.2] versus 9.3 [7.6-10.5] versus 4.0 [2.4-9.3] months) and overall survival (32.0 [29.2-41.5] versus 21.7 [19.3-27.0] versus 15.5 [10.5-33.7] months, respectively), providing evidence for initial stratification. A total of 63.7% of the patients achieved week 8 ctDNA clearance, with significant difference noted among the three subgroups (74.5% versus 64.0% versus 29.4%, respectively, p = 0.004, Fisher's exact test). Patients without week 8 ctDNA clearance had worse progression-free survival (clearance versus nonclearance 11.2 [9.9-13.2] versus 7.4 [5.6-9.6] months, p = 0.016, Cox regression], especially in the second subgroup [5.8 (5.6-11.5) months], suggesting the necessity of adaptive stratification during treatment. During follow-up, 56.0% and 20.8% of the patients eventually harbored p.T790M and non-p.T790M mutations, respectively, with a significant difference in non-p.T790M mutations among the three subgroups (7.5% versus 15.7% versus 80.0%, respectively, p < 0.001, Fisher's exact test), giving clues to postline treatment. CONCLUSIONS: The patients with baseline comutations and ctDNA nonclearance at first visit might require combined therapy because of the limited survival benefit of EGFR tyrosine kinase inhibitor monotherapy. We proposed a refined stratification mode for the whole-course management of EGFR-mutant LUAD.

Genomic characteristics in Chinese non-small cell lung cancer patients and its value in prediction of postoperative prognosis.

BACKGROUND: The genomic profile of non-small cell lung cancer (NSCLC) in Asians is distinct from that of Caucasians, but comprehensive genetic profiling reports have been limited for Asian patients. We aimed to elucidate genomic characteristics of Chinese NSCLC patients and develop potential model including genomic characteristics to predict postoperative prognosis. METHODS: Resected tumor samples from 511 patients with stage I-IV lung cancer were subjected to targeted sequencing using a panel of 295 cancer-related genes. Based on the molecular profiles and clinical features, we established nomogram models with predictors consisting of integrated clinical and genomic characteristics to provide post-operative risk stratification. RESULTS: Compared to the TCGA population (mainly Caucasians), there was a significantly higher frequency of EGFR (53.7% vs. 14.4%) and NOTCH3 (8.4% vs. 1.3%) mutations and less mutated KRAS (11.0% vs. 32.6%), KEAP1 (4.4% vs. 17.4%) and LRP1B (16.3% vs. 29.6%) in Chinese lung adenocarcinomas (LUAD). Distinct patterns of mutually exclusive and co-occurring mutations were identified between LUAD and lung squamous cell carcinoma (LUSC), indicating the unique histology-specific tumorigenesis mechanism of each subtype. We observed alterations in pathways correlated with clinical characteristics. Additionally, we constructed nomogram model with predictors consisting of clinical and genomic characteristics, which were more accurate than models with clinical characteristics or TNM staging only both in stage I-IIIA patients and T1-2N0M0 sub-cohort. CONCLUSIONS: This study revealed Chinese NSCLC patients have unique genomic profile. Furthermore, the nomogram model combining clinical features with genomic characteristics could improve risk stratification in early-stage NSCLC.

Tumor mutation burden in Chinese cancer patients and the underlying driving pathways of high tumor mutation burden across different cancer types.

BACKGROUND: Tumor mutation burden (TMB) has an important association with immunotherapy responses. TMB in the Chinese population has not been well established. Finding differences between the Chinese and Caucasian populations and elucidating the underlying biological mechanisms of high TMB might help develop more precise and effective means for TMB and immunotherapy response prediction. METHODS: Chinese cancer patients fresh tissue (n=2,177), formalin-fixed, paraffin-embed (FFPE) specimens (n=3,294), and pleural fluid (n=189) were profiled using a 295- or 520-gene next-generation sequencing (NGS) panel. The association of the TMB status with a series of molecular features and biological pathways was determined using bootstrapping. RESULTS: TMB, measured by 295- or 520-cancer-related gene panels, was correlated with whole-exome sequencing (WES) TMB based on the in silico simulation in The Cancer Genome Atlas cohort. The median TMB of our data was slightly higher than that from the Foundation Medicine Inc. (FMI) dataset. TMB was also slightly different within the same cancer type between the Chinese and Caucasian population. We discovered that the underlying pathways of TMB status varied greatly and sometimes had an opposite association with TMB across different cancer types. Moreover, we developed a 23-gene and a 16-gene signature to predict TMB prediction for lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC), respectively, indicating a histology-specific mechanism for driving high-TMB in lung cancer. CONCLUSIONS: TMB varies among different ethnic populations. Our findings extend the knowledge of the underlying biological mechanisms for high TMB and might be helpful for developing more precise and accessible TMB assessment panels and algorithms in more cancer types.

High-level gain of mesenchymal-epithelial transition factor (MET) copy number using next-generation sequencing as a predictive biomarker for MET inhibitor efficacy.

BACKGROUND: In clinical oncology, targeted next-generation sequencing (NGS) has become an integral part of the routine molecular diagnostics repertoire. However, a consensus is yet to be agreed on the optimal mesenchymal-epithelial transition factor (MET) copy number (CN) cut-off value based on NGS data that could predict the MET-amplified non-small cell lung cancer (NSCLC) patients who could benefit from MET tyrosine kinase inhibitor (TKI) therapy. In this study, we aimed to identify the criteria to define MET amplification derived from NGS data. METHODS: Sequencing data from matched plasma and tissue samples from 40 MET-amplified NSCLC patients were used to derive a normalization method, referred to as adjusted copy number (adCN). Clinical outcomes from an additional 18 MET TKI-treated NSCLC patients with solely MET-amplified cancers were analyzed to validate the adCN cut-offs. RESULTS: AdCN, calculated as the absolute CN generated from NGS relative to the maximum mutant allele fraction (maxMAF) per sample, was demonstrated to have a high correlation with MET CN in tissue and plasma samples (R2=0.73). Using a cut-off value of 5.5 and 13, tertile stratification of adCN was able to distinguish patients with high-level MET amplification. The MET TKI-treated patients with adCN >13, categorized as high-level amplification, had significantly longer progression-free survival (PFS) than those with adCN <13 (P=0.009), suggesting that adCN positively correlated with the response to MET TKI. CONCLUSIONS: We derived a normalization method that could reflect the relative CN and distinguish MET-amplified NSCLC patients with high-level gene amplification who were sensitive to crizotinib, suggesting adCN could potentially serve as a predictive biomarker for MET TKI response.

The prognostic value of a Methylome-based Malignancy Density Scoring System to predict recurrence risk in early-stage Lung Adenocarcinoma.

Current NCCN guidelines do not recommend the use of adjuvant chemotherapy for stage IA lung adenocarcinoma patients with R0 surgery. However, 25% to 40% of patients with stage IA disease experience recurrence. Stratifying patients according to the recurrence risk may tailor adjuvant therapy and surveillance imaging for those with a higher risk. However, prognostic markers are often identified by comparing high-risk and low-risk cases which might introduce bias due to the widespread interpatient heterogeneity. Here, we developed a scoring system quantifying the degree of field cancerization in adjacent normal tissues and revealed its association with disease-free survival (DFS). Methods: We recruited a cohort of 44 patients with resected stage IA lung adenocarcinoma who did not receive adjuvant therapy. Both tumor and adjacent normal tissues were obtained from each patient and subjected to capture-based targeted genomic and epigenomic profiling. A novel methylome-based scoring system namely malignancy density ratio (MD ratio) was developed based on 39 patients by comparing tumor and corresponding adjacent normal tissues of each patient. A MD score was then obtained by Wald statistics. The correlations of MD ratio, MD score, and genomic features with clinical outcome were investigated. Results: Patients with a high-risk MD ratio showed a significantly shorter postsurgical DFS compared with those with a low-risk MD ratio (HR=4.47, P=0.01). The MD ratio was not associated with T stage (P=1), tumor cell fraction (P=0.748) nor inflammatory status (p=0.548). Patients with a high-risk MD score also demonstrated an inferior DFS (HR=4.69, P=0.039). In addition, multivariate analysis revealed EGFR 19 del (HR=5.39, P=0.012) and MD score (HR= 7.90, P=0.01) were independent prognostic markers. Conclusion: The novel methylome-based scoring system, developed by comparing the signatures between tumor and corresponding adjacent normal tissues of individual patients, largely minimizes the bias of interpatient heterogeneity and reveals a robust prognostic value in patients with resected lung adenocarcinoma.