METTL3-dependent m6A modification of PSEN1 mRNA regulates craniofacial development through the Wnt/ß-catenin signaling pathway.

Craniofacial malformations, often associated with syndromes, are prevalent birth defects. Emerging evidence underscores the importance of m6A modifications in various bioprocesses such as stem cell differentiation, tissue development, and tumorigenesis. Here, in vivo, experiments with zebrafish models revealed that mettl3-knockdown embryos at 144 h postfertilization exhibited aberrant craniofacial features, including altered mouth opening, jaw dimensions, ethmoid plate, tooth formation and hypoactive behavior. Similarly, low METTL3 expression inhibited the proliferation and migration of BMSCs, HEPM cells, and DPSCs. Loss of METTL3 led to reduced mRNA m6A methylation and PSEN1 expression, impacting craniofacial phenotypes. Co-injection of mettl3 or psen1 mRNA rescued the level of Sox10 fusion protein, promoted voluntary movement, and mitigated abnormal craniofacial phenotypes induced by mettl3 knockdown in zebrafish. Mechanistically, YTHDF1 enhanced the mRNA stability of m6A-modified PSEN1, while decreased METTL3-mediated m6A methylation hindered ß-catenin binding to PSEN1, suppressing Wnt/ß-catenin signaling. Pharmacological activation of the Wnt/ß-catenin pathway partially alleviated the phenotypes of mettl3 morphant and reversed the decreases in cell proliferation and migration induced by METTL3 silencing. This study elucidates the pivotal role of METTL3 in craniofacial development via the METTL3/YTHDF1/PSEN1/ß-catenin signaling axis.

Directed evolution of a neutrophilic and mesophilic methanol dehydrogenase based on high-throughput and accurate measurement of formaldehyde.

Methanol is a promising one-carbon feedstock for biomanufacturing, which can be sustainably produced from carbon dioxide and natural gas. However, the efficiency of methanol bioconversion is limited by the poor catalytic properties of nicotinamide adenine dinucleotide (NAD+)-dependent methanol dehydrogenase (Mdh) that oxidizes methanol to formaldehyde. Herein, the neutrophilic and mesophilic NAD+-dependent Mdh from Bacillus stearothermophilus DSM 2334 (MdhBs) was subjected to directed evolution for enhancing the catalytic activity. The combination of formaldehyde biosensor and Nash assay allowed high-throughput and accurate measurement of formaldehyde and facilitated efficient selection of desired variants. MdhBs variants with up to 6.5-fold higher Kcat/KM value for methanol were screened from random mutation libraries. The T153 residue that is spatially proximal to the substrate binding pocket has significant influence on enzyme activity. The beneficial T153P mutation changes the interaction network of this residue and breaks the α-helix important for substrate binding into two short α-helices. Reconstructing the interaction network of T153 with surrounding residues may represent a promising strategy to further improve MdhBs, and this study provides an efficient strategy for directed evolution of Mdh.

A TP63 mutation identified in a Han Chinese family with ectodermal dysplasia.

OBJECTIVE: The purpose of this study was to identify a pathogenic mutation located in TP63 in a nuclear Han Chinese family. DESIGN: Whole-exome sequencing and Sanger sequencing were performed to identify candidate variants. The AlphaFold and PyMOL predicted the three-dimensional structure of the protein. Single-cell RNA-sequencing data and spatiotemporal transcriptomic atlas were used to generate the dissection of candidate gene expression at single-cell resolution. Significant genes (Pearson's coefficient ≥0.8 and P < 0.05) were identified for Gene Ontology (GO) analysis and Kyoto encyclopedia of genes and genomes (KEGG) pathways analysis. RESULTS: A heterozygous missense variant at TP63 exon 8 (c.1010 G>A:p.Arg337Gln) was identified in the proband. This variant was predicted deleterious and likely to impair the local stability of the protein. In addition, single-cell RNA-sequencing indicated that TP63 was highly expressed in skin tissues. Furthermore, spatial transcriptome data of mice embryos showed TP63 was mainly enriched in the mucosal epithelium, thymus, epidermis, mesenchyme, and surface ectoderm. GO and KEGG pathway annotation analysis revealed that TP63 played a positive role in the process of ectoderm via the TGF-beta signaling pathway. CONCLUSIONS: The missense variant of TP63 (c.1010 G>A:p.Arg337Gln) was associated with ectodermal dysplasia.

Effectiveness and safety of first-line immune checkpoint inhibitors for patients with extensive-stage small cell lung carcinoma: A systematic review and network meta-analysis.

Objective: In recent years, the introduction of immune checkpoint inhibitors (ICIs) has revolutionized the treatment of extensive-stage small cell lung carcinoma (ES-SCLC), but the optimal combination of ICI and standard chemotherapy strategy is yet to be established. The aim of this network meta-analysis (NMA) was to identify which first-line combination strategy is optimal for patients with ES-SCLC. Methods: PubMed, Embase, Cochrane Library, and the proceedings of international conferences, including American Society of Clinical Oncology and European Society for Medical Oncology meetings, were searched for randomized controlled trials (RCTs) published through October 31, 2022. The collected primary outcomes were overall survival (OS), progression-free survival (PFS), and grade 3-5 treatment-related adverse events (TRAEs). Results: Our NMA study included six phase 3 and three phase 2 RCTs including 4037 patients and 10 first-line regimens. Regarding effectiveness, the addition of programmed cell death 1 (PD-1) or programmed cell death ligand 1 (PD-L1) inhibitors to standard chemotherapy provided greater efficacy than chemotherapy alone. However, cytotoxic T lymphocyte-associated antigen-4 inhibitors were not associated with satisfactory prognoses. Serplulimab plus carboplatin-etoposide (vs. standard chemotherapy, hazard ratio [HR] = 0.63; 95% CI = 0.49-0.82) and nivolumab plus platinum-etoposide (HR = 0.65; 95% confidence interval [CI] = 0.46-0.91) displayed the greatest benefit regarding OS. In terms of PFS, serplulimab plus carboplatin-etoposide yielded the best benefit of all treatments (HR = 0.48; 95% CI = 0.39-0.6). The combination of ICIs and chemotherapy caused more toxicity in general, but durvalumab plus platinum-etoposide (odds ratio [OR] = 0.98; 95% CI = 0.68-1.4), atezolizumab plus carboplatin-etoposide (OR = 1.04; 95% CI = 0.68-1.6), and adebrelimab plus platinum-etoposide (OR = 1.02; 95% CI = 0.52-2) displayed similar safety as standard chemotherapy. Subgroup analysis by race illustrated that serplulimab plus carboplatin-etoposide was associated with the best OS in Asian patients. And in non-Asian patients, the combination of PD-1/PD-L1 inhibitors and chemotherapy (pembrolizumab plus platinum-etoposide, durvalumab plus platinum-etoposide, and durvalumab and tremelimumab plus platinum-etoposide) displayed superiority to standard chemotherapy. Conclusions: The results of our NMA study suggested that serplulimab plus carboplatin-etoposide and nivolumab plus platinum-etoposide are associated with the best OS as first-line treatments for patients with ES-SCLC. Serplulimab plus carboplatin-etoposide was associated with the best PFS. In Asian patients, serplulimab plus carboplatin-etoposide had the best OS. Systematic review registration: This study is registered with PROSPERO, number CRD42022345850.

Novel diagnostic biomarkers of T cell-mediated tumor killing characteristics for early-stage triple negative breast cancer: A SEER analysis and molecular portraits.

The primary objective was to investigate the epidemiology, molecular characteristics, and clinical survival to identify potential transcriptome biomarkers to promote early diagnosis and screening of triple-negative breast cancer patients. Early-stage triple-negative breast cancer patients (E-TNBC) and late-stage triple-negative breast cancer patients (L-TNBC) were identified from the Surveillance, Epidemiology, and End Results database from 2010 to 2019. The difference in cancer specific survival (CSS) and overall survival (OS) between E-TNBC and L-TNBC was analyzed via a Kaplan-Meier plotter. 118 triple-negative breast cancer samples and 114 normal samples with the RNA sequencing expression data were selected from the cohort of TCGA breast cancer from UCSC Xena Database. The study involved 13,690 patients with L-TNBC and 44,994 patients with E-TNBC. L-TNBC patients were more frequently to be ≤ 60 years old (54.9% vs 52.2%), multiple primary site (44.0% vs 30.1%), and were more likely to receive radiotherapy (49.6% vs 47.4%) and chemotherapy (81.1% vs 72.1%), while L-TNBC patients were less likely to be white (68.7% vs 73.0%), married or with domestic partner (46.7% vs 54.7%), poorly differentiated grade (54.0% vs 61.9%), < 3 months from diagnosis to treatment (91.6% vs 96.4%), and were less likely to receive surgery (72.3% vs 95.4%). Stage-stratified survival analysis revealed that the prognosis of L-TNBC was worse when compared to E-TNBC, Kaplan-Meier analysis demonstrated that there were striking differences in OS and CSS between E-TNBC and L-TNBC. In the multivariable regression models, L-TNBC was the single highest risk factor, with a death risk that was 4.741 and 6.074 times higher than E-TNBC in terms of OS and CSS, respectively. The results also showed that treatment with surgery, radiation, or chemotherapy was effective for a better prognosis. Transcriptome analyses revealed that the top 5 upregulated genes in L-TNBC were, respectively, ISX, ALOX15B, MADCAM1, TP63, and ARG1 compared with E-TNBC. And the top 5 downregulated genes were, respectively, CTAG1B, CT45A1, MAGEC2, TFF2, and TNFRSF11B. The L-TNBC exhibited a lower rate of survival than E-TNBC, and the 2 groups differed in terms of transcriptome characteristics. To date, the diagnostic value of T cell-mediated tumor killing portraits on E-TNBC may not be completely recognized.

Genetic architecture of non-syndromic skeletal class III malocclusion.

Non-syndromic skeletal Class III malocclusion is a major craniofacial disorder characterized by genetic and environmental factors. Patients with severe skeletal Class III malocclusion require orthognathic surgery to obtain aesthetic facial appearance and functional occlusion. Recent studies have demonstrated that susceptible chromosomal regions and genetic variants of candidate genes play important roles in the etiology of skeletal Class III malocclusion. Here, we provide a comprehensive review of our current understanding of the genetic factors that affect non-syndromic skeletal Class III malocclusion, including the patterns of inheritance and multiple genetic approaches. We then summarize the functional studies on related loci and genes using cell biology and animal models, which will help to implement individualized therapeutic interventions.

Nanobiotherapeutic strategies to target immune microenvironment of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is the subtype with the least favourable outcomes in breast cancer. Besides chemotherapy, there is a chronic lack of other effective treatments. Advances in omic technologies have liberated us from the ambiguity of TNBC heterogeneity in terms of cancer cell and immune microenvironment in recent years. This new understanding of TNBC pathology has already led to the exploitation of novel nanoparticulate systems, including tumor vaccines, oncolytic viruses, and antibody derivatives. The revolutionary ideas in the therapeutic landscape provide new opportunities for TNBC patients. Translating these experimental medicines into clinical benefit is both appreciated and challenging. In this review, we describe the prospective nanobiotherapy of TNBC that has been developed to overcome clinical obstacles, and provide our vision for this booming field at the overlap of cancer biotherapy and nanomaterial design.

IL2RA is a prognostic indicator and correlated with immune characteristics of pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive and incurable cancer with a dismal prognosis. In this study, we aimed to explore potential predictors for the prognosis and immunological characteristics of PDAC. Estimation of stromal and immune cells in malignant tumors, using expression data (ESTIMATE) method was applied to calculate the immune and stromal scores of 206 PDAC samples from GSE71729. R package of "limma" was utilized to identify differentially expressed genes (DEGs). Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analyses were conducted for functional exploration. Protein-protein interaction (PPI) network and Univariate Cox analysis were conducted to select key prognostic genes of PDAC. Gene set enrichment analysis (GSEA) was applied to investigate the roles of IL2RA in PDAC. Single sample GSEA (ssGSEA) was performed to evaluate the immunological characteristics of PDAC samples. Wilcoxon rank sum test was conducted to compare the difference of immunological characteristics of PDAC samples between low IL2RA and high IL2RA. Spearman correlation analysis was used to explore the correlations of IL2RA expression and immune checkpoint genes. A total of 747 DEGs were identified between low and high immune/stromal groups. Functional exploration revealed upregulated DEGs were associated with immune-related activities, whereas downregulated DEGs were involved in inflammatory-related activities. IL2RA was selected as the critical gene by overlapping the hub genes in PPI network and prognostic genes. Significantly, IL2RA expression was significantly elevated in PDAC and patients with higher IL2RA expression had worse prognoses. The immunological and oncogenic roles of IL2RA in PDAC were evidenced by GSEA. Furthermore, PDAC samples with high IL2RA expression exhibited increased immune infiltration and better immunotherapy responses. IL2RA expression was positively correlated with PDCD1, CD274, CTLA4, IDO1, TDO2, and TIGT. Higher expression of IL2RA predicts worse survival outcomes and increased immune infiltration in PDAC. PDAC patients with high IL2RA expression might potentially benefit from immunotherapy.

The improved success rate and reduced complications of a novel localization device vs. hookwire for thoracoscopic resection of small pulmonary nodules: a single-center, open-label, randomized clinical trial.

Background: In our previous study, we developed a 4-hook claw-suture localization device for pulmonary nodule resection, which acheived satifisfactory results. Following this, we conducted this single-center, open-label, randomized clinical trial to compare the success rate and complication rate of this novel localization device and currently widely-used hookwire. Methods: Patients with small pulmonary nodules (0.4-1 cm) who received preoperative localization and thoracoscopic resection at Shanghai Chest Hospital were randomly assigned (1:2 ratio, via computer-generated randomized numbers) to undergo localization using either a novel claw-suture system (claw group) or classical (hookwire group) localization device. The primary endpoint of this study was localization success rate, and the secondary endpoints included complications, localization-related time, and pain. Results: A total of 411 patients were randomly assigned to the claw group (n=136) or the hookwire group (n=275) before thoracoscopic resection of small pulmonary nodules and analyzed. Compared with the hookwire group, the claw group had a significantly higher success rate (133/136, 97.8% vs. 254/275, 92.4%, P=0.027), less asymptomatic hemorrhage (16.9% vs. 37.5%, P=0.003) and pleural reaction (0% vs. 5.1%, P=0.017), as well as better pain alleviation 10 min after localization (measured using the difference between two visual analog scale scores, 0.84±0.98 vs. 0.35±0.79, P<0.001). In contrast, the hookwire group was associated with a shorter localization procedure duration than the claw group (7.2±2.9 vs. 14.4±6.6 min, P<0.001). In the multiple localization subgroup, the claw group compared to the hookwire group also achieved higher success (32/33, 97.0% vs. 70/86, 81.4%) and less pleural reaction (0% vs. 16.3%). Conclusions: The new claw-suture localization device is superior to traditional hookwire, with a higher success rate, fewer complications, and better patient tolerance for preoperative localization of small pulmonary nodules. Trial Registration: Chinese Clinical Trial Registry ChiCTR1900027346.

Multi-scale characterization of tumor-draining lymph nodes in resectable lung cancer treated with neoadjuvant immune checkpoint inhibitors.

BACKGROUND: Regional lymph node (LN) acts as a pivotal organ for antitumor immunity. Paradoxically, tumor-draining LNs (TDLNs) are usually the first site of tumor metastasis in lung cancer. It is largely unknown about the association between the status of TDLNs and the response of primary tumor beds to immune checkpoint inhibitors (ICIs) in lung cancer patients. Also, studies characterizing the TDLNs in response to ICIs are scarce. METHODS: We characterized and compared the radiological, metabolic (18F-FDG) and pathologic responses between primary tumor beds and paired TDLNs (invaded/non-invaded) from 68 lung cancer patients who underwent neoadjuvant ICIs plus surgery. Additionally, we performed the spatial profiling of immune and non-immune cells within TDLNs using multiplexed immunofluorescence. Therapy responses (e.g., pathologic complete (pCR) or major response (MPR)) of primary lung tumor beds and paired TDLNs were investigated separately. FINDINGS: We observed that responses of TDLNs to ICIs markedly differ from their paired primary lung tumors regarding the radiological, metabolic (18F-FDG uptake), and pathologic alterations. Neoadjuvant ICIs therapy specifically decreased 18F-FDG-reflected metabolic activity in the primary tumor beds with pCR/MPR but not their TDLNs counterparts. Furthermore, the presence of invaded TDLNs was associated with poor pathologic responses in the matched primary tumor beds and predictive of rapid post-treatment tumor relapse. Spatial profiling demonstrated exclusion of T cell infiltrates within the metastatic lesions of invaded TDLNs, and diminished multiple immune and non-immune compositions in non-involved regions surrounding the metastatic lesions. INTERPRETATION: These results provide the first clinically-relevant evidence demonstrating unique response patterns of TDLNs under ICIs treatment and revealing the underappreciated association of TDLNs status with the response of their paired primary tumors to ICIs in lung cancer. FUNDING: This work was supported by the National Natural Science Foundation of China (82072570 to F. Yao; 82002941 to B. Sun), the excellent talent program of Shanghai Chest Hospital (to F.Y), the Basic Foundation Program for Youth of Shanghai Chest Hospital (2021YNJCQ2 to H.Yang), and the Innovative Research Team of High-level Local Universities in Shanghai (SHSMU-ZLCX20212302 to F. Yao).

Cyclophilin A contributes to shikonin-induced glioma cell necroptosis and promotion of chromatinolysis.

Shikonin induces glioma cell death via necroptosis, a caspase-independent programmed cell death pathway that is chiefly regulated by receptor-interacting serine/threonine protein kinase1 (RIP1) and 3 (RIP3). Chromatinolysis is considered as one of the key events leading to cell death during necroptosis. It is usually accompanied with nuclear translocation of AIF and formation of γ-H2AX. Cyclophilin A (CypA) is reported to participate in the nuclear translocation of AIF during apoptosis. However, it remains unclear whether CypA contributes to necroptosis and regulation of chromatinolysis. In this study, our results revealed for the first time that shikonin promoted time-dependent CypA activation, which contributed to nuclear translocation of AIF and γ-H2AX formation. In vitro studies showed that knockdown of CypA by siRNA or inhibition of CypA by its specific inhibitor, cyclosporine A (CsA), not only significantly mitigated shikonin-induced glioma cell death, but also prevented chromatinolysis. Mechanistically, activated CypA targeted mitochondria and triggered mitochondrial superoxide overproduction, which then promoted AIF translocation from mitochondria into the nucleus by depolarizing the mitochondria and intensified the formation of γ-H2AX by promoting intracellular accumulation of ROS. Additionally, the CypA in the nucleus can form DNA degradation complexes with AIF and γ-H2AX, which also promote the execution of chromatinolysis. Thus, we demonstrate that CypA contributes to shikonin-induced glioma cell necroptosis and promotion of chromatinolysis.

Analysis of the prognostic role and biological characteristics of circulating tumor cell-associated white blood cell clusters in non-small cell lung cancer.

Background: Recently, circulating tumor-cell-associated white blood cell (CTC-WBC) clusters have been reported to have prognostic value in some cancers. The prognostic role of CTC-WBC clusters in lung cancer has not yet been elucidated. Very little information is available about the biological characteristics of CTC-WBC clusters. Methods: A total of 82 patients with non-small cell lung cancer (NSCLC) were included in this study, and 61 patients with advanced-stage disease were closely followed-up. All patients had blood drawn prior to treatment. Subtraction enrichment and immunostaining-fluorescence in situ hybridization (SE-iFISH) platform was used to isolate and identify CTCs and CTC-WBC clusters. Kaplan-Meier survival analysis and Cox regression analysis were applied to assess patient progression-free survival (PFS). Further, qualitative and quantitative analyses the size and ploidy characteristics of CTC-WBC clusters. Results: Firstly, CTC-WBC clusters appeared more in the advanced (stage III and IV) stage (P=0.043) than in the early stage. Furthermore, the multivariable analysis (Cox proportional hazards model) revealed that the high-CTC (≥7/6 mL) group and CTC-WBC clusters (≥1/6 mL) positive group both had significantly worse PFS, with a hazard ratio (HR) of 2.89 [95% confidence interval (CI): 1.36-6.17, P=0.006] and 2.18 (95% CI: 1.07-4.43, P=0.031), respectively. In the conjoint analysis, compared to patients with <7 CTCs/6 mL without CTC-WBC clusters, patients with ≥7 CTCs/6 mL with CTC-WBC clusters had the highest risk of progression (HR =7.13, 95% CI: 2.51-20.23, P<0.001). In addition, the presence of ≥3-cell CTC-WBC clusters in patients may indicate a shorter PFS (P<0.05) and a higher risk of progression (HR =2.90, 95% CI: 1.06-7.89, P=0.037). Furthermore, compared with the characteristics of the total CTCs, almost all of the CTCs that could recruit WBCs were large cells (≥5 µm) and exhibited polyploidy (≥ tetraploid) (both P<0.01). Conclusions: The presence of CTC-WBC clusters was an independent prognostic factor for advanced NSCLC. The joint analysis of CTCs and CTC-WBC clusters could provide additional prognostic value to the enumeration of CTCs alone. Besides, most of the CTCs in CTC-WBC clusters were large polyploid cells.

Multi-scale integrative analyses identify THBS2+ cancer-associated fibroblasts as a key orchestrator promoting aggressiveness in early-stage lung adenocarcinoma.

Rationale: Subsets of patients with early-stage lung adenocarcinoma (LUAD) have a poor post-surgical course after curative surgery. However, biomarkers stratifying this high-risk subset and molecular underpinnings underlying the aggressive phenotype remain unclear. Methods: We integrated bulk and single-cell transcriptomics, proteomics, secretome and spatial profiling of clinical early-stage LUAD samples to identify molecular underpinnings that promote the aggressive phenotype. Results: We identified and validated THBS2, at multi-omic levels, as a tumor size-independent biomarker that robustly predicted post-surgical survival in multiple independent clinical cohorts of early-stage LUAD. Furthermore, scRNA-seq data revealed that THBS2 is exclusively derived from a specific cancer-associated fibroblast (CAF) subset that is distinct from CAFs defined by classical markers. Interestingly, our data demonstrated that THBS2 was preferentially secreted via exosomes in early-stage LUAD tumors with high aggressiveness, and its levels in the peripheral plasma associated with short recurrence-free survival. Further characterization showed that THBS2-high early-stage LUAD was characterized by suppressed antitumor immunity. Specifically, beyond tumor cells, THBS2+ CAFs mainly interact with B and CD8+ T lymphocytes as well as macrophages within tumor microenvironment of early-stage LUAD, and THBS2-high LUAD was associated with decreased immune cell infiltrates but increased immune exhaustion marker. Clinically, high THBS2 expression predicted poor response to immunotherapies and short post-treatment survival of patients. Finally, THBS2 recombinant protein suppressed ex vivo T cells proliferation and promoted in vivo LUAD tumor growth and distant micro-metastasis. Conclusions: Our multi-level analyses uncovered tumor-specific THBS2+ CAFs as a key orchestrator promoting aggressiveness in early-stage LUAD.

Smoking signature is superior to programmed death-ligand 1 expression in predicting pathological response to neoadjuvant immunotherapy in lung cancer patients.

BACKGROUND: There is a paucity of biomarkers that can predict the degree of pathological response [e.g., pathological complete response (pCR) or major response (pMR)] to immunotherapy. Neoadjuvant immunotherapy provides an ideal setting for exploring responsive biomarkers because the pathological responses can be directly and accurately evaluated. METHODS: We retrospectively collected the clinicopathological characteristics and treatment outcomes of non-small cell lung cancer (NSCLC) patients who received neoadjuvant immunotherapy or chemo-immunotherapy followed by surgery between 2018 and 2020 at a large academic thoracic cancer center. Clinicopathological factors associated with pathological response were analyzed. RESULTS: A total of 39 patients (35 males and 4 females) were included. The most common histological subtype was lung squamous cell carcinoma (LUSC) (n=28, 71.8%), followed by lung adenocarcinoma (LUAD) (n=11, 28.2%). After neoadjuvant treatment, computed tomography (CT) scan-based evaluation showed poor agreement with the postoperatively pathological examination (weighted kappa =0.0225; P=0.795), suggesting the poor performance of CT scans in evaluating the response to immunotherapy. Importantly, we found that the smoking signature displayed a better performance than programmed death-ligand 1 (PD-L1) expression in predicting the pathological response (area under the curve: 0.690 vs. 0.456; P=0.0259), which might have resulted from increased tumor mutational burden (TMB) and/or microsatellite instability (MSI) relating to smoking exposure. CONCLUSIONS: These findings suggest that CT scan-based evaluation is not able to accurately reflect the pathological response to immunotherapy and that smoking signature is a superior marker to PD-L1 expression in predicting the benefit of immunotherapy in NSCLC patients.

Surgical Resection of Primary Tumors Provides Survival Benefits for Lung Cancer Patients With Unexpected Pleural Dissemination.

Background: Surgery is not generally recommended for non-small cell lung cancer (NSCLC) patients with malignant pleural dissemination (PD). However, in some cases, PD is found unexpectedly during surgery. There is no consensus on whether surgical intervention can provide survival benefit for them. We investigated the role of surgery in NSCLC patients with unexpected PD by a cohort study. Methods: Clinical data of consecutive patients who intended to undergo radical surgery for NSCLC between January 2010 and December 2015 at Shanghai Chest Hospital and Huadong Hospital were collected from a lung cancer database. Patients diagnosed with unexpected malignant pleural nodules intraoperatively were enrolled in this retrospective study. Results: A total of 181 NSCLC patients were diagnosed with unexpected malignant PD intraoperatively and confirmed with postoperatively histological examinations. Out of these, 80 (44.2%) patients received pleural nodule biopsies alone, and 101 (55.8%) received primary tumor resection (47 with sublobar resection and 54 with lobectomy). The median progression-free survival and overall survival for all patients were 13 and 41 months respectively. Patients in the resection group had significantly better progression-free survival (19.0 vs. 10.0 months, P < 0.0001) and overall survival (48.0 vs. 33.0 months, P < 0.0001) than patients in the biopsy group. In the resection group, there was no statistical difference between patients with sublobar resection and lobectomy (P = 0.34). Univariate and multivariate analyses identified primary tumor resection, targeted adjuvant therapy, and tumor size (≤ 3 cm) as independent prognostic factors. Conclusions: NSCLC patients with unexpected intraoperative PD potentially benefited from surgical resection of the primary tumor and multidisciplinary targeted therapy, particularly when tumor size did not exceed 3 cm. Our data demonstrated that the resection type was not associated with survival differences, which remains to be defined with a larger sample size.

pN1 but not pN0/N2 predicts survival benefits of prophylactic cranial irradiation in small-cell lung cancer patients after surgery.

BACKGROUND: Prophylactic cranial irradiation has been shown to reduce brain metastases and provide survival benefits in small-cell lung cancer (SCLC). However, its role in limited-stage SCLC patients after surgery remains unclear. Further, it is unknown whether the effect of prophylactic cranial irradiation is generalizable in these patients with different pathological nodal (N0-N2) stages, a state indicating the presence of tumor metastases. METHODS: We combined data from a single medical center and Surveillance, Epidemiology, and End Results database. Propensity score matching analyses were performed (1:2) to evaluate the role of prophylactic cranial irradiation in SCLC patients after surgery. Cox proportional hazards regression model was used to identify predictors of survival. RESULTS: 124 (18.7%) out of 664 surgically-treated SCLC patients received prophylactic cranial irradiation treatment. Within the entire cohort, multivariate Cox regression analysis identified dataset source, age, pathological T and N stages, adjuvant chemotherapy, resection type, and histology as independent prognostic factors for overall survival. Prophylactic cranial irradiation appeared to be associated with a better overall survival, but the difference is marginally significant (P=0.063). Further, we stratified patients based on the pathological N0-N2 stages using propensity score matching analyses, which showed that prophylactic cranial irradiation treatment was superior to non-prophylactic cranial irradiation treatment for surgically-treated SCLC patients with N1 stage only (univariate analysis: P=0.026; multivariate Cox: P=0.004), but not N0/N2 stage (univariate analysis: P=0.65 and P=0.28, respectively; multivariate Cox: P=0.99 and P=0.35, respectively). CONCLUSIONS: Prophylactic cranial irradiation provides survival benefits for SCLC patients with pN1 after surgery but not with pathological N0/N2 stage. Our findings may provide helpful stratifications for clinical decision-making of prophylactic cranial irradiation intervention in SCLC patients.

Cisplatin nanoparticles boost abscopal effect of radiation plus anti-PD1 therapy.

The abscopal effect of radiation therapy (RT) is clinically significant but occurs rarely. Although anti-programmed cell death protein 1 antibody (anti-PD1) is likely to enhance the abscopal effect in patients receiving RT, the incidence rate remains less than 30%. One major limitation is the paucity of CD8+ T cells within non-irradiated tumors. Here, cisplatin (CDDP) loaded poly(l-glutamic acid)-graft-methoxy poly(ethylene glycol) complex nanoparticles (CDDP-NPs) are confirmed to increase CD8+ T cells within non-irradiated tumors and boost the abscopal effect of RT plus anti-PD1, and more strongly than CDDP. Compared to RT and RT + CDDP, RT + CDDP-NPs induced greater immunogenic cell death (ICD) with enhanced proportion of Calreticulin+ Lewis lung cancer (LLC) cells (16.47%, 20.53% and 27.03%), along with which more CD8+ T cells were infiltrated into CDDP-NP treated irradiated tumors in the unilateral LLC tumor model. In the bilateral LLC tumor model, RT + CDDP-NPs significantly induced more chemokine (C-X-C motif) ligand 10 (CXCL10) secretion (36.3, 44.19 and 56.37 pg mL-1), which corresponded to greater CD8+ T cell infiltration in the non-irradiated tumors (0.19%, 0.20% and 0.72%). Finally, compared to RT + anti-PD1 and RT + anti-PD1 + CDDP, RT + anti-PD1 + CDDP-NPs significantly inhibited the growth of non-irradiated tumors more forcefully, as indicated by the respective tumor volumes of 1141, 1146 and 585 mm3. This is the first study to show that CDDP-NPs can amplify RT-induced immune activation and break through the efficiency limitation of the RT plus anti-PD1 induced abscopal effect.

Pulmonary benign metastasizing leiomyoma simultaneously diagnosed with uterine leiomyoma at first visit before hysteromyomectomy.

Pulmonary benign metastasizing leiomyoma (PBML) represents a rare disease which is mostly diagnosed in reproductive-age women with a hysteromyomectomy history. This disease could occur in any age group, but more common in women of late fertility. And its clinical processes are closely linked to sex hormone levels. Here we describe a case of PBML simultaneously diagnosed with uterine leiomyoma at patient's first visit without a hysteromyomectomy history. The patient was a 37-year-old woman presenting with recurrent lower abdominal discomfort for more than half a year. No nausea, cough, dyspnea, hemoptysis, or abdominal distension was found. Additionally, the menstruation of the patient was regular. Image examinations confirmed a single solid pulmonary nodule in the right upper lobe, multiple fibroids of the uterus, and left adnexal nodules at her first visit. Laparoscopic hysteromyomectomy and pulmonary segmentectomy were sequentially performed with an interval of one month. Pathological findings were consistent with benign metastasizing leiomyomas. Estrogen and progesterone receptors were positive by immunohistostaining of the tumors. In conclusion, when confronted with female patients, who are at childbearing age with or without a history of uterine fibroids surgery, with single or multiple lesions in the lung that highly resemble metastatic tumors, but fail to find the primary malignant tumor clinically, the possibility of PBML must be considered. And sequential surgical treatment could be arranged.

Procyanidins mediates antineoplastic effects against non-small cell lung cancer via the JAK2/STAT3 pathway.

BACKGROUND: Lung cancer is a malignant tumor with one of the highest rates of cancer-related morbidity and mortality worldwide. Non-small cell lung cancer (NSCLC) account for 85% of all lung cancers and have a poor prognosis. Proanthocyanidins (PCs) are polyphenolic compounds that are found widely in natural plants. The present study aimed to determine the effects of PC on lung cancer and identify its possible mechanism. METHODS: A cell growth assay was used to detect the cell growth ability of A549 cancer cells, and a clonal formation assay was used to detect the cloning ability of A549 cancer cells. Flow cytometry was used to detect the effect of PCs on apoptosis and the cell cycle. The wound healing test, Transwell migration, and invasion test were used to detect the migration and invasion of human NSCLC A549 cells. Western blotting was utilized to detect the expression levels of N-cadherin, E-cadherin, vimentin, Janus kinase 2 (JAK2), p-signal transducer and activator of transcription 3 (p-STAT3), STAT3, matrix metalloproteinase 2 (MMP-2), MMP-9, and the apoptosis-related proteins, B-cell lymphoma-2 (Bcl-2) and BCL2-associated X (Bax). Cell immunofluorescence was used to detect the expression levels of the p-STAT3 primary antibody. RESULTS: PCs reduced the proliferation and cloning ability of A549 cells and significantly inhibited the migration and invasion of A549 cells in a dose-dependent manner. At the same time, PCs induced apoptosis in A549 cells and G2/M cell cycle arrest. PCs increased the pro-apoptotic protein expression, Bax, and down-regulated the anti-apoptotic protein expression, Bcl-2. PCs also inhibited the epithelial-mesothermal transition (EMT) process of A549 cells. We also found that the JAK2/STAT3 signaling pathway inhibitor, AG490, cooperated with PCs to inhibit A549 cell invasion and migration. Our results demonstrated that PCs could mediate the antitumor effect of NSCLC via the JAK2/STAT3 pathway. CONCLUSIONS: PCs can inhibit NSCLC A549 cell proliferation, invasion, metastasis, clone formation, EMT, and induced apoptosis and G2/M cell cycle arrest. They work by inhibiting the JAK2/STAT3 signaling pathway. As a novel antitumor drug, PCs have broad application prospects for the treatment of NSCLC.

Neoadjuvant immunotherapy facilitates resection of surgically-challenging lung squamous cell cancer.

BACKGROUND: Locally-advanced lung squamous cell carcinoma represents a special subset that is challenging to resect completely with surgery alone. Immunotherapy has achieved great success in treating late-stage lung cancer. However, whether neoadjuvant immunotherapy can facilitate resection of initially locally-advanced and surgically-difficult locally-advanced lung squamous cell carcinoma remains to be investigated. METHODS: We retrospectively collected clinical records of locally-advanced lung squamous cell carcinoma patients who received neoadjuvant immunotherapy followed by surgery between 2018 and 2020 at a large academic thoracic cancer center. RESULTS: A total of 23 patients (22 males, 1 female) with locally-advanced locally-advanced lung squamous cell carcinoma were included, initially clinically staged at IIIA (16, 69.6%), IIIB (n=4, 17.4%), IIB (n=2, 8.7%) and IIIC (n=1, 4.3%). The median interval between final treatment to surgery was 36 days (range, 25-93 days), without treatment-related delay in surgery. The neoadjuvant treatment resulted in a high rate of radical resection (n=20, 87.0%). The final histopathological examination demonstrated 6 (26.1%) cases with pathological complete response and 8 (34.8%) with pathological major response. Comparing with the computed tomography scan-based response, we observed a very low consistency (weighted kappa =0.122, P=0.315) between the computed tomography scan-based and final pathological evaluation. The median follow-up time was 510 days (range, 217-920 days). At the end of the follow-up, 1 patient died. CONCLUSIONS: Our findings showed the clinical promise of neoadjuvant immunotherapy plus surgery for locally-advanced lung squamous cell carcinoma. Computed tomography scan displays a poor role in assessing the resectability after neoadjuvant immunotherapy.