Traditional Chinese herbal medicine complex supplementation improves reproductive performance, serum biochemical parameters, and anti-oxidative capacity in periparturient dairy cows.

This study was conducted to investigate the effects of a traditional Chinese herbal medicine complex (TCHMC) on the productive performance of periparturient dairy cows. Eighteen non-lactating pregnant Holstein dairy cows with similar body conditions with 1 to 2 parity were randomly divided into three groups (n = 6), receiving a basal diet with 0 (CON group), 200 (T-200 group), and 300 (T-300 group) g TCHMC per day from 14 to 9 days prepartum. The results demonstrated that TCHMC treatments decreased the days of gestation, calving to first service, and calving to first visible estrus. Compared with CON at specific time points, TCHMC treatments increased the concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2), whereas progesterone (P4) and E2 concentrations decreased. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and creatine kinase (CK) concentrations were downregulated, whereas that of globulin (GLB) and immunoglobulin G (IgG) were upregulated by TCHMC treatments around the time of calving. Compared with CON and T-200 treatments, the T-300 treatment increased the serum concentrations of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), catalase (CAT) and total antioxidant capacity (T-AOC) and decreased the malondialdehyde (MDA) concentration from 7 d prepartum to 21 d postpartum when. In addition, although TCHMC treatment had no effect on average birth weight, heart rate, respiratory rate, and body temperature of calves, the T-300 treatment increased serum albumin (ALB) and IgG concentrations in calves from 3 to 14 days postpartum. The addition of TCHMC used in the present study could serve as a potential effective strategy to improve the health and productive performance of periparturient dairy cows, and the optimal dose should be set at 300 g per day.

Methylation patterns of Lys9 and Lys27 on histone H3 correlate with patient outcome and tumor progression in lung cancer.

BACKGROUNDS: Histone methylation is recognized as an important component of the epigenetic mechanisms of cancer initiation and progression. Previous studies have demonstrated that aberrant alterations in histone methylation are associated with lung cancer. However, novel and specific epigenetic biomarkers for monitoring lung adenocarcinoma remain unknown. METHODS: A retrospective clinicopathological analysis was performed on 71 lung adenocarcinoma (LUAD) patients who received complete ablative surgical treatment. Tissue arrays were made from the paraffin-embedded LUAD tumor tissues, and these, together with corresponding normal tissues, were examined through immunohistochemistry for several markers: histone 3 lysine 9 di-methylation (H3K9me2), histone 3 lysine 9 tri-methylation (H3K9me3), and histone 3 lysine 27 tri-methylation (H3K27me3). The expression level of each marker was analyzed according to the histological classification and clinical prognosis data. RESULTS: Compared with peri-cancerous tissues, cancerous tissues distinctly expressed higher proportions of H3K9me2, H3K9me3, and H3K27me3. A higher expression pattern of H3K27me3 was associated with the poorly differentiation and unfavorable prognosis in LUAD. Based on histological types, it was found that the H3K27me3 level of patients with micropapillary type is high, and it is related to worse prognosis. CONCLUSIONS: The findings of this study show that the H3K27me3 and micropapillary type are malignant clinical factors of LUAD. H3K27me3 reduction is a novel epigenetic biomarker for defining high-risk LUAD and predicting worse prognosis. Immunohistochemical evaluation of H3K27me3 expression is an economic, easily available, and readily adaptable method.

Four transcription profile-based models identify novel prognostic signatures in oesophageal cancer.

Oesophageal cancer (ESCA) is a clinically challenging disease with poor prognosis and health-related quality of life. Here, we investigated the transcriptome of ESCA to identify high risk-related signatures. A total of 159 ESCA patients of The Cancer Genome Atlas (TCGA) were sorted by three phases. In the discovery phase, differentially expressed transcripts were filtered; in the training phase, two adjusted Cox regressions and two machine leaning models were used to construct and estimate signatures; and in the validation phase, prognostic signatures were validated in the testing dataset and the independent external cohort. We constructed two signatures from three types of RNA markers by Akaike information criterion (AIC) and least absolute shrinkage and selection operator (LASSO) Cox regressions, respectively, and all candidate markers were further estimated by Random Forest (RFS) and Support Vector Machine (SVM) algorithms. Both signatures had good predictive performances in the independent external oesophageal squamous cell carcinoma (ESCC) cohort and performed better than common clinicopathological indicators in the TCGA dataset. Machine learning algorithms predicted prognosis with high specificities and measured the importance of markers to verify the risk weightings. Furthermore, the cell function and immunohistochemical (IHC) staining assays identified that the common risky marker FABP3 is a novel oncogene in ESCA.

A novel circular RNA, hsa-circ-0000211, promotes lung adenocarcinoma migration and invasion through sponging of hsa-miR-622 and modulating HIF1-α expression.

Recently, several studies have evaluated the role of circular RNAs in the metastasis and development of multiple cancers. In our earlier microarray-based study, we had reported the aberrant expression of a novel circular RNA, hsa-circ-0000211 in lung adenocarcinoma (LAC) tissues. However, the roles of hsa-circ-0000211 in LAC have not been studied. Here hsa-circ-0000211 expression in the LAC tissues and cell lines was determined by quantitative real-time PCR (qRT-PCR). The function of hsa-circ-0000211 was evaluated by transwell assay and wound healing. Mechanisms of hsa-circ-0000211 was measured by luciferase reporter assay and western blot. Results revealed the expression of hsa-circ-0000211 in the human LAC tissues and LAC cell lines was higher than that in normal tissue and human lung normal epithelial cells, respectively. The knockdown of hsa-circ-0000211 could inhibit the migration and invasion properties of LAC. Furthermore, hsa-circ-0000211 promoted the migration and invasion of LAC by sponging miR-622. Moreover, hsa-circ-0000211 upregulated the HIF1-α expression by targeting miR-622. hsa-circ-0000211 promoted LAC cell migration and invasion by modulating the miR-622/HIF1-α network. Our study demonstrated that hsa-circ-0000211 can be a potential novel therapeutic target for LAC.

Synaptotagmin 12 (SYT12) Gene Expression Promotes Cell Proliferation and Progression of Lung Adenocarcinoma and Involves the Phosphoinositide 3-Kinase (PI3K)/AKT/Mammalian Target of Rapamycin (mTOR) Pathway.

BACKGROUND This study aimed to use bioinformatics analysis to compare data from tissue microarrays from patients with lung adenocarcinoma (LUAD) and normal lung tissue, and human lung adenocarcinoma cells with normal lung epithelial cells in vitro to investigate the role of synaptotagmin 12 (SYT12) gene expression in LUAD. MATERIAL AND METHODS Human lung adenocarcinoma cell lines (A549, SPC-A-1, H1299, H1975, and PC9) and the normal HBE cell line were compared, and tumor xenografts were developed in mice. The Cancer Genome Atlas (TCGA) tissue microarray data were used to compare SYT12 expression and overall survival (OS). The in vivo and in vitro effects of down-regulation and upregulation of SYT12 were studied using short-interfering RNA (si-RNA) and overexpression plasmids, respectively. The Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) pathway analysis, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), and Western blot investigated the molecular mechanisms of SYT12 expression in LUAD. RESULTS SYT12 expression was increased in tissues from patients with LUAD from TCGA and was associated with advanced tumor stage and reduced prognosis. Knockdown of SYT12 suppressed the proliferation and migration of LUAD cells, and upregulation of SYT12 increased the proliferation and migration of LUAD cells in vitro. Phosphorylation of PIK3R3 activated the PI3K/AKT/mTOR pathway. In the mouse xenograft model, expression of SYT12 increased the volume and weight of the xenograft tumors. CONCLUSIONS Bioinformatics analysis, human LUAD cells, and mouse xenograft studies showed that SYT12 acted as a possible oncogene by phosphorylation of PIK3R3 to activate the PI3K/AKT/mTOR signaling pathway.

Upregulation of the long noncoding RNA TUG1 promotes proliferation and migration of esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is one of the prevalent and deadly cancers worldwide, especially in Eastern Asia. The prognosis of ESCC remains poor; thus, it is still necessary to further dissect the underlying mechanisms and explore therapeutic targets of ESCC. Recent studies show that long noncoding RNAs (lncRNAs) have critical roles in diverse biological processes, including tumorigenesis. Some lncRNAs, such as HOTAIR and POU3F3, were reported to play important roles in ESCC. Here, we characterized the expression profile of taurine-upregulated gene 1 (TUG1), a lncRNA recruiting and binding to polycomb repressive complex 2 (PRC2), in ESCC. In a cohort of 62 patients, TUG1 was significantly overexpressed in ESCC tissues compared with paired adjacent normal tissues, and high expression level of TUG1 was associated with family history and upper segment of esophageal cancer (p < 0.05). Further, in vitro silencing TUG1 via siRNA inhibited the proliferation and migration of ESCC cells and blocked the progression of cell cycle. Therefore, our study indicates that TUG1 promotes proliferation and migration of ESCC cells and is a potential oncogene of ESCC.

Differentially expressed protein-coding genes and long noncoding RNA in early-stage lung cancer.

Due to the application of low-dose computed tomography screening, more and more early-stage lung cancers have been diagnosed. Thus, it is essential to characterize the gene expression profile of early-stage lung cancer to develop potential biomarkers for early diagnosis and therapeutic targets. Here, we analyzed microarray data of 181 early-stage lung cancer patients. By comparing gene expression between different tumor and lymph node metastasis stages, we identified various differentially expressed protein-coding genes and long noncoding RNA (lncRNA) in the comparisons of T2 vs. T2 and N1- vs. N0-stage lung cancer. Functional analyses revealed that these differentially expressed genes were enriched in various tumorigenesis or metastasis-related pathways. Survival analysis indicated that two protein-coding genes, C7 and SCN7A, were significantly associated survival of lung cancer. Notably, a novel lncRNA, LINC00313, was highly expressed in both T2- and N1-stage lung cancers. On the other hand, LINC00313 was also upregulated in lung cancer and metastasized lung cancer tissues, compared with adjacent lung tissues and primary lung cancer tissues. Additionally, higher expression level of LINC00313 indicated poor prognosis of lung cancer (hazard ratio = 0.658). Overall, we characterized the expression profiles of protein-coding genes and lncRNA in early-stage lung cancer and found that LINC00313 could be a biomarker for lung cancer.

Long noncoding RNA CCAT2 correlates with smoking in esophageal squamous cell carcinoma.

Esophageal cancer is one of the leading causes of cancer-related mortality, and most esophageal squamous cell carcinoma (ESCC) cases are located in Asian area. Recent studies about long noncoding RNAs (lncRNA) have offered a new perspective for cancer research and provided new approaches to understand the complex regulation network in cancer. Our group has reported that the novel lncRNA colon cancer-associated transcript 2 (CCAT2) has important biological function and could be a potential biomarkers in lung cancer. Here, we performed in silico analysis and characterized the expression profile of CCAT2 in a cohort of esophageal squamous cell carcinoma (ESCC) patients and cell lines. In silico analysis showed that no CpG island is found in the chromosome region of CCAT2, indicating that the expression of CCAT2 is possibly not regulated by DNA methylation. Compared with paired adjacent normal esophageal tissues, CCAT2 was significantly overexpressed in ESCC tissues with an average fold of 7.18. In ESCC cell lines, CCAT2 was mostly upregulated in KYSE410 cell (24.7-fold upregulation) when normalized to normal esophageal epithelium cell line (HEEC) and most CCAT2 transcripts were located in nucleus (> 95 %). Statistical analysis showed that CCAT2 expression level was significantly associated with smoking status (P = 0.036). Receiver operative curve and the area under curve were calculated to assess the diagnostic potential of CCAT2. Measured by area under curve (AUC), CCAT2 showed higher diagnostic performance than conventional serum biomarkers, like AFP, CA153, and NSE. In this study, we firstly characterized the expression profile of CCAT2 in ESCC and evaluated its potential diagnostic value as a biomarker.

Clinical outcome and expression of mutant P53, P16, and Smad4 in lung adenocarcinoma: a prospective study.

BACKGROUND: Whole-exome sequencing has shown that lung adenocarcinoma (LAC) can be driven by mutant genes, including TP53, P16, and Smad4. The aim of this study was to clarify protein alterations of P53, P16, and Smad4 and to explore their correlations between the protein alterations and clinical outcome. METHODS: We investigated associations among P53 mutant (P53(Mut)) expression, and P16 and Smad4 loss-of-expression, with clinical outcome in 120 LAC patients who underwent curative resection, using immunohistochemical (IHC) methods. RESULTS: Of the 120 patients, 76 (63.3%) expressed P53(Mut) protein, whereas 54 (45.0%) loss of P16 expressed and 75 (62.5%) loss of Smad4 expressed. P53(Mut) expression was associated with tumor size (P = 0.041) and pathological stage (P = 0.025). Loss of P16 expression was associated with lymph node metastasis (P = 0.001) and pathological stage (P < 0.001). Loss of Smad4 expression was associated with tumor size (P = 0.033), lymph node metastasis (P = 0.014), pathological stage (P = 0.017), and tumor differentiation (P = 0.022). Kaplan-Meier survival analysis showed that tumor size (P = 0.031), lymph node metastasis (P < 0.001), pathological stage (P < 0.001), P53(Mut) protein expression (P = 0.038), and loss of p16 or Smad4 expression (P < 0.001) were significantly associated with shorter overall survival(OS), whereas multivariate analysis indicated that lymph node metastasis (P = 0.014) and loss of p16 or Smad4 expression (P < 0.001) were independent prognostic factors. Analysis of protein combinations showed patients with more alterations had poorer survival (P < 0.001). Spearman correlation analysis showed that loss of Smad4 expression inversely correlated with expression of P53(Mut) (r = (-)0.196, P = 0.032) and positively with lost P16 expression (r =0.182, P = 0.047). CONCLUSIONS: The findings indicate that IHC status of P53(Mut), P16, and Smad4 may predict patient outcomes in LAC.

CCAT2 is a lung adenocarcinoma-specific long non-coding RNA and promotes invasion of non-small cell lung cancer.

The prognosis of non-small cell lung cancer (NSCLC) is still poor, and it is necessary to identify effectively diagnostic and prognostic biomarkers for NSCLC. Recent evidence demonstrates that long non-coding RNA (lncRNA) is actively transcribed from human genome. Some lncRNAs show a time- or tissue-specific expression manner and play important roles in diverse biological processes. Additionally, various cancer-associated lncRNAs have been identified, such as metastasis-associated lung adenocarcinoma transcript 1 for lung cancer. Here, we characterized the expression profile of a novel lncRNA, colon cancer-associated transcript 2 (CCAT2), in lung cancer and found that CCAT2 was significantly over-expressed in NSCLC tissues compared with paired adjacent normal tissues, with an average up-regulation fold of 7.5. Intriguingly, over-expression of CCAT2 was significantly associated with lung adenocarcinoma (p=0.033) but not squamous cell cancer. Silencing CCAT2 by siRNA led to inhibition of proliferation and invasion in NSCLC cell lines in vitro. Additionally, CCAT2 combined with CEA could predict lymph node metastasis. Our findings indicate that CCAT2 is a lung adenocarcinoma-specific lncRNA and promotes invasion of NSCLC and highlight its potential as a biomarker for lymph node metastasis.

Value of dual-layer spectral detector CT in predicting lymph node metastasis of non-small cell lung cancer.

Background: The accurate assessment of lymph node metastasis (LNM) is crucial for the staging, treatment, and prognosis of lung cancer. In this study, we explored the potential value of dual-layer spectral detector computed tomography (SDCT) quantitative parameters in the prediction of LNM in non-small cell lung cancer (NSCLC). Methods: In total, 91 patients presenting with solid solitary pulmonary nodules (8 mm < diameter ≤30 mm) with pathologically confirmed NSCLC (57 without LNM, and 34 with LNM) were enrolled in the study. The patients' basic clinical data and the SDCT morphological features were analyzed using the chi-square test or Fisher's exact test. The Mann-Whitney U-test and independent sample t-test were used to analyze the differences in multiple SDCT quantitative parameters between the non-LNM and LNM groups. The diagnostic efficacy of the corresponding parameters in predicting LNM in NSCLC was evaluated by plotting the receiver operating characteristic (ROC) curves. A multivariate logistic regression analysis was conducted to determine the independent predictive factors of LNM in NSCLC. Interobserver agreement was assessed using intraclass correlation coefficients (ICCs) and Bland-Altman plots. Results: There were no significant differences between the non-LNM and LNM groups in terms of age, sex, and smoking history. Lesion size and vascular convergence sign differed significantly between the two groups (P<0.05), but there were no significant differences in the six tumor markers. The SDCT quantitative parameters [SAR40keV, SAR70keV, Δ40keV, Δ70keV, CER40keV, CER70keV, NEF40keV, NEF70keV, λ, normalized iodine concentration (NIC) and NZeff] were significantly higher in the non-LNM group than the LNM group (P<0.05). The ROC analysis showed that CER40keV, NIC, and CER70keV had higher diagnostic efficacy than other quantitative parameters in predicting LNM [areas under the curve (AUCs) =0.794, 0.791, and 0.783, respectively]. The multivariate logistic regression analysis showed that size, λ, and NIC were independent predictive factors of LNM. The combination of size, λ, and NIC had the highest diagnostic efficacy (AUC =0.892). The interobserver repeatability of the SDCT quantitative and derived quantitative parameters in the study was good (ICC: 0.801-0.935). Conclusions: The SDCT quantitative parameters combined with the clinical data have potential value in predicting LNM in NSCLC. The size + λ + NIC combined parameter model could further improve the prediction efficacy of LNM.

[Mutational Signatures Analysis of Micropapillary Components and Exploration of ZNF469 Gene in Early-stage Lung Adenocarcinoma with Ground-glass Opacities].

BACKGROUND: In China, lung cancer remains the cancer with the highest incidence and mortality rate. Among early-stage lung adenocarcinomas (LUAD), the micropapillary (MPP) component is prevalent and typically exhibits high aggressiveness, significantly correlating with early metastasis, lymphatic infiltration, and reduced five-year survival rates. Therefore, the study is to explore the similarities and differences between MPP and non-micropapillary (non-MPP) components in malignant pulmonary nodules characterized by GGOs in early-stage LUAD, identify unique mutational features of the MPP component and analyze the relationship between the ZNF469 gene, a member of the zinc-finger protein family, and the prognosis of early-stage LUAD, as well as its correlation with immune infiltration. METHODS: A total of 31 malignant pulmonary nodules of LUAD were collected and dissected into paired MPP and non-MPP components using microdissection. Whole-exome sequencing (WES) was performed on the components of early-stage malignant pulmonary nodules. Mutational signatures analysis was conducted using R packages such as maftools, Nonnegative Matrix Factorization (NMF), and Sigminer to unveil the genomic mutational characteristics unique to MPP components in invasive LUAD compared to other tumor tissues. Furthermore, we explored the expression of the ZNF469 gene in LUAD using The Cancer Genome Atlas (TCGA) database to investigate its potential association with the prognosis. We also investigated gene interaction networks and signaling pathways related to ZNF469 in LUAD using the GeneMANIA database and conducted Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Lastly, we analyzed the correlation between ZNF469 gene expression and levels of immune cell infiltration in LUAD using the TIMER and TISIDB databases. RESULTS: MPP components exhibited a higher number of genomic variations, particularly the 13th COSMIC (Catalogue of Somatic Mutations in Cancer) mutational signature characterized by the activity of the cytidine deaminase APOBEC family, which was unique to MPP components compared to non-MPP components in tumor tissues. This suggests the potential involvement of APOBEC in the progression of MPP components in early-stage LUAD. Additionally, MPP samples with high similarity to APOBEC signature displayed a higher tumor mutational burden (TMB), indicating that these patients may be more likely to benefit from immunotherapy. The expression of ZNF469 was significantly upregulated in LUAD compared to normal tissue, and was associated with poor prognosis in LUAD patients (P<0.05). Gene interaction network analysis and GO/KEGG enrichment analysis revealed that COL6A1, COL1A1, COL1A2, TGFB2, MMP2, COL8A2 and C2CD4C interacted with ZNF469 and were mainly involved in encoding collagen proteins and participating in the constitution of extracellular matrix. ZNF469 expression was positively correlated with immune cell infiltration in LUAD (P<0.05). CONCLUSIONS: The study has unveiled distinctive mutational signatures in the MPP components of early-stage invasive LUAD in the Asian population. Furthermore, we have identified that the elevated expression of mutated ZNF469 impacts the prognosis and immune infiltration in LUAD, suggesting its potential as a diagnostic and prognostic biomarker in LUAD.

Cancer-associated fibroblasts: Key criminals of tumor pre-metastatic niche.

Cancer-associated fibroblasts (CAFs) are abundant and important components of the tumour mesenchyme, and have been extensively studied for their role in primary tumours. CAFs provide biomechanical support for tumour cells and play key roles in immunosuppression and tumour metastasis. CAFs can promote epithelial-mesenchymal transition (EMT) of the primary tumour by secreting extracellular vesicles (EVs), increasing adhesion to tumour cells, remodelling the extracellular matrix (ECM) of the primary tumour, and changing its mechanical stiffness, which provides a pathway for tumour metastasis. Moreover, CAFs can form cell clusters with circulating tumour cells (CTCs) to help them resist blood shear forces and achieve colonisation of distant host organs. Recent studies have revealed their roles in pre-metastatic niche (PMN) formation and prevention. In this review, we discuss the role of CAFs in PMN formation and therapeutic interventions targeting PMN and CAFs to prevent metastasis.

Value of spectral computed tomography-derived quantitative parameters based on full volume analysis in the diagnosis of benign/malignant and pathological subtypes of solitary pulmonary nodules.

Background: Conventional dynamic computed tomography (CT) has a low specificity for the distinction between benign and malignant solitary pulmonary nodules (SPNs), and spectral CT has been proposed as a potential alternative. We aimed to investigate the role of quantitative parameters based on full-volume spectral CT in the differential diagnosis of SPNs. Methods: This retrospective study included spectral CT images of 100 patients with pathologically confirmed SPNs (78 and 22 in the malignant and benign groups, respectively). All cases were confirmed by postoperative pathology, percutaneous biopsy, and bronchoscopic biopsy. Multiple quantitative parameters derived from spectral CT were extracted from whole-tumor volume and standardized. Differences in quantitative parameters between groups were statistically analyzed. Diagnostic efficiency was evaluated by generating a receiver operating characteristic (ROC) curve. Between-group differences were evaluated using an independent sample t-test or Mann-Whitney U test. Interobserver repeatability was assessed using intraclass correlation coefficients (ICCs) and Bland-Altman plots. Results: Spectral CT-derived quantitative parameters, except attenuation difference between the SPN in the 70 keV and arterial enhancement [ΔS-A(70 keV)], were significantly higher for malignant SPNs than for benign nodules (P<0.05). In the subgroup analysis, most parameters could distinguish between the benign and adenocarcinoma groups, and between the benign and squamous cell carcinoma groups (P<0.05). Only 1 parameter could differentiate the adenocarcinoma and squamous cell carcinoma groups (P=0.020). ROC curve analysis indicated that normalized arterial enhancement fraction in the 70 keV (NEF70 keV), normalized iodine concentration (NIC), and Δ70 keV had high diagnostic efficacy for differentiating SPNs between the benign and malignant SPNs [area under the curve (AUC): 0.867, 0.866, and 0.848, respectively] and between the benign and adenocarcinoma groups (AUC: 0.873, 0.872, and 0.874, respectively). The multiparameters derived from spectral CT exhibited satisfactory interobserver repeatability (ICC: 0.856-0.996). Conclusions: Our study suggests that quantitative parameters derived from whole-volume spectral CT may be useful to improve discrimination of SPNs.

Value of dual-layer spectral detector computed tomography in the diagnosis of benign/malignant solid solitary pulmonary nodules and establishment of a prediction model.

Objective: This study aimed to investigate the role of spectral detector computed tomography (SDCT) quantitative parameters and their derived quantitative parameters combined with lesion morphological information in the differential diagnosis of solid SPNs. Methods: This retrospective study included basic clinical data and SDCT images of 132 patients with pathologically confirmed SPNs (102 and 30 patients in the malignant and benign groups, respectively). The morphological signs of SPNs were evaluated and the region of interest (ROI) was delineated from the lesion to extract and calculate the relevant SDCT quantitative parameters, and standardise the process. Differences in qualitative and quantitative parameters between the groups were statistically analysed. A receiver operating characteristic (ROC) curve was constructed to evaluate the efficacy of the corresponding parameters in the diagnosis of benign and malignant SPNs. Statistically significant clinical data, CT signs and SDCT quantitative parameters were analysed using multivariate logistic regression to determine the independent risk factors for predicting benign and malignant SPNs, and the best multi-parameter regression model was established. Inter-observer repeatability was assessed using the intraclass correlation coefficient (ICC) and Bland-Altman plots. Results: Malignant SPNs differed from benign SPNs in terms of size, lesion morphology, short spicule sign, and vascular enrichment sign (P< 0.05). The SDCT quantitative parameters and their derived quantitative parameters of malignant SPNs (SAR40keV, SAR70keV, Δ40keV, Δ70keV, CER40keV, CER70keV, NEF40keV, NEF70keV, λ, NIC, NZeff) were significantly higher than those of benign SPNs (P< 0.05). In the subgroup analysis, most parameters could distinguish between benign and adenocarcinoma groups (SAR40keV, SAR70keV, Δ40keV, Δ70keV, CER40keV, CER70keV, NEF40keV, NEF70keV, λ, NIC, and NZeff), and between benign and squamous cell carcinoma groups (SAR40keV, SAR70keV, Δ40keV, Δ70keV, NEF40keV, NEF70keV, λ, and NIC). However, there were no significant differences between the parameters in the adenocarcinoma and squamous cell carcinoma groups. ROC curve analysis indicated that NIC, NEF70keV, and NEF40keV had higher diagnostic efficacy for differentiating benign and malignant SPNs (area under the curve [AUC]:0.869, 0.854, and 0.853, respectively), and NIC was the highest. Multivariate logistic regression analysis showed that size (OR=1.138, 95% CI 1.022-1.267, P=0.019), Δ70keV (OR=1.060, 95% CI 1.002-1.122, P=0.043), and NIC (OR=7.758, 95% CI 1.966-30.612, P=0.003) were independent risk factors for the prediction of benign and malignant SPNs. ROC curve analysis showed that the AUC of size, Δ70keV, NIC, and a combination of the three for differential diagnosis of benign and malignant SPNs were 0.636, 0.846, 0.869, and 0.903, respectively. The AUC for the combined parameters was the largest, and the sensitivity, specificity, and accuracy were 88.2%, 83.3% and 86.4%, respectively. The SDCT quantitative parameters and their derived quantitative parameters in this study exhibited satisfactory inter-observer repeatability (ICC: 0.811-0.997). Conclusion: SDCT quantitative parameters and their derivatives can be helpful in the differential diagnosis of benign and malignant solid SPNs. The quantitative parameter, NIC, is superior to the other relevant quantitative parameters and when NIC is combined with lesion size and Δ70keV value for comprehensive diagnosis, the efficacy could be further improved.

Energy stress-induced circZFR enhances oxidative phosphorylation in lung adenocarcinoma via regulating alternative splicing.

BACKGROUND: Circular RNAs (circRNAs) contribute to multiple biological functions and are also involved in pathological conditions such as cancer. However, the role of circRNAs in metabolic reprogramming, especially upon energy stress in lung adenocarcinoma (LUAD), remains largely unknown. METHODS: Energy stress-induced circRNA was screened by circRNA profiling and glucose deprivation assays. RNA-seq, real-time cell analyzer system (RTCA) and measurement of oxygen consumption rate (OCR) were performed to explore the biological functions of circZFR in LUAD. The underlying mechanisms were investigated using circRNA pull-down, RNA immunoprecipitation, immunoprecipitation and bioinformatics analysis of alternative splicing. Clinical implications of circZFR were assessed in 92 pairs of LUAD tissues and adjacent non-tumor tissues, validated in established patient-derived tumor xenograft (PDTX) model. RESULTS: CircZFR is induced by glucose deprivation and is significantly upregulated in LUAD compared to adjacent non-tumor tissues, enhancing oxidative phosphorylation (OXPHOS) for adaptation to energy stress. CircZFR is strongly associated with higher T stage and poor prognosis in patients with LUAD. Mechanistically, circZFR protects heterogeneous nuclear ribonucleoprotein L-like (HNRNPLL) from degradation by ubiquitination to regulate alternative splicing, such as myosin IB (MYO1B), and subsequently activates the AKT-mTOR pathway to facilitate OXPHOS. CONCLUSION: Our study provides new insights into the role of circRNAs in anticancer metabolic therapies and expands our understanding of alternative splicing.

Whole-Exome Sequencing Reveals the Genomic Features of the Micropapillary Component in Ground-Glass Opacities.

BACKGROUND: Micropapillary components are observed in a considerable proportion of ground-glass opacities (GGOs) and contribute to the poor prognosis of patients with invasive lung adenocarcinoma (LUAD). However, the underlying mutational processes related to the presence of micropapillary components remain obscure, limiting the development of clinical interventions. METHODS: We collected 31 GGOs, which were separated into paired micropapillary and non-micropapillary components using microdissection. Whole-exome sequencing (WES) was performed on the GGO components, and bioinformatics analysis was conducted to reveal the genomic features of the micropapillary component in invasive LUAD. RESULTS: The micropapillary component had more genomic variations, including tumor mutation burden, intratumoral heterogeneity, and copy number variation. We also observed the enrichment of AID/APOBEC mutation signatures and an increased activation of the RTK/Ras, Notch, and Wnt oncogenic pathways within the micropapillary component. A phylogenetic analysis further suggested that ERBB2/3/4, NCOR1/2, TP53, and ZNF469 contributed to the micropapillary component's progression during the early invasion of LUAD, a finding that was validated in the TCGA cohort. CONCLUSIONS: Our results revealed specific mutational characteristics of the micropapillary component of invasive LUAD in an Asian population. These characteristics were associated with the formation of high-grade invasive patterns. These preliminary findings demonstrated the potential of targeting the micropapillary component in patients with early-stage LUAD.

Circulating tumor DNA integrating tissue clonality detects minimal residual disease in resectable non-small-cell lung cancer.

BACKGROUND: Circulating tumor DNA (ctDNA) has been proven as a marker for detecting minimal residual diseases following systemic therapies in mid-to-late-stage non-small-cell lung cancers (NSCLCs) by multiple studies. However, fewer studies cast light on ctDNA-based MRD monitoring in early-to-mid-stage NSCLCs that received surgical resection as the standard of care. METHODS: We prospectively recruited 128 patients with stage I-III NSCLCs who received curative surgical resections in our Lung Cancer Tempo-spatial Heterogeneity prospective cohort. Plasma samples were collected before the surgery, 7 days after the surgery, and every 3 months thereafter. Targeted sequencing was performed on a total of 628 plasma samples and 645 matched tumor samples using a panel covering 425 cancer-associated genes. Tissue clonal phylogeny of each patient was reconstructed and used to guide ctDNA detection. RESULTS: The results demonstrated that ctDNA was more frequently detected in patients with higher stage diseases pre- and postsurgery. Positive ctDNA detection at as early as 7 days postsurgery identified high-risk patients with recurrence (HR = 3.90, P < 0.001). Our results also show that longitudinal ctDNA monitoring of at least two postsurgical time points indicated a significantly higher risk (HR = 7.59, P < 0.001), preceding radiographic relapse in 73.5% of patients by a median of 145 days. Further, clonal ctDNA mutations indicated a high-level specificity, and subclonal mutations informed the origin of tumor recurrence. CONCLUSIONS: Longitudinal ctDNA surveillance integrating clonality information may stratify high-risk patients with disease recurrence and infer the evolutionary origin of ctDNA mutations.

90-Gene Expression Profiling for Tissue Origin Diagnosis of Cancer of Unknown Primary.

Cancer of unknown primary (CUP), in which metastatic diseases exist without an identifiable primary location, accounts for about 3-5% of all cancer diagnoses. Successful diagnosis and treatment of such patients are difficult. This study aimed to assess the expression characteristics of 90 genes as a method of identifying the primary site from CUP samples. We validated a 90-gene expression assay and explored its potential diagnostic utility in 44 patients at Jiangsu Cancer Hospital. For each specimen, the expression of 90 tumor-specific genes in malignant tumors was analyzed, and similarity scores were obtained. The types of malignant tumors predicted were compared with the reference diagnosis to calculate the accuracy. In addition, we verified the consistency of the expression profiles of the 90 genes in CUP secondary malignancies and metastatic malignancies in The Cancer Genome Atlas. We also reported a detailed description of the next-generation coding sequences for CUP patients. For each clinical medical specimen collected, the type of malignant tumor predicted and analyzed by the 90-gene expression assay was compared with its reference diagnosis, and the overall accuracy was 95.4%. In addition, the 90-gene expression profile generally accurately classified CUP into the cluster of its primary tumor. Sequencing of the exome transcriptome containing 556 high-frequency gene mutation oncogenes was not significantly related to the 90 genes analysis. Our results demonstrate that the expression characteristics of these 90 genes can be used as a powerful tool to accurately identify the primary sites of CUP. In the future, the inclusion of the 90-gene expression assay in pathological diagnosis will help oncologists use precise treatments, thereby improving the care and outcomes of CUP patients.

FAM83H-AS1 is a noncoding oncogenic driver and therapeutic target of lung adenocarcinoma.

BACKGROUND: Little is known about noncoding oncogenes of lung adenocarcinoma (LUAD), and these potential drivers might provide novel therapeutic targets. METHODS: Since abnormally overexpression of oncogenic drivers is induced by genomic variation, we here utilized genomic, transcriptomic, and clinical prognosis data of The Cancer Genome Atlas (TCGA) LUAD datasets to discover novel drivers from long noncoding RNAs. We further used zebrafish models to validate the biological function of candidates in vivo. The full length of FAM83H-AS1 was obtained by rapid amplification of the cDNA ends assay. RNA pull-down, RNA immunoprecipitation, quantitative mass spectrometry, and RNA sequencing assays were conducted to explore the potential mechanisms. Additionally, we used CRISPR interference (CRISPRi) method and patient-derived tumor xenograft (PDTX) model to evaluate the therapeutic potential of targeting FAM83H-AS1. RESULTS: The results suggest that FAM83H-AS1 is a potential oncogenic driver due to chromosome 8q24 amplification. Upregulation of FAM83H-AS1 results in poor prognosis of LUAD patients in both Jiangsu Cancer Hospital (JSCH) and TCGA cohorts. Functional assays revealed that FAM83H-AS1 promotes malignant progression and inhibits apoptosis. Mechanistically, FAM83H-AS1 binds HNRNPK to enhance the translation of antiapoptotic oncogenes RAB8B and RAB14. Experiments using CRISPRi-mediated xenografts and PDTX models indicated that targeting FAM83H-AS1 inhibited LUAD progression in vivo. CONCLUSIONS: Our work demonstrates that FAM83H-AS1 is a noncoding oncogenic driver that inhibits LUAD apoptosis via the FAM83H-AS1-HNRNPK-RAB8B/RAB14 axis, which highlights the importance and potential roles that FAM83H-AS1 may serve as a novel therapeutic target for LUAD.