A nuclease-dead Cas9-derived tool represses target gene expression.

Manipulation of gene expression is central to understanding gene function, engineering cell behavior, and altering biological traits according to production demands. Nuclease-dead Cas9 (dCas9), a variant of active Cas9, offers a versatile platform for the precise control of genome function without DNA cleavage. Notably, however, an effective and universal dCas9-based transcriptional repression system remains unavailable in plants. The noncanonical histone acetyltransferase TENDRIL-LESS (CsTEN) is responsible for chromatin loosening and histone modification in cucumber (Cucumis sativus). In this study, we engineered a gene regulation tool by fusing TEN and its truncated proteins with dCas9. The full-length dCas9-TEN protein substantially repressed gene expression, with the N-terminal domain identified as the core repression domain. We subsequently validated the specificity and efficacy of this system through both transient infection and genetic transformation in cucumber and Arabidopsis (Arabidopsis thaliana). The electrophoretic mobility shift assay (EMSA) revealed the ability of the N-terminal domain of TEN to bind to chromatin, which may promote target binding of the dCas9 complex and enhance the transcriptional repression effect. Our tool enriches the arsenal of genetic regulation tools available for precision breeding in crops.

A cytosine base editor toolkit with varying activity windows and target scopes for versatile gene manipulation in plants.

CRISPR/Cas-derived base editing tools empower efficient alteration of genomic cytosines or adenines associated with essential genetic traits in plants and animals. Diversified target sequences and customized editing products call for base editors with distinct features regarding the editing window and target scope. Here we developed a toolkit of plant base editors containing AID10, an engineered human AID cytosine deaminase. When fused to the N-terminus or C-terminus of the conventional Cas9 nickase (nSpCas9), AID10 exhibited a broad or narrow activity window at the protospacer adjacent motif (PAM)-distal and -proximal protospacer, respectively, while AID10 fused to both termini conferred an additive activity window. We further replaced nSpCas9 with orthogonal or PAM-relaxed Cas9 variants to widen target scopes. Moreover, we devised dual base editors with AID10 located adjacently or distally to the adenine deaminase ABE8e, leading to juxtaposed or spaced cytosine and adenine co-editing at the same target sequence in plant cells. Furthermore, we expanded the application of this toolkit in plants for tunable knockdown of protein-coding genes via creating upstream open reading frame and for loss-of-function analysis of non-coding genes, such as microRNA sponges. Collectively, this toolkit increases the functional diversity and versatility of base editors in basic and applied plant research.

De novo transcriptome assembly and molecular response mechanism analysis of a diatom Cyclotella meneghiniana Kützing exposed to cadmium.

Cadmium is a persistent heavy metal commonly found in aquatic ecosystems and has a strong toxic effect on organisms. The sensitivity of phytoplankton to environmental changes and its role as an indicator of aquatic ecosystem health have been well-established. However, the mechanisms by which phytoplankton respond to cadmium remain incompletely understood. In this study, we chose the typical planktonic diatom Cyclotella meneghiniana Kützing, by integrating physiological-biochemical data and transcriptome analysis, to reveal the molecular mechanisms of C. meneghiniana responing to cadmium. Under cadmium stress, the cell density and chlorophyll-a content of C. meneghiniana significantly decreased, while MDA content and SOD activity gradually increased. At 72 h of cadmium stress, we found that at this time point, cell abundance and physiological variation were very significant, therefore we selected 72 h for subsequent analysis. To better understand the cadmium stress response mechanisms of C. meneghiniana, a de novo transcriptome method was used to analyse C. meneghiniana under cadmium stress for 72 h, and 1704 (M vs. CK) and 4788 (H vs. CK) differentially expressed genes were found. Our results showed that the changes in gene expression were closely correlated to the physiological-biochemical changes. Although cadmium stress could promote the nitrogen metabolism pathway, ROS scavenging system, and photosynthesis. While, C. meneghiniana under medium and high concentrations of cadmium can also limit various intracellular metabolic pathways, such as the MAPK pathway and phosphatidylinositol metabolic pathway, and the degree of inhibition increases with the increase of stress concentration. In present study, the complete molecular mechanism of the planktonic diatom response to cadmium has been established, which provided important information for further studies on heavy metal pollutants and the multiple functional genes responsible for cadmium sensitivity and tolerance in planktonic diatoms.

Fault Diagnosis Method for Space Fluid Loop Systems Based on Improved Evidence Theory.

Addressing the challenges posed by the complexity of the structure and the multitude of sensor types installed in space application fluid loop systems, this paper proposes a fault diagnosis method based on an improved D-S evidence theory. The method first employs the Gaussian affiliation function to convert the information acquired by sensors into BPA functions. Subsequently, it utilizes a pignistic probability transformation to convert the multiple subset focal elements into single subset focal elements. Finally, it comprehensively evaluates the credibility and uncertainty factors between evidences, introducing Bray-Curtis dissimilarity and belief entropy to achieve the fusion of conflicting evidence. The proposed method is initially validated on the classic Iris dataset, demonstrating its reliability. Furthermore, when applied to fault diagnosis in space application fluid circuit loop pumps, the results indicate that the method can effectively fuse multiple sensors and accurately identify faults.

Molecular subtyping of small-cell lung cancer based on mutational signatures with different genomic features and therapeutic strategies.

Small-cell lung cancer (SCLC) is an exceptionally lethal malignancy characterized by extremely high alteration rates and tumor heterogeneity, which limits therapeutic options. In contrast to non-small-cell lung cancer that develops rapidly with precision oncology, SCLC still remains outside the realm of precision medicine. No recurrent and actionable mutations have been detected. Additionally, a paucity of substantive tumor specimens has made it even more difficult to classify SCLC subtypes based on genetic background. We therefore carried out whole-exome sequencing (WES) on the largest available Chinese SCLC cohort. For the first time, we partitioned SCLC patients into three clusters with different genomic alteration profiles and clinical features based on their mutational signatures. We showed that these clusters presented differences in intratumor heterogeneity and genome instability. Moreover, a wide existence of mutually exclusive gene alterations, typically within similar biological functions, was detected and suggested a high SCLC intertumoral heterogeneity. Particularly, Cluster 1 presented the greatest potential to benefit from immunotherapy, and Cluster 3 constituted recalcitrant SCLC, warranting biomarker-directed drug development and targeted therapies in clinical trials. Our study would provide an in-depth insight into the genome characteristics of the Chinese SCLC cohort, defining distinct molecular subtypes as well as subtype-specific therapies and biomarkers. We propose tailoring differentiated therapies for distinct molecular subgroups, centering on a personalized precision chemotherapy strategy combined with immunization or targeted therapy for patients with SCLC.

Effect of thoracic radiotherapy dose on the prognosis of advanced lung adenocarcinoma harboring EGFR mutations.

BACKGROUND: The aim of this study was to investigate the effects of different thoracic radiotherapy doses on OS and incidence of radiation pneumonia which may provide some basis for optimizing the comprehensive treatment scheme of these patients with advanced EGFR mutant lung adenocarcinoma. METHODS: Data from 111 patients with EGFR-mutant lung adenocarcinoma who received thoracic radiotherapy were included in this retrospective study. Overall survival (OS) was the primary endpoints of the study. Kaplan-Meier method was used for the comparison of OS. The Cox proportional-hazard model was used for the multivariate and univariate analyses to determine the prognostic factors related to the disease. RESULTS: The mOS rates of the patients, who received radiotherapy dose scheme of less than 50 Gy, 50-60 Gy (including 50 Gy), and 60 Gy or more were 29.1 months, 34.4 months, and 51.0 months, respectively (log-rank P = 0.011). Although trend suggested a higher levels of pneumonia cases with increasing radiation doses, these lack statistical significance (χ2 = 1.331; P = 0.514). The multivariate analysis showed that the thoracic radiotherapy dose schemes were independently associated with the improved OS of patients (adjusted hazard ratio [HR], 0.606; 95% CI, 0.382 to 0.961; P = 0.033). CONCLUSIONS: For the patients with advanced EGFR-mutant lung adenocarcinoma, the radical thoracic radiotherapy dose scheme (≥ 60 Gy) could significantly prolong the OS of patients during the whole course management.

Dynamic changes in the systemic immune-inflammation index predict the prognosis of EGFR-mutant lung adenocarcinoma patients receiving brain metastasis radiotherapy.

BACKGROUND: The systemic immune-inflammation index (SII) has recently emerged as a predictor of survival in non-small cell lung cancer patients. There is also tight correlation between radiotherapy and immune status, and brain metastases (BM) radiotherapy is an important treatment in patients with BM from lung adenocarcinoma harboring epidermal growth factor receptor (EGFR) mutations. Hence, this study aimed to present the prognostic value of SII and its dynamic changes during BM radiotherapy in EGFR-mutant lung adenocarcinoma patients with BM. METHODS: Patients with EGFR-mutant lung adenocarcinoma who received BM radiotherapy between November 2011 and April 2021 were included in this retrospective study. The SII was calculated using data acquired within 1 week before the start of radiation treatment and 1 week before its completion. According to the cutoff value of SII before radiation treatment determined using receiver operating characteristic curve analyses, we divided the patients into a high group and a low group. Patients were further classified into high-high, high-low, low-low, and low-high groups based on dynamic changes in SII. Prognostic values of the SII and other factors were determined using the Kaplan-Meier method, as well as univariate and multivariate Cox analysis. RESULTS: A total of 202 patients met the inclusion criteria, and the median overall survival (OS) of the entire cohort was 36 months. According to the SII cutoff of 859.79, an SII value below this cutoff was associated with longer OS (hazard ratio 0.6653, 95% confidence interval 0.4708-0.9402, P < 0.05). The patients in the low-low group, whose SII within 1 week before the start and end of BM radiotherapy were below the cutoff, had a median OS of 55.2 months, which was significantly longer than the OS in all other groups (P < 0.05). Univariate and multivariate analyses confirmed that dynamic SII change (P = 0.032), Lung-molGPA (P < 0.001), and thoracic radiation (P = 0.048) were independently correlated with OS. CONCLUSIONS: The SII and its dynamic change may have a prognostic value in patients with EGFR-mutant lung adenocarcinoma treated with BM radiotherapy.

EPHA5 mutation was associated with adverse outcome of atezolizumab treatment in late-stage non-small cell lung cancers.

BACKGROUND: The aim of the study was to investigate predictive value of gene mutation for atezolizumab treatment response from OAK and POPLAR cohorts. METHODS: Several public databases were used for analyzing gene mutation type of EPHA5 and association with alterations of other genes. Survival analysis was performed for patients receiving atezolizumab from OAK and POPLAR cohorts. RESULTS: EPHA5 mutation have high frequency to harbor TP53 and KEAP1 mutations. The bTMB value has significant difference between EPHA5 mutant and wild-type cases. Patients with EPHA5 mutation got worse survival compared to those without gene mutations receiving atezolizumab (P = 0.0186). CONCLUSIONS: EPHA5 mutant NSCLC may represent a subpopulation which showed worse response after treatment of atezolizumab compared to wild-type ones.

Pine Pollen Polysaccharides' and Sulfated Polysaccharides' Effects on UC Mice through Modulation of Cell Tight Junctions and RIPK3-Dependent Necroptosis Pathways.

The purpose of this study is to explore the effects of pine pollen polysaccharides and sulfated polysaccharides on mice with ulcerative colitis and whether they could protect mice from inflammation by regulating the tight junctions of colonic epithelial cells and regulating the RIPK3-dependent necroptosis pathways. Pine pollen polysaccharides were prepared by water boiling and ethanol precipitation. After deproteinedization with trichloroacetic acid, the UV spectrum showed that there were no proteins. One polysaccharide component (PPM60-III) was made by gel filtration chromatography, and then sulfated polysaccharide (SPPM60-III) was derived using the chlorosulfonic acid-pyridine method. After treatment with PPM60-III and SPPM60-III, the body weight of mice with ulcerative colitis induced by dextran sodium sulfate increased, the DAI score decreased, the levels of pro-inflammatory factors and inflammation-related enzymes decreased, and the level of anti-inflammatory factors increased. In addition, after treatment, the expressions levels of tight junction proteins increased, the expressions levels of key proteins of programmed necroptosis decreased, while the level of Caspase-8 increased. The results indicated that pine pollen polysaccharides and sulfated polysaccharides have a certain therapeutic effect on UC mice, and the therapeutic effect may be achieved by regulating the tight junction of colonic epithelial cells and regulating the RIPK3-dependent necroptosis pathways.

Effect of brain radiotherapy strategies on prognosis of patients with EGFR-mutant lung adenocarcinoma with brain metastasis.

PURPOSE: Epidermal growth factor receptor (EGFR)-mutant lung cancers have a high risk of developing brain metastases (BM). Whole brain radiotherapy (WBRT), local radiotherapy, and WBRT + Boost are frequently used for treatment of BM. This retrospective study aimed to evaluate the difference in efficacy of these radiotherapy modes in patients with EGFR-mutant lung adenocarcinoma with BMs. Further, we determined the optimal radiotherapy regimen for patients based on Lung-molGPA. METHODS AND MATERIALS: We retrospectively enrolled 232 patients with EGFR-mutant lung adenocarcinoma with BMs. Patients were divided into three groups based on the different modes of brain radiotherapy: WBRT group, local radiotherapy group, and WBRT + Boost group. Graded prognostic assessment for lung cancer using molecular markers (Lung molGPA), overall survival (OS), and intracranial progression-free survival (iPFS) were calculated. Kaplan-Meier was used to compare iPFS and OS in different groups. RESULTS: The median OS for the WBRT (n = 84), local radiotherapy (n = 65), and WBRT + Boost (n = 83) cohorts was 32.8, 59.1, and 41.7 months, respectively (P = 0.0002). After stratification according to the Lung-molGPA score, the median OS for the WBRT (n = 56), local radiotherapy (n = 19), and WBRT + Boost (n = 28) cohorts was 32.5, 30.9, and 30.8 months, respectively, in subgroup with score 1-2 (P = 0.5097). In subgroup with score 2.5-4, the median OS for the WBRT (n = 26), local radiotherapy (n = 45), and WBRT + Boost (n = 54) cohorts was 32, 68.4, and 51 months, respectively (P = 0.0041). CONCLUSION: The present study showed that in patients with EGFR-mutant lung adenocarcinoma with BM, local radiotherapy and WBRT + Boost perform similarly well both in the subgroups with low and high scores of Lung-molGPA. Considering the side effect caused by whole brain radiotherapy, we recommended local radiotherapy as optimal brain radiation mode for those subtype lung cancer patients.

Barriers to accessing internet-based home Care for Older Patients: a qualitative study.

BACKGROUND: Due to the increasingly ageing society and the shortage of nursing human resources in China, the imbalance between the home care needs of older patients and the inadequate supply of nursing services is increasing. Based on this medical situation, China is implementing internet-based home care (with the nurses who provide this care called online nurses or sharing nurses) based on the concept of the sharing economy, internet technology and knowledge from the home care experience in other countries. Internet-based home care follows an online application/offline service model. Patients place orders through an app, nurses grab orders instantly, and managers dispatch orders through a web platform based on various factors such as nurses' qualifications, professionalism and distance from the patient. In this way, home care is provided for patients with limited mobility, such as older or disabled patients, patients in rehabilitation and terminal patients. Only by fully understanding the barriers to accessing internet-based home care can we provide quality nursing services to older patients and achieve the sustainable development of internet-based home care. OBJECTIVE: The goal of this study was to use qualitative methods to explore barriers to accessing internet-based home care for older patients. METHODS: Based on Levesque's access to health care framework, semi-structured personal interviews were conducted with 19 older patients in a descriptive qualitative study using directed content analysis. RESULTS: We identified four barriers to accessing internet-based home care for older patients: barriers to perceiving, seeking, paying for, and engaging in internet-based home care. Specific barriers included traditional perceptions, barriers to internet use, high payment costs, uneven quality of services, and concerns about privacy and patient safety. CONCLUSIONS: Internet-based home care brings new risks and challenges. In order to enable older patients to better enjoy it, it is necessary to strengthen publicity, optimize the network application process, improve the health insurance system, formulate unified nursing service standards, and address safety risks.

Multiplex and optimization of dCas9-TV-mediated gene activation in plants.

Synthetic gene activators consisting of nuclease-dead Cas9 (dCas9) for single-guide RNA (sgRNA)-directed promoter binding and a transcriptional activation domain (TAD) represent new tools for gene activation from endogenous genomic locus in basic and applied plant research. However, multiplex gene coactivation by dCas9-TADs has not been demonstrated in whole plants. There is also room to optimize the performance of these tools. Here, we report that our previously developed gene activator, dCas9-TV, could simultaneously upregulate OsGW7 and OsER1 in rice by up to 3,738 fold, with one sgRNA targeting to each promoter. The gene coactivation could persist to at least the fourth generation. Astonishingly, the polycistronic tRNA-sgRNA expression under the maize ubiquitin promoter, a Pol II promoter, could cause enormous activation of these genes by up to >40,000-fold in rice. Moreover, the yeast GCN4 coiled coil-mediated dCas9-TV dimerization appeared to be promising for enhancing gene activation. Finally, we successfully introduced a self-amplification loop for dCas9-TV expression in Arabidopsis to promote the transcriptional upregulation of AtFLS2, a previously characterized dCas9-TV-refractory gene with considerable basal expression. Collectively, this work illustrates the robustness of dCas9-TV in multigene coactivation and provides broadly useful strategies for boosting transcriptional activation efficacy of dCas9-TADs in plants.

Comprehensive analysis of multiple parameters associated with tumor immune microenvironment in ARID1A mutant cancers.

Aim: To verify the relationship between ARID1A and tumor immune microenvironment thus immune checkpoint inhibitors (ICIs) response. Material & methods: Several public databases were used to characterize the association between ARID1A gene alteration and tumor immunity. Results: The gene mutation frequency was 8.2% in all cancer types. The ARID1A-mutated cancers have higher scores of mutation count, tumor mutational burden, neoantigen load (p < 0.001) and T cell repertoire, B cell repertoire diversity (p < 0.05). The gene mutation has tight association with multiple-activated immune cells. Survival analysis suggested that patients with ARID1A mutant cancers benefit more from ICIs treatment (p = 0.013). Conclusion: The ARID1A gene mutation was correlated with higher tumor immunogenicity and activated antitumor immune microenvironment, resulting in superior cohort that respond well to ICIs.

Gene disruption through base editing-induced messenger RNA missplicing in plants.

Gene knockout tools are highly desirable for basic and applied plant research. Here, we leverage the Cas9-derived cytosine base editor to introduce precise C-to-T mutations to disrupt the highly conserved intron donor site GT or acceptor site AG, thereby inducing messenger RNA (mRNA) missplicing and gene disruption. As proof of concept, we successfully obtained Arabidopsis null mutant of MTA gene in the T2 generation and rice double null mutant of GL1-1 and NAL1 genes in the T0 generation by this strategy. Elimination of the original intron donor site or acceptor site could trigger aberrant splicing at a new specific exonic site, but not at the closest GT or AG site, suggesting cryptic rules governing splice site recognition. The strategy presented expands the applications of base editing technologies in plants by providing a new means for gene inactivation without generating DNA double-strand breaks, and it can potentially serve as a useful tool for studying the biology of mRNA splicing.

A hypothesized TNM staging system based on the number and location of positive lymph nodes may better reflect the prognosis for patients with NSCLC.

BACKGROUND: This study aimed to evaluate the feasibility and prognostic accuracy of incorporating the number of positive lymph nodes (PLN) into the TNM staging system for non-small cell lung cancer (NSCLC) patients. METHODS: We screened a total of 9539 patients with resected stage IA-IIIB non-small cell cancer between 2010 and 2015 from SEER database. The chi-square test was used to compare patient baseline characteristics and the X-tile model was applied to determine cut-off values for the number of PLN (nN). The X-tile model was used to screen three different cut-off values including nN = 0, nN1-3 and nN4-. Univariate and multivariate Cox proportional hazards regression models were used to analyze the influence of different variables on overall survival (OS). Kaplan-Meier and log-rank test were used to compare survival differences. RESULTS: Based on the nN cutoffs, we conducted the univariate and multivariate Cox proportional hazards regression. The result showed that nN stage was a significant prognostic factor affecting patients' OS (all P <  0.001). We reclassified the seventh edition TNM stages of the enrolled patients with stage IA-IIIB NSCLC according to the 5-year OS rate. Hypothesized TNM substage based on the location and the number of PLN was further calculated. Then we drew survival curves for each substage, including for the current TNM stage and the hypothesized TNM stage. From the comparison of survival curves, we found that the survival curve of each substage of the hypothesized TNM classification was proportional and well distributed compared with the current TNM classification (P <  0.001). CONCLUSION: Revised TNM staging integrating locational pN stage and numerical nN stage was a more accurate prognostic determinant in patients with NSCLC.

Poor survival of non-small-cell lung cancer patients with main bronchus tumor: a large population-based study.

Aim: In this study, we evaluated the association between tumor location and prognosis in non-small-cell lung cancer patients. Patients & methods: The SEER database was used to screen for suitable patients using our inclusion criteria. The χ2 test was used to compare baseline patient characteristics and the Kaplan-Meier method as well as the log-rank test were used to compare survival differences. At last, univariate and multivariate Cox proportional hazards regression models were used to analyze the influence of different variables on overall survival. Results: The results found no significant difference in overall survival between patients in laterality (p = 0.071). However, patients with main bronchial tumors had worse prognosis than tumors at other locations (p < 0.001). Our results also showed that tumor location including main bronchus, upper lobe, middle lobe, lower lobe and overlapping lesion was a significant factor affecting survival (p < 0.001). Subgroup analysis revealed that regardless of histology or M stage, patients with main bronchial tumors had a worse survival compared with other tumor locations (all; p < 0.001). Interestingly, we found that patients with tumor main bronchial tumors were more likely to be squamous carcinoma and terminal Tumor, Node, Metastasis stage (all; p < 0.001). Conclusion: Non-small-cell lung cancer patients' prognosis was related to the tumor location. And patients with tumors located in main bronchus had worse outcomes than those located in other locations. Tumor primary site should be considered in treatment management and prognosis assessment.

The prognostic analysis of different metastatic patterns in extensive-stage small-cell lung cancer patients: a large population-based study.

AIM: To analyze the metastasis patterns and prognosis differences for extensive-stage small-cell lung cancer patients. METHODS: Log-rank tests were used to calculate and compare survival estimates. Cox regression analyses were used to evaluate the prognosis factors. RESULTS: The liver was the most common metastatic site, and lung was the least common. In two metastatic sites, liver and bone metastases were the most common combination of sites. An isolated liver metastasis had the worst overall survival (OS) and cancer-specific survival (CSS) among metastatic sites (both p < 0.001). Liver and lung metastases were associated with worse CSS (p < 0.039) and OS (p < 0.015). However, for patients with three metastatic sites showed no statistical differences in their CSS and OS (all, p > 0.05). CONCLUSION: Extensive-stage small-cell lung cancer patients with metastasis to the liver alone or in combination with other organs appear to have worse outcomes.

Activation of the BMP-BMPR pathway conferred resistance to EGFR-TKIs in lung squamous cell carcinoma patients with EGFR mutations.

The empirical criteria for defining a clinical subtype of lung cancer are gradually transiting from histopathology to genetic variations in driver genes. Targeting these driver mutations, such as sensitizing epidermal growth factor receptor (EGFR) mutations, has dramatically improved the prognosis of advanced non-small cell lung cancer (NSCLC). However, the clinical benefit of molecularly targeted therapy on NSCLC appears to be different between lung adenocarcinomas and squamous cell carcinomas (SqCCs). We report here that the resistance of lung SqCC harboring EGFR mutations to EGFR tyrosine kinase inhibitors (EGFR-TKIs) was due to the activation of BMP-BMPR-Smad1/5-p70S6K. The combined treatment of these tumor cells with EGFR-TKI, together with inhibitors specific to BMPR or downstream mTOR, effectively reversed the resistance to EGFR-TKI. Moreover, blocking the whole PI3K-AKT-mTOR pathway with the PI3K/mTOR dual inhibitor BEZ235 also showed efficacy in treating this subtype of lung SqCC. This study details the empirical basis for a feasible clinical solution for squamous cell carcinomas with EGFR mutations.

Prognostic significance of the lymphocyte-to-monocyte ratio and the tumor-infiltrating lymphocyte to tumor-associated macrophage ratio in patients with stage T3N0M0 esophageal squamous cell carcinoma.

PURPOSE: We assessed the prognostic significance of, and the relationship between, the pretreatment lymphocyte-to-monocyte ratio (LMR) and the TILs/tumor-associated macrophages (TAMs) ratio, in patients with esophageal squamous cell carcinoma (ESCC) of pathological stage T3N0M0 (pT3N0M0). METHODS: A total of 220 newly diagnosed ESCC patients of stage pT3N0M0 who had not undergone neoadjuvant therapy were included. Densities of CD8+ TILs, CD4+ TILs, CD45RO+ TILs, and CD68+ TAMs were assessed by immunohistochemical staining of tissue microarray cores from all 220 pT3N0M0 ESCC patients (who underwent radical resection). Hematological biomarkers including lymphocyte and monocyte counts were obtained from routine preoperative blood test data, and the LMR and TILs/TAMs ratios calculated. Cutoff finder for survival prediction was plotted to find out the optimal cutoff point for each parameter. RESULTS: The LMR and TILs/TAMs ratios were interrelated. On univariate analyses of data from the entire cohort, the LMR, CD45RO/CD68 ratio, and CD8/CD68 ratio were significantly associated with both OS and disease-free survival. Only the CD45RO/CD68 ratio was independently prognostic of survival on multivariate analysis. CONCLUSIONS: The prognostic significance of the CD45RO/CD68 ratio was higher than that of the LMR. The CD45RO/CD68 ratio is a useful independent prognostic marker in patients with pT3N0M0 ESCC who have undergone complete resection without neoadjuvant therapy.