Neoadjuvant chemoradiotherapy (neoCRT) followed by surgery is the most common approach for locally advanced resectable esophageal squamous cell carcinoma (ESCC) patients. How neoCRT impacts ESCC tumor immune microenvironment (TIME) has not been fully understood. Single-cell RNA sequencing (scRNA-seq) was conducted to examine the neoCRT-driven cellular and molecular dynamics in 8 pre- and 7 post-neoCRT ESCC samples from 8 male patients. scRNA-seq data of about 112,000 cells were obtained. Expression programs of cell cycle, epithelium development, immune response, and extracellular structure in pre-treatment tumor cells were related to neoCRT response. Spearman correlation between CD8⁺ T cells' cytotoxicity and expression of checkpoint molecules was prominent in pre-neoCRT intermediate activated/exhausted CD8⁺ T cells. NeoCRT increased CD8⁺ T cells’ infiltration but promoted their exhaustion in both major and minor responders. NeoCRT promoted differentiation of Th but demoted that of Treg cells in major responders. Maturation of cDC1s and expression of M2 macrophage markers increased while the number of cDC2s decreased after neoCRT. Higher activities of immune-related pathways in pre-neoCRT CD8⁺ T cells and macrophages, as well as a pronounced decrease of them after neoCRT, correlated with better neoCRT response. Interactions between intermediate activated/exhausted CD8⁺ T and macrophages, cDC1s, and LAMP3⁺ cDCs decreased after neoCRT. Our comprehensive picture of the neoCRT-related immune changes provides deeper insights into immunological mechanisms associated with ESCC response to neoCRT, which may aid in future development of immune-strategies for improving ESCC treatment. This work was supported by the National Natural Science Foundation of China (82072607).

Identification of molecular subtypes that reflect different prognoses and treatment responses, especially immune checkpoint inhibitors (ICIs) in esophageal squamous cell carcinoma (ESCC), is essential for treatment decisions. We performed targeted sequencing in 201 patients with ESCC to discover genetic subtypes and validate our findings via multiple data sets. We identified 3 driver genes ( FCGBP , GRIN2B , and FRY ), and recurrent truncating mutations in FRY impaired its tumor-suppressive function and promoted tumor proliferation. A 3-gene mutation signature ( FAT1 , FAT3 , and FRY ) recognized a molecular subtype named “FAT/FRY” with frequent Hippo pathway–related mutations. In multiple ESCC cohorts, the patients with the FAT/FRY subtype had poorer prognosis than did patients in the WT group. Transcriptome analysis indicated that the FAT/FRY subtype was characterized by inactivation of the Hippo pathway, hypoxia, chemoresistance, higher infiltration of CD8 ⁺ T cells and activated DCs, and a transcriptome similar to that of cancer responders. Furthermore, the 3-gene signature predicted better survival for patients treated with ICIs, partially explained by its positive correlation with the tumor mutation burden and neoantigen burden. The 3-gene signature is a biomarker to recognize the FAT/FRY molecular subtype, evaluate prognosis, and select potential beneficiaries of ICIs in ESCC.

Esophageal squamous cell carcinoma (ESCC) is one of the deadliest malignancies in China. The prognostic value of mutations, especially those in minor tumor clones, has not been systematically investigated. We conducted targeted deep sequencing to analyze the mutation status and the cancer cell fraction (CCF) of mutations in 201 ESCC patients. Our analysis showed that the prognostic effect of mutations was relevant to the CCF, and it should be considered in prognosis prediction. EP300 was a promising biomarker for overall survival, impairing prognosis in a CCF dose-dependent manner. We constructed a CCF-based predictor using a smooth clipped absolute deviation Cox model in the training set of 143 patients. The 3-year disease-free survival rates were 6.3% (95% CI: 1.6–23.9%), 29.8% (20.9–42.6%) and 70.5% (56.6–87.7%) in high-, intermediate- and low-risk patients, respectively, in the training set. The prognostic accuracy was verified in a validation set of 58 patients and the TCGA-ESCC cohort. The eight-gene model predicted prognosis independent of clinicopathological factors and the combination of our model and pathological staging markedly improved the prognostic accuracy of pathological staging alone. Our study describes a novel recurrence predictor for ESCC patients and provides a new perspective for the clinical translation of genomic findings.

Background: The role of adjuvant chemotherapy in patients with pathological lymph node‐positive (pN+) resectable esophageal squamous cell carcinoma (ESCC) remains unclear. We aimed to explore whether adjuvant chemotherapy could improve the overall survival (OS) of patients with pN+ ESCC and whether oral chemotherapy could be used as an alternative to intravenous chemotherapy.Methods: The patients were divided into two groups: a surgery plus chemotherapy group (S + CT group, 400 patients) and a surgery alone group (S group, 582 patients). Propensity score matching (PSM) was used to create patient groups that were balanced across several covariates (n = 331 in each group). The survival rates of patients receiving oral chemotherapy (69 patients with S‐1 and 68 patients with tegafur tablets) and intravenous chemotherapy (263 patients) were compared using the Kaplan–Meier method.Results: In the overall study cohort, the 3‐year OS was significantly higher in the S + CT group than in the S group (66.3% vs. 49.9%, p < 0.001). These data were confirmed in the matched groups (3‐year OS, 72.9% vs. 62.0%, p < 0.001). Multivariate Cox regression analysis in the matched samples showed that adjuvant chemotherapy was an independent prognostic factor for ESCC (HR: 0.62, 95% CI: 0.50–0.76, p < 0.001). Patients who received oral chemotherapy had a similar OS as patients who received intravenous chemotherapy.Conclusions: Adjuvant chemotherapy could significantly improve the OS of patients with pN+ ESCC, and oral chemotherapy drugs might be a better option because of their similar efficacy but fewer side effects than intravenous chemotherapy. This conclusion warrants further study in prospective, randomized controlled trials.

Background: The existing studies on primary bladder lymphoma (PBL) are retrospective analyses based on individual cases or small series studies, and the research on PBL is not unified and in-depth enough at present because of the scarcity of PBL and the lack of relevant literature. This study is designed to develop and validate nomograms for overall survival (OS) and cancer-specific survival (CSS) prediction in patients with PBL. Methods: According to the Surveillance, Epidemiology, and End Results (SEER) database, 405 patients diagnosed with PBL from 1975 to 2016 were collected and randomly assigned to training (n = 283) and validation (n = 122) cohort. After the multivariable Cox regression, the OS and CSS nomograms were developed. The discrimination, calibration and clinical usefulness of the nomograms were assessed and validated, respectively, by the training and validation cohort. Furthermore, all of the patients were reclassified into high- and low-risk groups and their survival were compared through Kaplan-Meier method and log-rank test. Results: Age, subtype, Ann Arbor stage, radiation and chemotherapy were identified as independent prognostic factors for OS and age, sex, and subtype for CSS, then corresponding nomograms predicting the 3- and 5-year survival were constructed. The presented nomograms demonstrated good discrimination and calibration, which the C-index in the training and validation cohort were 0.744 (95% confidence interval [CI], 0.705–0.783) and 0.675 (95% CI, 0.603–0.747) for OS nomogram and 0.692 (95% CI, 0.632–0.752) and 0.646 (95% CI, 0.549–0.743) for CSS nomogram, respectively. Furthermore, the nomograms can be used to effectively distinguish Patients with PBL at high risk of death. The clinical usefulness of the nomograms was visually displayed by decision curve analysis. Conclusion: We updated the baseline characteristics of patients with PBL and constructed OS and CSS nomograms to predict their 3- and 5-year survival. Using these nomograms, it would be convenient to individually predict the prognosis of patients with PBL and provide guidance for clinical treatment.