Yuchen Jiao

Background: Glucose restriction is a hallmark of the tumor microenvironment (TME), yet how tumor cells adapt to this metabolic stress and the impact of metabolic reprogramming on the TME remains incompletely understood. Methods: A genome-wide CRISPR knockout positive screen was performed to identify key mediators of cellular adaptation to glucose restriction. Using biochemistry, molecular biology, metabolomics and confocal immunofluorescent microscopy to elucidate the underlying molecular mechanisms. Additionally, single-cell RNA sequencing and orthotopic tumor models were used to characterize TME remodeling and assess therapeutic efficacy. Results: We identified zinc and ring finger 3 (ZNRF3) as a key mediator of cellular adaptation to glucose restriction through genome-wide CRISPR/Cas9 screening. Multiple tumor cells adaptively survived in glucose-restricted conditions with downregulated ZNRF3. Mechanistically, low glucose suppresses ZNRF3 expression, leading to Wnt pathway activation and subsequent transcriptional repression of stearoyl-CoA desaturase (SCD). This metabolic rewiring enhances antitumor immunity by increasing T-cell infiltration and cytotoxicity. In multiple preclinical models, dietary glucose restriction synergizes with immune checkpoint blockade to suppress tumor growth. Conclusion: These findings establish the ZNRF3-Wnt-SCD axis as a metabolic checkpoint controlling tumor cell fate under glucose restriction and provide a rationale for combining dietary intervention with immunotherapy.

Peritoneal metastasis (PM) remains a major cause of mortality in gastric cancer, yet current diagnostic methods lack sensitivity for early detection. This study aimed to validate the clinical value of minimal residual cancer cells detection in peritoneal lavage fluid (PLF) for predicting and monitoring PM. This study extended follow‐up analysis of a previously reported exploratory cohort ( n = 104) and validated findings in a new validation cohort ( n = 76). Using personalized mutation profiling, we detected cancer cell fraction (CCF) in pre‐resection PLF, post‐resection PLF, and circulating tumor DNA (ctDNA) in matched blood samples. Additionally, we monitored CCF and ctDNA dynamics in five patients receiving intraperitoneal chemotherapy (IPC). In the combined cohort ( n = 180), pre‐resection PLF CCF status showed 98% sensitivity and 80% specificity for PM prediction, while post‐resection PLF demonstrated 82% sensitivity and 90% specificity. Combining pre‐ and post‐resection PLF analysis achieved 100% sensitivity with 80% specificity. Compared to PLF cytology and plasma ctDNA, PLF CCF status emerged as the strongest independent predictor of PM (HR = 177.78, 95% CI: 23.14–22,968.19, p <.0001). In IPC‐treated patients, PLF CCF correlated with peritoneal tumor burden reduction and survival outcomes, highlighting its potential for monitoring therapeutic response. This study establishes PLF CCF detection as a robust and clinically valuable method for early prediction of PM and risk stratification in gastric cancer. In addition, PLF CCF monitoring holds potential for identifying patients who may benefit from prophylactic IPC before clear evidence of PM emerges, as well as for evaluating the efficacy of IPC treatment.

: Liver metastasis remains the primary cause of mortality in patients with colon cancer. Identifying specific driver gene mutations that contribute to metastasis may offer viable therapeutic targets. To explore clonal evolution and genetic heterogeneity within the metastasis, we conducted single-cell exome sequencing on 150 single cells isolated from the primary tumor, liver metastasis, and lymphatic metastasis from a stage IV colon cancer patient. The genetic landscape of the tumor samples revealed that both lymphatic and liver metastases originated from the same region of the primary tumor. Notably, the liver metastasis was derived directly from the primary tumor, bypassing the lymph nodes. Comparative analysis of the sequencing data for individual cell pairs within different tumors demonstrated that the genetic heterogeneity of both liver and lymphatic metastases was also greater than that of the primary tumor. This finding indicates that liver and lymphatic metastases arose from clusters of circulating tumor cell (CTC) of a polyclonal origin, rather than from a single cell from the primary tumor. Single-cell transcriptome analysis suggested that higher EMT score and CNV scores were associated with more polyclonal metastasis. Additionally, a mutation in the TRPS1 (Transcriptional repressor GATA binding 1) gene, TRPS1 R544Q, was enriched in the single cells from the liver metastasis. The mutation significantly increased CRC invasion and migration both in vitro and in vivo through the TRPS1^R544Q/ZEB1 axis. Further TRPS1 mutations were detected in additional colon cancer cases, correlating with advanced-stage disease and inferior prognosis. These results reveal polyclonal seeding and TRPS1 mutation as potential mechanisms driving the development of liver metastases in colon cancer.

Background: Early detection of colorectal cancer (CRC) significantly enhances patient outcomes. Conventional CRC screening tools, like endoscopy and stool-based tests, have constraints due to their invasiveness or suboptimal patient adherence. Recently, liquid biopsy employing plasma cell-free DNA (cfDNA) has emerged as a potential noninvasive screening technique for various malignancies. Methods: In this research, we harnessed the Mutation Capsule Plus (MCP) technology to profile an array of genomic characteristics from cfDNA procured from a single blood draw. This profiling encompassed DNA methylation, the 5’ end motif, copy number variation (CNV), and genetic mutations. An integrated model built upon selected multiomics biomarkers was trained using a cohort of 93 CRC patients and 96 healthy controls. Results: This model was subsequently validated in another cohort comprising 89 CRC patients and 95 healthy controls. Remarkably, the model achieved an area under the curve (AUC) of 0.981 (95% confidence interval (CI), 0.965–0.998) in the validation set, boasting a sensitivity of 92.1% (95% CI, 84.5%-96.8%) and a specificity of 94.7% (95% CI, 88.1%-98.3%). These numbers surpassed the performance of any single genomic feature. Importantly, the sensitivities reached 80% for stage I, 89.2% for stage II, and were 100% for stages III and IV. Conclusion: Our findings underscore the clinical potential of our multiomics liquid biopsy test, indicating its prospective role as a noninvasive method for early-stage CRC detection. This multiomics approach holds promise for further refinement and broader clinical application.

Background Immune checkpoint blockade (ICB) therapy provides remarkable clinical benefits for multiple cancer types. However, the overall response rate to ICB therapy remains low in esophageal squamous cell carcinoma (ESCC). This study aimed to identify biomarkers of ICB therapy for ESCC and interrogate its potential clinical relevance. Methods We investigated gene expression in 42 treatment-naïve ESCC tumor tissues and identified differentially expressed genes, tumor-infiltrating lymphocytes and immune-related genes signatures associated with differential immunotherapy responses. We systematically assessed the tumor microenvironment using the NanoString GeoMx digital spatial profiler, single-cell RNA-seq and multiplex immunohistochemistry in ESCC. Finally, we evaluated the associations between HLA-A-positive tertiary lymphoid structures (TLSs) and patients’ responses to ICB in 60 ESCC patients. Results Tumor infiltrating B lymphocytes and several immune-related gene signatures, such as the antigen presenting machinery (APM) signature, are significantly elevated in ICB treatment responders. Multiplex immunohistochemistry identified the presence of HLA-A⁺ TLSs and showed that TLS-resident cells increasingly express HLA-A as TLSs mature. Most TLS-resident HLA-A⁺ cells are tumor-infiltrating T (TIL-T) or tumor-infiltrating B (TIL-B) lymphocytes. Digital spatial profiling of spatially distinct TIL-T lymphocytes and single-cell RNA-seq data from 60 ESCC tumor tissues revealed that CXCL13-expressing exhausted TIL-Ts inside TLSs are reactivated with elevated expression of the APM signature as TLSs mature. Finally, we demonstrated that HLA-A⁺ TLSs and their major cellular components, TIL-Ts and TIL-Bs, are associated with a clinical benefit from ICB treatment for ESCC. Conclusions HLA-A⁺ TLSs are present in ESCC tumor tissues. TLS-resident TIL-Ts with elevated expression of the APM signature may be reactivated. HLA-A⁺ TLSs and their major cellular components, TIL-Ts and TIL-Bs, may serve as biomarkers for ICB-treated ESCC patients.

Accurate presurgical prediction of pathological complete response (pCR) can guide treatment decisions, potentially avoiding unnecessary surgeries and improving the quality of life for cancer patients. We developed a minimal residual disease (MRD) profiling approach with enhanced sensitivity and specificity for detecting minimal tumor DNA from cell-free DNA (cfDNA). The approach was validated in two independent esophageal squamous cell carcinoma (ESCC) cohorts. In a cohort undergoing neoadjuvant, surgical, and adjuvant therapy (NAT cohort), presurgical MRD status precisely predicted pCR. All MRD-negative cases (10/10) were confirmed as pCR by pathological evaluation on the resected tissues. In contrast, MRD-positive cases included all the 27 non-pCR cases and only one pCR case (10/10 vs 1/28, P < 0.0001, Fisher’s exact test). In a definitive radiotherapy cohort (dRT cohort), post-dRT MRD status was closely correlated with patient prognosis. All MRD-negative patients (25/25) remained progression-free during the follow-up period, while 23 of the 26 MRD-positive patients experienced disease progression (25/25 vs 3/26, P < 0.0001, Fisher’s exact test; progression-free survival, P < 0.0001, log-rank test). The MRD profiling approach effectively predicted the ESCC patients who would achieve pCR with surgery and those likely to remain progression-free without surgery. This suggests that the cancer cells in these MRD-negative patients have been effectively eliminated and they could be suitable candidates for a watch-and-wait strategy, potentially avoiding unnecessary surgery.

Pancreatic intraepithelial neoplasias (PanINs) are the most common precursors of pancreatic cancer, but their small size and inaccessibility in humans make them challenging to study¹. Critically, the number, dimensions and connectivity of human PanINs remain largely unknown, precluding important insights into early cancer development. Here, we provide a microanatomical survey of human PanINs by analysing 46 large samples of grossly normal human pancreas with a machine-learning pipeline for quantitative 3D histological reconstruction at single-cell resolution. To elucidate genetic relationships between and within PanINs, we developed a workflow in which 3D modelling guides multi-region microdissection and targeted and whole-exome sequencing. From these samples, we calculated a mean burden of 13 PanINs per cm³ and extrapolated that the normal intact adult pancreas harbours hundreds of PanINs, almost all with oncogenic KRAS hotspot mutations. We found that most PanINs originate as independent clones with distinct somatic mutation profiles. Some spatially continuous PanINs were found to contain multiple KRAS mutations; computational and in situ analyses demonstrated that different KRAS mutations localize to distinct cell subpopulations within these neoplasms, indicating their polyclonal origins. The extensive multifocality and genetic heterogeneity of PanINs raises important questions about mechanisms that drive precancer initiation and confer differential progression risk in the human pancreas. This detailed 3D genomic mapping of molecular alterations in human PanINs provides an empirical foundation for early detection and rational interception of pancreatic cancer.