Tumor-Host Cometabolism Collaborates to Shape Cancer Immunity.

SUMMARY: Nutrients are essential for supporting tumor growth and immune cell function in the tumor microenvironment, but emerging evidence reveals a paradoxical competition and collaboration between the metabolic demands of proliferating cancer cells and immune cell activation. Dietary interventions and metabolic immunoengineering offer promise to selectively modulate cancer and immune cell metabolism by targeting metabolic sensing processes rather than pathways directly, moving beyond conventional ideas and heralding an exciting new era of immunometabolism discovery and translation.

Neutrophil profiling illuminates anti-tumor antigen-presenting potency.

Neutrophils, the most abundant and efficient defenders against pathogens, exert opposing functions across cancer types. However, given their short half-life, it remains challenging to explore how neutrophils adopt specific fates in cancer. Here, we generated and integrated single-cell neutrophil transcriptomes from 17 cancer types (225 samples from 143 patients). Neutrophils exhibited extraordinary complexity, with 10 distinct states including inflammation, angiogenesis, and antigen presentation. Notably, the antigen-presenting program was associated with favorable survival in most cancers and could be evoked by leucine metabolism and subsequent histone H3K27ac modification. These neutrophils could further invoke both (neo)antigen-specific and antigen-independent T cell responses. Neutrophil delivery or a leucine diet fine-tuned the immune balance to enhance anti-PD-1 therapy in various murine cancer models. In summary, these data not only indicate the neutrophil divergence across cancers but also suggest therapeutic opportunities such as antigen-presenting neutrophil delivery.

An Inflammatory Checkpoint Generated by IL1RN Splicing Offers Therapeutic Opportunity for KRAS-Mutant Intrahepatic Cholangiocarcinoma.

KRAS mutations are causally linked to protumor inflammation and are identified as driving factors in tumorigenesis. Here, using multiomics data gathered from a large set of patients, we showed that KRAS mutation was associated with a specific landscape of alternative mRNA splicing that connected to myeloid inflammation in intrahepatic cholangiocarcinoma (iCCA). Then, we identified a negative feedback mechanism in which the upregulation of interleukin 1 receptor antagonist (IL1RN)-201/203 due to alternative splicing confers vital anti-inflammatory effects in KRAS-mutant iCCA. In KRAS-mutant iCCA mice, both IL1RN-201/203 upregulation and anakinra treatment ignited a significant antitumor immune response by altering neutrophil recruitment and phenotypes. Furthermore, anakinra treatment synergistically enhanced anti-PD-1 therapy to activate intratumoral GZMB+ CD8+ T cells in KRAS-mutant iCCA mice. Clinically, we found that high IL1RN-201/203 levels in patients with KRAS-mutant iCCA were significantly associated with superior response to anti-PD-1 immunotherapy. SIGNIFICANCE: This work describes a novel inflammatory checkpoint mediated by IL1RN alternative splicing variants that may serve as a promising basis to develop therapeutic options for KRAS-mutant iCCA and other cancers. This article is featured in Selected Articles from This Issue, p. 2109.

Pharmaco-proteogenomic characterization of liver cancer organoids for precision oncology.

Organoid models have the potential to recapitulate the biological and pharmacotypic features of parental tumors. Nevertheless, integrative pharmaco-proteogenomics analysis for drug response features and biomarker investigation for precision therapy of patients with liver cancer are still lacking. We established a patient-derived liver cancer organoid biobank (LICOB) that comprehensively represents the histological and molecular characteristics of various liver cancer types as determined by multiomics profiling, including genomic, epigenomic, transcriptomic, and proteomic analysis. Proteogenomic profiling of LICOB identified proliferative and metabolic organoid subtypes linked to patient prognosis. High-throughput drug screening revealed distinct response patterns of each subtype that were associated with specific multiomics signatures. Through integrative analyses of LICOB pharmaco-proteogenomics data, we identified the molecular features associated with drug responses and predicted potential drug combinations for personalized patient treatment. The synergistic inhibition effect of mTOR inhibitor temsirolimus and the multitargeted tyrosine kinase inhibitor lenvatinib was validated in organoids and patient-derived xenografts models. We also provide a user-friendly web portal to help serve the biomedical research community. Our study is a rich resource for investigation of liver cancer biology and pharmacological dependencies and may help enable functional precision medicine.

Multidimensional immune profiling in Gastric Cancer Multiplex Immunohistochemistry Atlas from Peking University Cancer Hospital project informs PD-1/PD-L1 blockade efficacy.

BACKGROUND: Immunotherapy has resulted in impressive objective response rates and durable tumour remission, but only in a subset of gastric cancer (GC) patients. The PD-L1 combined positive score is the most widely used tissue-based biomarker for anti-PD-1/PD-L1 therapy; however, this unidimensional method has limitations. Next-generation exploration of tissue-based biomarkers for GC requires characterisation of various cellular markers and key immunoregulatory molecule expression in situ. Thus, a complete, stepwise solution covering the entire process from staining samples to cross-site utilisation of pathomics data is urgently needed. METHODS: With the advanced multispectral imaging analysis method, web-based data repository, and interactive sharing technology, we conducted a project entitled Gastric Cancer Multiplex Immunohistochemistry Atlas from Peking University Cancer Hospital (GMAP). We propose a standard pipeline covering sample collection, staining, scanning multispectral images, constructing a spectral library, identifying and phenotyping cells, positioning each element, and quantitatively extracting immune features. We designed an open-access relational database to explore tissue-based biomarkers to determine PD-1/PD-L1 blockade efficacy. RESULTS: The GMAP project detected the functional status and spatial location of more than 50 million cells using 15 markers in 80 GC patients, based on which billions of cell pairs were recognised, highlighting the rich spatial arrangement information and the fine tumour microenvironment structure. We generated a tumour-immune atlas using the count and spatial features of 65 immune cell types. We eventually selected the indicators and built a comprehensive risk-scoring system. Patients with higher risk score showed superior immunotherapy-related progression-free survival (irPFS) (hazard ratio [HR]: 3.19; P < 0.001; median irPFS: 4.87 versus 19.87months, respectively) and immunotherapy-related overall survival (HR: 3.10; P = 0.001; median irPFS: 10.03 versus 24.87months, respectively) compared with lower risk patients, demonstrating their potential for guiding anti-PD-1/PD-L1-based immunotherapy. Importantly, an easy-to-use and versatile web server was built to promote tissue-based biomarker exploration in GC. CONCLUSION: The GMAP project highlighted the clinical value of tissue-based immune features as biomarkers for immunotherapeutic decision-making. We present a well-designed, detailed workflow for the orderly generation and use of a high-quality, spatially resolved pathological database.

Lactylome analysis suggests lactylation-dependent mechanisms of metabolic adaptation in hepatocellular carcinoma.

Enhanced glycolysis and accumulation of lactate is a common feature in various types of cancer. Intracellular lactate drives a recently described type of posttranslational modification, lysine lactylation (Kla), on core histones. However, the impact of lactylation on biological processes of tumour cells remains largely unknown. Here we show a global lactylome profiling on a prospectively collected hepatitis B virus-related hepatocellular carcinoma (HCC) cohort. Integrative lactylome and proteome analysis of the tumours and adjacent livers identifies 9,275 Kla sites, with 9,256 sites on non-histone proteins, indicating that Kla is a prevalent modification beyond histone proteins and transcriptional regulation. Notably, Kla preferentially affects enzymes involved in metabolic pathways, including the tricarboxylic acid cycle, and carbohydrate, amino acid, fatty acid and nucleotide metabolism. We further verify that lactylation at K28 inhibits the function of adenylate kinase 2, facilitating the proliferation and metastasis of HCC cells. Our study therefore reveals that Kla plays an important role in regulating cellular metabolism and may contribute to HCC progression.

FUT8 is regulated by miR-122-5p and promotes malignancies in intrahepatic cholangiocarcinoma via PI3K/AKT signaling.

BACKGROUND: Intrahepatic cholangiocarcinoma (iCCA) is the second-most lethal primary liver cancer and its prognosis remains dismal. N-glycosylation, which is biosynthesized by a number of glycosyltransferases, plays an important role in a variety of biological processes and is associated with cancer development and progression. METHODS: Based on our previous proteogenomic resources from an iCCA cohort of 262 patients, fucosyltransferases 8 (FUT8) showed significant prognosis relevance in iCCA. Tumor tissues from iCCA patients were used to evaluate the correlation between its expression and clinical information. Gain/loss-of-function experiments in iCCA cell lines were performed to elucidate the biological function of FUT8. In addition, its downstream pathways and post-transcriptional regulators were inferred and verified. RESULTS: Elevated FUT8 expression was clinically associated with worse overall survival in iCCA patients. Its overexpression promoted migration, invasion and proliferation ability of iCCA cells. In addition, miR-122-5p was found to act as a post-transcriptional regulator of FUT8 and proved to inhibit FUT8 expression and then suppress the proliferation and migration ability of iCCA cell lines. Furthermore, FUT8 was observed to promote iCCA development through PI3K/AKT signaling pathway. CONCLUSIONS: These findings demonstrated that FUT8, regulated by miR-122-5p, could be a tumor promoter of iCCA through PI3K/AKT signaling pathway.

Multi-omics characterization reveals the pathogenesis of liver focal nodular hyperplasia.

The molecular landscape and pathogenesis of focal nodular hyperplasia (FNH) have yet to be elucidated. We performed multi-omics approaches on FNH and paired normal liver tissues from 22 patients, followed by multi-level bioinformatic analyses and experimental validations. Generally, FNH had low mutation burden with low variant allele frequencies, and the mutation frequency significantly correlated with proliferation rate. Although no recurrently deleterious genomic events were found, some putative tumor suppressors or oncogenes were involved. Mutational signatures indicated potential impaired mismatch function and possible poison contact. Integrated analyses unveiled a group of FNH specific endothelial cells that uniquely expressed SOST and probably had strong interaction with fibroblasts through PDGFB/PDGFRB pathway to promote fibrosis. Notably, in one atypical FNH (patient No.11) with pronounced copy number variations, we observed a unique immune module. Most FNH are benign, but molecularly atypical FNH still exist; endothelial cell derived PDGFB probably promotes the fibrogenic process in FNH.

Single-cell immune ecosystem and metabolism reprogramming imprinted by psoriasis niche.

Background: A major challenge of psoriasis is its dysfunctional immune niche. Remarkable gaps remain in understanding how immune cell state transitions are linked to clinical outcomes in psoriasis. Thus, there is a pressing need to discover immunomodulatory programs governing psoriasis progression. Methods: Here, by using the state-of-the-art single-cell RNA-sequencing (RNA-seq) data, we observed the unique immune cell profile inside the psoriasis niche compared with the normal skins. Results: In detail, the immunosuppressive T cells such as regulatory T (Treg) cells and CTLA4+ CD8 T cells showed higher infiltration in the psoriasis niche, indicating the immunosuppressive state was imprinted by such disease. Interestingly, unbiased trajectory and pathway enrichment analysis showed that those suppressive T cells potentially showed developmental and metabolic abnormalities. Intercellular crosstalk modeling shows that exhausted CTLA4+ CD8 T cells can send out cytokine signaling via utilizing CXCL13-CXCR3 ligand-receptor pair. We finally quantified the metabolism profile of T cells and strikingly observed their enhanced metabolic activity. Conclusions: Taken together, these data highlight cell-type specific reprogramming within the psoriasis microenvironment and provide evidence for immune-related biomarkers of psoriasis clinical outcome. Our work not only revealed the unique immune ecosystem of psoriasis, but also opened new opportunities for targeting immunometabolism in treating such skin diseases.

Single-cell transcriptomic analysis suggests two molecularly subtypes of intrahepatic cholangiocarcinoma.

Intrahepatic cholangiocarcinoma (iCCA) is a highly heterogeneous cancer with limited understanding of its classification and tumor microenvironment. Here, by performing single-cell RNA sequencing on 144,878 cells from 14 pairs of iCCA tumors and non-tumor liver tissues, we find that S100P and SPP1 are two markers for iCCA perihilar large duct type (iCCAphl) and peripheral small duct type (iCCApps). S100P + SPP1- iCCAphl has significantly reduced levels of infiltrating CD4+ T cells, CD56+ NK cells, and increased CCL18+ macrophages and PD1+CD8+ T cells compared to S100P-SPP1 + iCCApps. The transcription factor CREB3L1 is identified to regulate the S100P expression and promote tumor cell invasion. S100P-SPP1 + iCCApps has significantly more SPP1+ macrophage infiltration, less aggressiveness and better survival than S100P + SPP1- iCCAphl. Moreover, S100P-SPP1 + iCCApps harbors tumor cells at different status of differentiation, such as ALB + hepatocyte differentiation and ID3+ stemness. Our study extends the understanding of the diversity of tumor cells in iCCA.

Potential Biomarkers for Liver Cancer Diagnosis Based on Multi-Omics Strategy.

Liver cancer is the fourth leading cause of cancer-related death worldwide. Hepatocellular carcinoma (HCC) accounts for about 85%-90% of all primary liver malignancies. However, only 20-30% of HCC patients are eligible for curative therapy mainly due to the lack of early-detection strategies, highlighting the significance of reliable and accurate biomarkers. The integration of multi-omics became an important tool for biomarker screening and unique alterations in tumor-associated genes, transcripts, proteins, post-translational modifications and metabolites have been observed. We here summarized the novel biomarkers for HCC diagnosis based on multi-omics technology as well as the clinical significance of these potential biomarkers in the early detection of HCC.

Spatial omics: Navigating to the golden era of cancer research.

The idea that tumour microenvironment (TME) is organised in a spatial manner will not surprise many cancer biologists; however, systematically capturing spatial architecture of TME is still not possible until recent decade. The past five years have witnessed a boom in the research of high-throughput spatial techniques and algorithms to delineate TME at an unprecedented level. Here, we review the technological progress of spatial omics and how advanced computation methods boost multi-modal spatial data analysis. Then, we discussed the potential clinical translations of spatial omics research in precision oncology, and proposed a transfer of spatial ecological principles to cancer biology in spatial data interpretation. So far, spatial omics is placing us in the golden age of spatial cancer research. Further development and application of spatial omics may lead to a comprehensive decoding of the TME ecosystem and bring the current spatiotemporal molecular medical research into an entirely new paradigm.

Proteogenomic characterization identifies clinically relevant subgroups of intrahepatic cholangiocarcinoma.

We performed proteogenomic characterization of intrahepatic cholangiocarcinoma (iCCA) using paired tumor and adjacent liver tissues from 262 patients. Integrated proteogenomic analyses prioritized genetic aberrations and revealed hallmarks of iCCA pathogenesis. Aflatoxin signature was associated with tumor initiation, proliferation, and immune suppression. Mutation-associated signaling profiles revealed that TP53 and KRAS co-mutations may contribute to iCCA metastasis via the integrin-FAK-SRC pathway. FGFR2 fusions activated the Rho GTPase pathway and could be a potential source of neoantigens. Proteomic profiling identified four patient subgroups (S1-S4) with subgroup-specific biomarkers. These proteomic subgroups had distinct features in prognosis, genetic alterations, microenvironment dysregulation, tumor microbiota composition, and potential therapeutics. SLC16A3 and HKDC1 were further identified as potential prognostic biomarkers associated with metabolic reprogramming of iCCA cells. This study provides a valuable resource for researchers and clinicians to further identify molecular pathogenesis and therapeutic opportunities in iCCA.

Spatiotemporal Immune Landscape of Colorectal Cancer Liver Metastasis at Single-Cell Level.

Liver metastasis, the leading cause of colorectal cancer mortality, exhibits a highly heterogeneous and suppressive immune microenvironment. Here, we sequenced 97 matched samples by using single-cell RNA sequencing and spatial transcriptomics. Strikingly, the metastatic microenvironment underwent remarkable spatial reprogramming of immunosuppressive cells such as MRC1 + CCL18 + M2-like macrophages. We further developed scMetabolism, a computational pipeline for quantifying single-cell metabolism, and observed that those macrophages harbored enhanced metabolic activity. Interestingly, neoadjuvant chemotherapy could block this status and restore the antitumor immune balance in responsive patients, whereas the nonresponsive patients deteriorated into a more suppressive one. Our work described the immune evolution of metastasis and uncovered the black box of how tumors respond to neoadjuvant chemotherapy. SIGNIFICANCE: We present a single-cell and spatial atlas of colorectal liver metastasis and found the highly metabolically activated MRC1 + CCL18 + M2-like macrophages in metastatic sites. Efficient neoadjuvant chemotherapy can slow down such metabolic activation, raising the possibility to target metabolism pathways in metastasis.This article is highlighted in the In This Issue feature, p. 1.

The endoribonuclease N4BP1 prevents psoriasis by controlling both keratinocytes proliferation and neutrophil infiltration.

Psoriasis is a common chronic skin disease, characterized by abnormal interplay between hyperproliferative epidermal keratinocytes and self-reactive immune cells with not fully addressed molecular mechanism. N4BP1 (NEDD4-binding protein 1) is considered as an immune regulator for a long time but its physiological role is not determined yet. Here, we found that the expression of N4BP1 in skin was highest among all 54 tested tissues, and its expression was further upregulated in psoriatic skin. N4BP1-deficient mice exhibited normal grossly, but developed severe and prolonged IMQ-induced psoriasis-like disease comparing to controls. N4BP1 mainly expressed in keratinocytes and located on nucleus. Up- but not downregulated genes in N4BP1-deficient skin were specifically enriched in keratinocyte proliferation and differentiation. The proliferation of N4BP1-deficient primary keratinocytes was faster compared to that of controls. The upregulated genes upon ablation of N4BP1 were highly enriched in targets of AP-1 transcription factor. Knocking out N4BP1 resulted in upregulation of JunB and FosB, and conversely, overexpression of N4BP1 greatly reduced their expression. Furthermore, N4BP1 binds with JunB and FosB encoding mRNAs and greatly reduces their stability. In addition, with a high expression in neutrophils, N4BP1 limits survival of neutrophils in blood and infiltration of neutrophils in psoriatic skin by targeting CXCL1, CCL20, and S100A8. These findings demonstrate that N4BP1 controls the proper function of keratinocytes and neutrophils by negatively regulating JunB, FosB, and CXCL1, respectively, and that is critical for psoriasis prevention.

Wide-ranging analysis of survival-related alternative splicing events in invasive breast carcinoma.

Invasive breast carcinoma (BRCA) is a serious disease that threatens the survival time of those affected. Alternative splicing (AS) involved in BRCA pathogenesis may be a potential therapeutic target. However, to the best of our knowledge, a systematic analysis of survival-related alternative splicing events (SREs) has not yet been reported. The aim of the present study was to identify SREs and analyze their potential biological functions as BRCA prognostic biomarkers. An UpSet plot demonstrated AS global characteristics. Cox's proportional hazards regression model quantitatively demonstrated the prognostic relevance of AS events. Functional enrichment analysis investigated the potential pathways through which AS events affect BRCA progression. The receiver operating characteristic curve model determined the clinical significance of AS events represented using percent-spliced-in (PSI) values. The regulatory network of splicing factors (SFs) and AS events laid the foundation for studying the role of SFs in BRCA. The present study identified 1,215 SREs and their distribution characteristics, suggesting that AS events in exon skipping (ES) primarily exerted normal physiological functions, while AS events in alternative terminator sites had the most significant prognostic effect. The present study demonstrated that survival-associated genes are involved primarily in certain biological processes of ribosomal proteins. In the diagnostic model, the alternative acceptor site, alternative donor site, alternative promoter site and ES performed well. ELAVL4 was the key gene associated with prognosis and SREs. In conclusion, a number of AS events affect BRCA initiation, progression and prognosis. The PSI value of AS events has the potential to diagnose BRCA and predict a prognosis; however, this must be confirmed in additional studies.

Identification of prognostic risk factors for pancreatic cancer using bioinformatics analysis.

BACKGROUND: Pancreatic cancer is one of the most common malignant cancers worldwide. Currently, the pathogenesis of pancreatic cancer remains unclear; thus, it is necessary to explore its precise molecular mechanisms. METHODS: To identify candidate genes involved in the tumorigenesis and proliferation of pancreatic cancer, the microarray datasets GSE32676, GSE15471 and GSE71989 were downloaded from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) between Pancreatic ductal adenocarcinoma (PDAC) and nonmalignant samples were screened by GEO2R. The Database for Annotation Visualization and Integrated Discovery (DAVID) online tool was used to obtain a synthetic set of functional annotation information for the DEGs. A PPI network of the DEGs was established using the Search Tool for the Retrieval of Interacting Genes (STRING) database, and a combination of more than 0.4 was considered statistically significant for the PPI. Subsequently, we visualized the PPI network using Cytoscape. Functional module analysis was then performed using Molecular Complex Detection (MCODE). Genes with a degree ≥10 were chosen as hub genes, and pathways of the hub genes were visualized using ClueGO and CluePedia. Additionally, GenCLiP 2.0 was used to explore interactions of hub genes. The Literature Mining Gene Networks module was applied to explore the cocitation of hub genes. The Cytoscape plugin iRegulon was employed to analyze transcription factors regulating the hub genes. Furthermore, the expression levels of the 13 hub genes in pancreatic cancer tissues and normal samples were validated using the Gene Expression Profiling Interactive Analysis (GEPIA) platform. Moreover, overall survival and disease-free survival analyses according to the expression of hub genes were performed using Kaplan-Meier curve analysis in the cBioPortal online platform. The relationship between expression level and tumor grade was analyzed using the online database Oncomine. Lastly, the eight snap-frozen tumorous and adjacent noncancerous adjacent tissues of pancreatic cancer patients used to detect the CDK1 and CEP55 protein levels by western blot. CONCLUSIONS: Altogether, the DEGs and hub genes identified in this work can help uncover the molecular mechanisms underlying the tumorigenesis of pancreatic cancer and provide potential targets for the diagnosis and treatment of this disease.