An atlas of epithelial cell states and plasticity in lung adenocarcinoma.

Understanding the cellular processes that underlie early lung adenocarcinoma (LUAD) development is needed to devise intervention strategies1. Here we studied 246,102 single epithelial cells from 16 early-stage LUADs and 47 matched normal lung samples. Epithelial cells comprised diverse normal and cancer cell states, and diversity among cancer cells was strongly linked to LUAD-specific oncogenic drivers. KRAS mutant cancer cells showed distinct transcriptional features, reduced differentiation and low levels of aneuploidy. Non-malignant areas surrounding human LUAD samples were enriched with alveolar intermediate cells that displayed elevated KRT8 expression (termed KRT8+ alveolar intermediate cells (KACs) here), reduced differentiation, increased plasticity and driver KRAS mutations. Expression profiles of KACs were enriched in lung precancer cells and in LUAD cells and signified poor survival. In mice exposed to tobacco carcinogen, KACs emerged before lung tumours and persisted for months after cessation of carcinogen exposure. Moreover, they acquired Kras mutations and conveyed sensitivity to targeted KRAS inhibition in KAC-enriched organoids derived from alveolar type 2 (AT2) cells. Last, lineage-labelling of AT2 cells or KRT8+ cells following carcinogen exposure showed that KACs are possible intermediates in AT2-to-tumour cell transformation. This study provides new insights into epithelial cell states at the root of LUAD development, and such states could harbour potential targets for prevention or intervention.

Atlas of Metastatic Gastric Cancer Links Ferroptosis to Disease Progression and Immunotherapy Response.

BACKGROUND & AIMS: Metastases from gastric adenocarcinoma (GAC) lead to high morbidity and mortality. Developing innovative and effective therapies requires a comprehensive understanding of the tumor and immune biology of advanced GAC. Yet, collecting matched specimens from advanced, treatment-naïve patients with GAC poses a significant challenge, limiting the scope of current research, which has focused predominantly on localized tumors. This gap hinders deeper insight into the metastatic dynamics of GAC. METHODS: We performed in-depth single-cell transcriptome and immune profiling on 68 paired, treatment-naïve, primary metastatic tumors to delineate alterations in cancer cells and their tumor microenvironment during metastatic progression. To validate our observations, we conducted comprehensive functional studies both in vitro and in vivo, using cell lines and multiple patient-derived xenograft and novel mouse models of GAC. RESULTS: Liver and peritoneal metastases exhibited distinct properties in cancer cells and dynamics of tumor microenvironment phenotypes, supporting the notion that cancer cells and their local tumor microenvironments co-evolve at metastatic sites. Our study also revealed differential activation of cancer meta-programs across metastases. We observed evasion of cancer cell ferroptosis via GPX4 up-regulation during GAC progression. Conditional depletion of Gpx4 or pharmacologic inhibition of ferroptosis resistance significantly attenuated tumor growth and metastatic progression. In addition, ferroptosis-resensitizing treatments augmented the efficacy of chimeric antigen receptor T-cell therapy. CONCLUSIONS: This study represents the largest single-cell dataset of metastatic GACs to date. High-resolution mapping of the molecular and cellular dynamics of GAC metastasis has revealed a rationale for targeting ferroptosis defense in combination with chimeric antigen receptor T-cell therapy as a novel therapeutic strategy with potential immense clinical implications.

TIGIT is the central player in T-cell suppression associated with CAR T-cell relapse in mantle cell lymphoma.

BACKGROUND: Chimeric antigen receptor (CAR) T-cell therapy using brexucabtagene autoleucel (BA) induces remission in many patients with mantle cell lymphoma (MCL), and BA is the only CAR T-cell therapy approved by the FDA for MCL. However, development of relapses to BA is recognized with poor patient outcomes. Multiple CAR T-cell therapies have been approved for other lymphomas and the resistance mechanisms have been investigated. However, the mechanisms underlying BA relapse in MCL have not been investigated and whether any previously reported resistance mechanisms apply to BA-relapsed patients with MCL is unknown. METHODS: To interrogate BA resistance mechanisms in MCL, we performed single-cell RNA sequencing on 39 longitudinally collected samples from 15 BA-treated patients, and multiplex cytokine profiling on 80 serial samples from 20 patients. RESULTS: We demonstrate that after BA relapse, the proportion of T cells, especially cytotoxic T cells (CTLs), decreased among non-tumor cells, while the proportion of myeloid cells correspondingly increased. TIGIT, LAG3, and CD96 were the predominant checkpoint molecules expressed on exhausted T cells and CTLs; only TIGIT was significantly increased after relapse. CTLs expanded during remission, and then contracted during relapse with upregulated TIGIT expression. Tumor cells also acquired TIGIT expression after relapse, leading to the enhanced interaction of tumor cell TIGIT with monocyte CD155/PVR. In myeloid cells, post-relapse HLA-II expression was reduced relative to pretreatment and during remission. Myeloid-derived suppressor cells (MDSCs) were enriched after relapse with elevated expression of activation markers, including CLU (clusterin) and VCAN (versican). Extracellular chemokines (CCL4, CXCL9, CXCL13), soluble checkpoint inhibitors (sPD-L1, sTIM3, s4-1BB), and soluble receptors (sIL-2R, sTNFRII) were decreased during remission but elevated after relapse. CONCLUSIONS: Our data demonstrate that multiple tumor-intrinsic and -extrinsic factors are associated with T-cell suppression and BA relapse. Among these, TIGIT appears to be the central player given its elevated expression after BA relapse in not only CTLs but also MCL cells. The acquisition of TIGIT expression on tumor cells is MCL-specific and has not been reported in other CAR T-treated diseases. Together, our data suggest that co-targeting TIGIT may prevent CAR T relapses and thus promote long-term progression-free survival in MCL patients.

In silico drug repositioning based on drug-miRNA associations.

Drug repositioning has become a prevailing tactic as this strategy is efficient, economical and low risk for drug discovery. Meanwhile, recent studies have confirmed that small-molecule drugs can modulate the expression of disease-related miRNAs, which indicates that miRNAs are promising therapeutic targets for complex diseases. In this study, we put forward and verified the hypothesis that drugs with similar miRNA profiles may share similar therapeutic properties. Furthermore, a comprehensive drug-drug interaction network was constructed based on curated drug-miRNA associations. Through random network comparison, topological structure analysis and network module extraction, we found that the closely linked drugs in the network tend to treat the same diseases. Additionally, the curated drug-disease relationships (from the CTD) and random walk with restarts algorithm were utilized on the drug-drug interaction network to identify the potential drugs for a given disease. Both internal validation (leave-one-out cross-validation) and external validation (independent drug-disease data set from the ChEMBL) demonstrated the effectiveness of the proposed approach. Finally, by integrating drug-miRNA and miRNA-disease information, we also explain the modes of action of drugs in the view of miRNA regulation. In summary, our work could determine novel and credible drug indications and offer novel insights and valuable perspectives for drug repositioning.

Pathway enrichment analysis approach based on topological structure and updated annotation of pathway.

Pathway enrichment analysis has been widely used to identify cancer risk pathways, and contributes to elucidating the mechanism of tumorigenesis. However, most of the existing approaches use the outdated pathway information and neglect the complex gene interactions in pathway. Here, we first reviewed the existing widely used pathway enrichment analysis approaches briefly, and then, we proposed a novel topology-based pathway enrichment analysis (TPEA) method, which integrated topological properties and global upstream/downstream positions of genes in pathways. We compared TPEA with four widely used pathway enrichment analysis tools, including database for annotation, visualization and integrated discovery (DAVID), gene set enrichment analysis (GSEA), centrality-based pathway enrichment (CePa) and signaling pathway impact analysis (SPIA), through analyzing six gene expression profiles of three tumor types (colorectal cancer, thyroid cancer and endometrial cancer). As a result, we identified several well-known cancer risk pathways that could not be obtained by the existing tools, and the results of TPEA were more stable than that of the other tools in analyzing different data sets of the same cancer. Ultimately, we developed an R package to implement TPEA, which could online update KEGG pathway information and is available at the Comprehensive R Archive Network (CRAN): https://cran.r-project.org/web/packages/TPEA/.

Accurate prediction and elucidation of drug resistance based on the robust and reproducible chemoresponse communities.

Selecting the available treatment for each cancer patient from genomic context is a core goal of precision medicine, but innovative approaches with mechanism interpretation and improved performance are still highly needed. Through utilizing in vitro chemotherapy response data coupled with gene and miRNA expression profiles, we applied a network-based approach that identified markers not as individual molecules but as functional groups extracted from the integrated transcription factor and miRNA regulatory network. Based on the identified chemoresponse communities, the predictors of drug resistance achieved high accuracy in cross-validation and were more robust and reproducible than conventional single-molecule markers. Meanwhile, as candidate communities not only enriched abundant cellular process but also covered a variety of drug enzymes, transporters, and targets, these resulting chemoresponse communities furnished novel models to interpret multiple kinds of potential regulatory mechanism, such as dysregulation of cancer cell apoptosis or disturbance of drug metabolism. Moreover, compounds were linked based on the enrichment of their common chemoresponse communities to uncover undetected response patterns and possible multidrug resistance phenotype. Finally, an omnibus repository named ChemoCommunity (http://www.jianglab.cn/ChemoCommunity/) was constructed, which furnished a user-friendly interface for a convenient retrieval of the detailed information on chemoresponse communities. Taken together, our method, and the accompanying database, improved the performance of classifiers for drug resistance and provided novel model to uncover the possible regulatory mechanism of individual response to drug.

ncDR: a comprehensive resource of non-coding RNAs involved in drug resistance.

SUMMARY: As a promising field of individualized therapy, non-coding RNA pharmacogenomics promotes the understanding of different individual responses to certain drugs and acts as a reasonable reference for clinical treatment. However, relevant information is scattered across the published literature, which is inconvenient for researchers to explore non-coding RNAs that are involved in drug resistance. To address this, we systemically identified validated and predicted drug resistance-associated microRNAs and long non-coding RNAs through manual curation and computational analysis. Subsequently, we constructed an omnibus repository named ncDR, which furnishes a user-friendly interface that allows for convenient browsing, visualization, querying and downloading of data. Given the rapidly increasing interest in precision medicine, ncDR will significantly improve our understanding of the roles of regulatory non-coding RNAs in drug resistance and has the potential to be a timely and valuable resource. AVAILABILITY AND IMPLEMENTATION: http://www.jianglab.cn/ncDR/. CONTACT: jiangwei@hrbmu.edu.cn or lw2247@yeah.net. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Identification of active transcription factor and miRNA regulatory pathways in Alzheimer's disease.

MOTIVATION: Alzheimer's disease (AD) is a severe neurodegenerative disease of the central nervous system that may be caused by perturbation of regulatory pathways rather than the dysfunction of a single gene. However, the pathology of AD has yet to be fully elucidated. RESULTS: In this study, we systematically analyzed AD-related mRNA and miRNA expression profiles as well as curated transcription factor (TF) and miRNA regulation to identify active TF and miRNA regulatory pathways in AD. By mapping differentially expressed genes and miRNAs to the curated TF and miRNA regulatory network as active seed nodes, we obtained a potential active subnetwork in AD. Next, by using the breadth-first-search technique, potential active regulatory pathways, which are the regulatory cascade of TFs, miRNAs and their target genes, were identified. Finally, based on the known AD-related genes and miRNAs, the hypergeometric test was used to identify active pathways in AD. As a result, nine pathways were found to be significantly activated in AD. A comprehensive literature review revealed that eight out of nine genes and miRNAs in these active pathways were associated with AD. In addition, we inferred that the pathway hsa-miR-146a→STAT1→MYC, which is the source of all nine significantly active pathways, may play an important role in AD progression, which should be further validated by biological experiments. Thus, this study provides an effective approach to finding active TF and miRNA regulatory pathways in AD and can be easily applied to other complex diseases.

SM2miR: a database of the experimentally validated small molecules' effects on microRNA expression.

UNLABELLED: The inappropriate expression of microRNAs (miRNAs) is closely related with disease diagnosis, prognosis and therapy response. Recently, many studies have demonstrated that bioactive small molecules (or drugs) can regulate miRNA expression, which indicates that targeting miRNAs with small molecules is a new therapy for human diseases. In this study, we established the SM2miR database, which recorded 2925 relationships between 151 small molecules and 747 miRNAs in 17 species after manual curation from nearly 2000 articles. Each entry contains the detailed information about small molecules, miRNAs and evidences of their relationships, such as species, miRBase Accession number, DrugBank Accession number, PubChem Compound Identifier (CID), expression pattern of miRNA, experimental method, tissues or conditions for detection. SM2miR database has a user-friendly interface to retrieve by miRNA or small molecule. In addition, we offered a submission page. Thus, SM2miR provides a fairly comprehensive repository about the influences of small molecules on miRNA expression, which will promote the development of miRNA therapeutics. AVAILABILITY: SM2miR is freely available at http://bioinfo.hrbmu.edu.cn/SM2miR/.

Oropharyngeal microbiome profiling and its association with age and heart failure in the elderly population from the northernmost province of China.

Respiratory tract infections are the most common triggers for heart failure in elderly people. The healthy respiratory commensal microbiota can prevent invasion by infectious pathogens and decrease the risk of respiratory tract infections. However, upper respiratory tract (URT) microbiome in the elderly is not well understood. To comprehend the profiles of URT microbiota in the elderly, and the link between the microbiome and heart failure, we investigated the oropharyngeal (OP) microbiome of these populations in Heilongjiang Province, located in the North-East of China, a high-latitude and cold area with a high prevalence of respiratory tract infection and heart failure. Taxonomy-based analysis showed that six dominant phyla were represented in the OP microbial profiles. Compared with young adults, the OP in the elderly exhibited a significantly different microbial community, mainly characterized by highly prevalent Streptococcus, unidentified_Saccharibacteria, Veillonella, unidentified_Pre votellaceae, and Neisseria. While unidentified_Prevotellaceae dominated in the young OP microbiome. There was competition for niche dominance between Streptococcus and member of Prevotellaceae in the OP. Correlation analysis revealed that the abundance of unidentified_Saccharibacteria was positive, while Streptococcus was negatively correlated to age among healthy elderly. The bacterial structure and abundance in the elderly with heart failure were much like healthy controls. Certain changes in microbial diversity indicated the potential OP microbial disorder in heart failure patients. These results presented here identify the respiratory tract core microbiota in high latitude and cold regions, and reveal the robustness of OP microbiome in the aged, supplying the basis for microbiome-targeted interventions.IMPORTANCETo date, we still lack available data on the oropharyngeal (OP) microbial communities in healthy populations, especially the elderly, in high latitude and cold regions. A better understanding of the significantly changed respiratory tract microbiota in aging can provide greater insight into characteristics of longevity and age-related diseases. In addition, determining the relationship between heart failure and OP microbiome may provide novel prevention and therapeutic strategies. Here, we compared OP microbiome in different age groups and elderly people with or without heart failure in northeastern China. We found that OP microbial communities are strongly linked to healthy aging. And the disease status of heart failure was not a powerful factor affecting OP microbiome. The findings may provide basic data to reveal respiratory bacterial signatures of individuals in a cold geographic region.

Blocking Ubiquitin-Specific Protease 7 Induces Ferroptosis in Gastric Cancer via Targeting Stearoyl-CoA Desaturase.

Gastric cancer (GC) presents a formidable global health challenge, and conventional therapies face efficacy limitations. Ubiquitin-specific protease 7 (USP7) plays pivotal roles in GC development, immune response, and chemo-resistance, making it a promising target. Various USP7 inhibitors have shown selectivity and efficacy in preclinical studies. However, the mechanistic role of USP7 has not been fully elucidated, and currently, no USP7 inhibitors have been approved for clinical use. In this study, DHPO is identified as a potent USP7 inhibitor for GC treatment through in silico screening. DHPO demonstrates significant anti-tumor activity in vitro, inhibiting cell viability and clonogenic ability, and preventing tumor migration and invasion. In vivo studies using orthotopic gastric tumor mouse models validate DHPO's efficacy in suppressing tumor growth and metastasis without significant toxicity. Mechanistically, DHPO inhibition triggers ferroptosis, evidenced by mitochondrial alterations, lipid Reactive Oxygen Species (ROS), Malondialdehyde (MDA) accumulation, and iron overload. Further investigations unveil USP7's regulation of Stearoyl-CoA Desaturase (SCD) through deubiquitination, linking USP7 inhibition to SCD degradation and ferroptosis induction. Overall, this study identifies USP7 as a key player in ferroptosis of GC, elucidates DHPO's inhibitory mechanisms, and highlights its potential for GC treatment by inducing ferroptosis through SCD regulation.

METI: deep profiling of tumor ecosystems by integrating cell morphology and spatial transcriptomics.

Recent advances in spatial transcriptomics (ST) techniques provide valuable insights into cellular interactions within the tumor microenvironment (TME). However, most analytical tools lack consideration of histological features and rely on matched single-cell RNA sequencing data, limiting their effectiveness in TME studies. To address this, we introduce the Morphology-Enhanced Spatial Transcriptome Analysis Integrator (METI), an end-to-end framework that maps cancer cells and TME components, stratifies cell types and states, and analyzes cell co-localization. By integrating spatial transcriptomics, cell morphology, and curated gene signatures, METI enhances our understanding of the molecular landscape and cellular interactions within the tissue. We evaluate the performance of METI on ST data generated from various tumor tissues, including gastric, lung, and bladder cancers, as well as premalignant tissues. We also conduct a quantitative comparison of METI with existing clustering and cell deconvolution tools, demonstrating METI's robust and consistent performance.

Evolution of immune and stromal cell states and ecotypes during gastric adenocarcinoma progression.

Understanding tumor microenvironment (TME) reprogramming in gastric adenocarcinoma (GAC) progression may uncover novel therapeutic targets. Here, we performed single-cell profiling of precancerous lesions, localized and metastatic GACs, identifying alterations in TME cell states and compositions as GAC progresses. Abundant IgA+ plasma cells exist in the premalignant microenvironment, whereas immunosuppressive myeloid and stromal subsets dominate late-stage GACs. We identified six TME ecotypes (EC1-6). EC1 is exclusive to blood, while EC4, EC5, and EC2 are highly enriched in uninvolved tissues, premalignant lesions, and metastases, respectively. EC3 and EC6, two distinct ecotypes in primary GACs, associate with histopathological and genomic characteristics, and survival outcomes. Extensive stromal remodeling occurs in GAC progression. High SDC2 expression in cancer-associated fibroblasts (CAFs) is linked to aggressive phenotypes and poor survival, and SDC2 overexpression in CAFs contributes to tumor growth. Our study provides a high-resolution GAC TME atlas and underscores potential targets for further investigation.

Pan-cancer T cell atlas links a cellular stress response state to immunotherapy resistance.

Tumor-infiltrating T cells offer a promising avenue for cancer treatment, yet their states remain to be fully characterized. Here we present a single-cell atlas of T cells from 308,048 transcriptomes across 16 cancer types, uncovering previously undescribed T cell states and heterogeneous subpopulations of follicular helper, regulatory and proliferative T cells. We identified a unique stress response state, TSTR, characterized by heat shock gene expression. TSTR cells are detectable in situ in the tumor microenvironment across various cancer types, mostly within lymphocyte aggregates or potential tertiary lymphoid structures in tumor beds or surrounding tumor edges. T cell states/compositions correlated with genomic, pathological and clinical features in 375 patients from 23 cohorts, including 171 patients who received immune checkpoint blockade therapy. We also found significantly upregulated heat shock gene expression in intratumoral CD4/CD8+ cells following immune checkpoint blockade treatment, particularly in nonresponsive tumors, suggesting a potential role of TSTR cells in immunotherapy resistance. Our well-annotated T cell reference maps, web portal and automatic alignment/annotation tool could provide valuable resources for T cell therapy optimization and biomarker discovery.

The Single-Cell Immunogenomic Landscape of B and Plasma Cells in Early-Stage Lung Adenocarcinoma.

Tumor-infiltrating B and plasma cells (TIB) are prevalent in lung adenocarcinoma (LUAD); however, they are poorly characterized. We performed paired single-cell RNA and B-cell receptor (BCR) sequencing of 16 early-stage LUADs and 47 matching multiregion normal tissues. By integrative analysis of ∼50,000 TIBs, we define 12 TIB subsets in the LUAD and adjacent normal ecosystems and demonstrate extensive remodeling of TIBs in LUADs. Memory B cells and plasma cells (PC) were highly enriched in tumor tissues with more differentiated states and increased frequencies of somatic hypermutation. Smokers exhibited markedly elevated PCs and PCs with distinct differentiation trajectories. BCR clonotype diversity increased but clonality decreased in LUADs, smokers, and with increasing pathologic stage. TIBs were mostly localized within CXCL13+ lymphoid aggregates, and immune cell sources of CXCL13 production evolved with LUAD progression and included elevated fractions of CD4 regulatory T cells. This study provides a spatial landscape of TIBs in early-stage LUAD. SIGNIFICANCE: While TIBs are highly enriched in LUADs, they are poorly characterized. This study provides a much-needed understanding of the transcriptional, clonotypic states and phenotypes of TIBs, unraveling their potential roles in the immunopathology of early-stage LUADs and constituting a road map for the development of TIB-targeted immunotherapies for the treatment of this morbid malignancy. This article is highlighted in the In This Issue feature, p. 2483.

Follicular Lymphoma Microenvironment Characteristics Associated with Tumor Cell Mutations and MHC Class II Expression.

Follicular lymphoma (FL) is a B-cell malignancy with a complex tumor microenvironment that is rich in nonmalignant immune cells. We applied single-cell RNA sequencing to characterize the diverse tumor and immune cell populations of FL and identified major phenotypic subsets of FL T cells, including a cytotoxic CD4 T-cell population. We characterized four major FL subtypes with differential representation or relative depletion of distinct T-cell subsets. By integrating exome sequencing, we observed that somatic mutations are associated with, but not definitive for, reduced MHC expression on FL cells. In turn, expression of MHCII genes by FL cells was associated with significant differences in the proportions and targetable immunophenotypic characteristics of T cells. This provides a classification framework of the FL microenvironment in association with FL genotypes and MHC expression, and informs different potential immunotherapeutic strategies based upon tumor cell MHCII expression. SIGNIFICANCE: We have characterized the FL-infiltrating T cells, identified cytotoxic CD4 T cells as an important component that is associated with tumor cell-intrinsic characteristics, and identified sets of targetable immune checkpoints on T cells that differed from FLs with normal versus low MHC expression. See related commentary by Melnick, p. 374. This article is highlighted in the In This Issue feature, p. 369.

Longitudinal single-cell profiling reveals molecular heterogeneity and tumor-immune evolution in refractory mantle cell lymphoma.

The mechanisms driving therapeutic resistance and poor outcomes of mantle cell lymphoma (MCL) are incompletely understood. We characterize the cellular and molecular heterogeneity within and across patients and delineate the dynamic evolution of tumor and immune cell compartments at single cell resolution in longitudinal specimens from ibrutinib-sensitive patients and non-responders. Temporal activation of multiple cancer hallmark pathways and acquisition of 17q are observed in a refractory MCL. Multi-platform validation is performed at genomic and cellular levels in PDX models and larger patient cohorts. We demonstrate that due to 17q gain, BIRC5/survivin expression is upregulated in resistant MCL tumor cells and targeting BIRC5 results in marked tumor inhibition in preclinical models. In addition, we discover notable differences in the tumor microenvironment including progressive dampening of CD8+ T cells and aberrant cell-to-cell communication networks in refractory MCLs. This study reveals diverse and dynamic tumor and immune programs underlying therapy resistance in MCL.

9p21 loss confers a cold tumor immune microenvironment and primary resistance to immune checkpoint therapy.

Immune checkpoint therapy (ICT) provides substantial clinical benefits to cancer patients, but a large proportion of cancers do not respond to ICT. To date, the genomic underpinnings of primary resistance to ICT remain elusive. Here, we performed immunogenomic analysis of data from TCGA and clinical trials of anti-PD-1/PD-L1 therapy, with a particular focus on homozygous deletion of 9p21.3 (9p21 loss), one of the most frequent genomic defects occurring in ~13% of all cancers. We demonstrate that 9p21 loss confers "cold" tumor-immune phenotypes, characterized by reduced abundance of tumor-infiltrating leukocytes (TILs), particularly, T/B/NK cells, altered spatial TILs patterns, diminished immune cell trafficking/activation, decreased rate of PD-L1 positivity, along with activation of immunosuppressive signaling. Notably, patients with 9p21 loss exhibited significantly lower response rates to ICT and worse outcomes, which were corroborated in eight ICT trials of >1,000 patients. Further, 9p21 loss synergizes with PD-L1/TMB for patient stratification. A "response score" was derived by incorporating 9p21 loss, PD-L1 expression and TMB levels in pre-treatment tumors, which outperforms PD-L1, TMB, and their combination in identifying patients with high likelihood of achieving sustained response from otherwise non-responders. Moreover, we describe potential druggable targets in 9p21-loss tumors, which could be exploited to design rational therapeutic interventions.

Resolving the Spatial and Cellular Architecture of Lung Adenocarcinoma by Multiregion Single-Cell Sequencing.

Little is known of the geospatial architecture of individual cell populations in lung adenocarcinoma (LUAD) evolution. Here, we perform single-cell RNA sequencing of 186,916 cells from five early-stage LUADs and 14 multiregion normal lung tissues of defined spatial proximities from the tumors. We show that cellular lineages, states, and transcriptomic features geospatially evolve across normal regions to LUADs. LUADs also exhibit pronounced intratumor cell heterogeneity within single sites and transcriptional lineage-plasticity programs. T regulatory cell phenotypes are increased in normal tissues with proximity to LUAD, in contrast to diminished signatures and fractions of cytotoxic CD8+ T cells, antigen-presenting macrophages, and inflammatory dendritic cells. We further find that the LUAD ligand-receptor interactome harbors increased expression of epithelial CD24, which mediates protumor phenotypes. These data provide a spatial atlas of LUAD evolution, and a resource for identification of targets for its treatment. SIGNIFICANCE: The geospatial ecosystem of the peripheral lung and early-stage LUAD is not known. Our multiregion single-cell sequencing analyses unravel cell populations, states, and phenotypes in the spatial and ecologic evolution of LUAD from the lung that comprise high-potential targets for early interception.This article is highlighted in the In This Issue feature, p. 2355.