Tumor cell plasticity contributes to intratumoral heterogeneity and therapy resistance. Through cell plasticity, lung adenocarcinoma (LUAD) cells transform into neuroendocrinal (NE) cell-like tumor cells. However, the mechanisms of NE cell plasticity remain unclear. CRACD, a capping protein inhibitor, is frequently inactivated in cancers. CRACD knock-out (KO) de-represses NE-related gene expression in the pulmonary epithelium and LUAD cells. In LUAD mouse models, Cracd KO increases intratumoral heterogeneity with NE gene expression. Single-cell transcriptomic analysis showed that Cracd KO-induced NE plasticity is associated with cell de-differentiation and activated stemness-related pathways. The single-cell transcriptomes of LUAD patient tumors recapitulate that the distinct LUAD NE cell cluster expressing NE genes is co-enriched with SOX2, OCT4, and NANOG pathway activation, and impaired actin remodeling. This study reveals an unexpected role of CRACD in restricting NE cell plasticity that induces cell de-differentiation, which provides new insights into cell plasticity of LUAD.

Spatiotemporal control of stem and progenitor cells is essential for lung regeneration, the failure of which leads to lung disease. However, the mechanism of alveolar cell plasticity during regeneration remains elusive. We previously found that PCLAF remodels the DREAM complex for cell cycle re-entry. PCLAF expression is specifically enriched in proliferating lung progenitor cells, along with the DREAM target genes by lung damage. Genetic ablation of Pclaf inhibited alveolar type I (AT1) cell regeneration from alveolar type II (AT2) cells, inducing lung fibrosis. Mechanistically, the PCLAF-DREAM complex directly transactivates CLIC4, promoting TGF-beta signaling that regulates the balance between AT1 and AT2 cells. Furthermore, a drug candidate that mimics the PCLAF-DREAM transcriptional signatures for lung regeneration was identified and validated in organoids and mice. Our study unveils an unexpected role of the PCLAF-DREAM axis in controlling alveolar cell plasticity for lung regeneration and proposes a viable option for lung fibrosis prevention.

Despite the promising outcome of immune checkpoint blockade (ICB), ICB resistance is a new challenge. Thus, selecting patients for specific ICB applications is crucial for maximizing therapeutic efficacy. Herein we curated 69 human esophageal squamous cell cancer (ESCC) patients’ tumor microenvironment (TME) single-cell transcriptomic datasets for ESCC subtyping. Notably, integrative analyses of the cellular network transcriptional signatures of T cells, myeloid cells, and fibroblasts define distinct ESCC subtypes characterized by T cell exhaustion, Interferon alpha and beta signaling, TIGIT enrichment, and specific marker genes. Furthermore, this approach classifies ESCC patients into ICB responders and non-responders, validated by liquid biopsy single-cell transcriptomics. This study stratifies ESCC patients by TME transcriptional network, which provides a novel insight into tumor niche remodeling and helps predict ICB responses of ESCC patients.

Diffuse-type gastric adenocarcinoma (DGAC) is a deadly cancer often diagnosed late and resistant to treatment. While hereditary DGAC is linked to CDH1 mutations, the role of CDH1/E-cadherin inactivation in sporadic DGAC tumorigenesis remains elusive. We discovered CDH1 inactivation in a subset of DGAC patient tumors. Analyzing single-cell transcriptomes in malignant ascites, we identified two DGAC subtypes: DGAC1 (CDH1 loss) and DGAC2 (lacking immune response). DGAC1 displayed distinct molecular signatures, activated DGAC-related pathways, and an abundance of exhausted T cells in ascites. Genetically engineered murine gastric organoids showed that Cdh1 knock-out (KO), KrasG12D, Trp53 KO (EKP) accelerates tumorigenesis with immune evasion compared to KrasG12D, Trp53 KO (KP). We also identified EZH2 as a key mediator promoting CDH1 loss-associated DGAC tumorigenesis. These findings highlight DGAC's molecular diversity and potential for personalized treatment in CDH1-inactivated patients.

Background and aims: Despite recent progress in identifying aberrant genetic and epigenetic alterations in esophageal squamous cell carcinoma (ESCC), the mechanism of ESCC initiation remains unknown. Methods: Using CRISPR/Cas 9-based genetic ablation, we targeted 9 genes (TP53, CDKN2A, NOTCH1, NOTCH3, KMT2D, KMT2C, FAT1, FAT4, and AJUBA) in murine esophageal organoids (EOs). Transcriptomic phenotypes of organoids and chemokine released by organoids were analyzed by single-cell RNA sequencing (scRNA-seq). Tumorigenicity of organoids and tumor-infiltrated immune cells were monitored by allograft transplantation. Human ESCC scRNA-seq datasets were analyzed to classify patients and find subsets relevant to organoid models and immune evasion. Results: We established 32 genetically engineered EOs and identified key genetic determinants that drive ESCC initiation. A single-cell transcriptomic analysis uncovered that Trp53, Cdkn2a, and Notch1 (PCN) triple knockout (KO) induces neoplastic features of ESCC by generating cell lineage heterogeneity and high cell plasticity. PCN KO also generates immunosuppressive niche enriched with exhausted T cells and M2 macrophages via the CCL2-CCR2 axis. Mechanistically, CDKN2A inactivation transactivates CCL2 via NF-B. Moreover, comparative single-cell transcriptomic analyses stratified ESCC patients and identified a specific subtype recapitulating the PCN-type ESCC signatures, including the high expression of CCL2 and CD274/PD-L1. Conclusions: Our study unveils that loss of TP53, CDKN2A, and NOTCH1 induces esophageal neoplasia and immune evasion for ESCC initiation and proposes the CCL2 blockade as a viable approach to target PCN-type ESCC.