Lymphotoxin-ß promotes breast cancer bone metastasis colonization and osteolytic outgrowth.

Bone metastasis is a lethal consequence of breast cancer. Here we used single-cell transcriptomics to investigate the molecular mechanisms underlying bone metastasis colonization-the rate-limiting step in the metastatic cascade. We identified that lymphotoxin-ß (LTß) is highly expressed in tumour cells within the bone microenvironment and this expression is associated with poor bone metastasis-free survival. LTß promotes tumour cell colonization and outgrowth in multiple breast cancer models. Mechanistically, tumour-derived LTß activates osteoblasts through nuclear factor-κB2 signalling to secrete CCL2/5, which facilitates tumour cell adhesion to osteoblasts and accelerates osteoclastogenesis, leading to bone metastasis progression. Blocking LTß signalling with a decoy receptor significantly suppressed bone metastasis in vivo, whereas clinical sample analysis revealed significantly higher LTß expression in bone metastases than in primary tumours. Our findings highlight LTß as a bone niche-induced factor that promotes tumour cell colonization and osteolytic outgrowth and underscore its potential as a therapeutic target for patients with bone metastatic disease.

Accurate Early Detection and EGFR Mutation Status Prediction of Lung Cancer Using Plasma cfDNA Coverage Patterns: A Proof-of-Concept Study.

Lung cancer is a major global health concern with a low survival rate, often due to late-stage diagnosis. Liquid biopsy offers a non-invasive approach to cancer detection and monitoring, utilizing various features of circulating cell-free DNA (cfDNA). In this study, we established two models based on cfDNA coverage patterns at the transcription start sites (TSSs) from 6X whole-genome sequencing: an Early Cancer Screening Model and an EGFR mutation status prediction model. The Early Cancer Screening Model showed encouraging prediction ability, especially for early-stage lung cancer. The EGFR mutation status prediction model exhibited high accuracy in distinguishing between EGFR-positive and wild-type cases. Additionally, cfDNA coverage patterns at TSSs also reflect gene expression patterns at the pathway level in lung cancer patients. These findings demonstrate the potential applications of cfDNA coverage patterns at TSSs in early cancer screening and in cancer subtyping.

HeteroTCR: A heterogeneous graph neural network-based method for predicting peptide-TCR interaction.

Identifying interactions between T-cell receptors (TCRs) and immunogenic peptides holds profound implications across diverse research domains and clinical scenarios. Unsupervised clustering models (UCMs) cannot predict peptide-TCR binding directly, while supervised predictive models (SPMs) often face challenges in identifying antigens previously unencountered by the immune system or possessing limited TCR binding repertoires. Therefore, we propose HeteroTCR, an SPM based on Heterogeneous Graph Neural Network (GNN), to accurately predict peptide-TCR binding probabilities. HeteroTCR captures within-type (TCR-TCR or peptide-peptide) similarity information and between-type (peptide-TCR) interaction insights for predictions on unseen peptides and TCRs, surpassing limitations of existing SPMs. Our evaluation shows HeteroTCR outperforms state-of-the-art models on independent datasets. Ablation studies and visual interpretation underscore the Heterogeneous GNN module's critical role in enhancing HeteroTCR's performance by capturing pivotal binding process features. We further demonstrate the robustness and reliability of HeteroTCR through validation using single-cell datasets, aligning with the expectation that pMHC-TCR complexes with higher predicted binding probabilities correspond to increased binding fractions.

VitTCR: A deep learning method for peptide recognition prediction.

This study introduces VitTCR, a predictive model based on the vision transformer (ViT) architecture, aimed at identifying interactions between T cell receptors (TCRs) and peptides, crucial for developing cancer immunotherapies and vaccines. VitTCR converts TCR-peptide interactions into numerical AtchleyMaps using Atchley factors for prediction, achieving AUROC (0.6485) and AUPR (0.6295) values. Benchmark analysis indicates VitTCR's performance is comparable to other models, with further comparative studies suggested to understand its effectiveness in varied contexts. Additionally, integrating a positional bias weight matrix (PBWM), derived from amino acid contact probabilities in structurally resolved pMHC-TCR complexes, slightly improves VitTCR's accuracy. The model's predictions show weak yet statistically significant correlations with immunological factors like T cell clonal expansion and activation percentages, underscoring the biological relevance of VitTCR's predictive capabilities. VitTCR emerges as a valuable computational tool for predicting TCR-peptide interactions, offering insights for immunotherapy and vaccine development.

Dysregulation of Wnt/ß-catenin signaling contributes to intestinal inflammation through regulation of group 3 innate lymphoid cells.

RORγt+ group 3 innate lymphoid cells (ILC3s) are essential for intestinal homeostasis. Dysregulation of ILC3s has been found in the gut of patients with inflammatory bowel disease and colorectal cancer, yet the specific mechanisms still require more investigation. Here we observe increased ß-catenin in intestinal ILC3s from inflammatory bowel disease and colon cancer patients compared with healthy donors. In contrast to promoting RORγt expression in T cells, activation of Wnt/ß-catenin signaling in ILC3s suppresses RORγt expression, inhibits its proliferation and function, and leads to a deficiency of ILC3s and subsequent intestinal inflammation in mice. Activated ß-catenin and its interacting transcription factor, TCF-1, cannot directly suppress RORγt expression, but rather alters global chromatin accessibility and inhibits JunB expression, which is essential for RORγt expression in ILC3s. Together, our findings suggest that dysregulated Wnt/ß-catenin signaling impairs intestinal ILC3s through TCF-1/JunB/RORγt regulation, further disrupting intestinal homeostasis, and promoting inflammation and cancer.

Glioma-derived ANXA1 suppresses the immune response to TLR3 ligands by promoting an anti-inflammatory tumor microenvironment.

A highly immunosuppressive tumor microenvironment (TME) and the presence of the blood‒brain barrier are the two major obstacles to eliciting an effective immune response in patients with high-grade glioma (HGG). Here, we tried to enhance the local innate immune response in relapsed HGG by intracranially injecting poly(I:C) to establish a robust antitumor immune response in this registered clinical trial (NCT03392545). During the follow-up, 12/27 (44.4%) patients who achieved tumor control concomitant with survival benefit were regarded as responders in our study. We found that the T-cell receptor (TCR) repertoire in the TME was reshaped after poly(I:C) treatment. Based on the RNA-seq analysis of tumor samples, the expression of annexin A1 (ANXA1) was significantly upregulated in the tumor cells of nonresponders, which was further validated at the protein level. In vitro and in vivo experiments showed that ANXA1 could induce the production of M2-like macrophages and microglia via its surface receptor formyl peptide receptor 1 (FPR1) to establish a Treg cell-driven immunosuppressive TME and suppress the antitumor immune response facilitated by poly(I:C). The ANXA1/FPR1 signaling axis can inhibit the innate immune response of glioma patients by promoting an anti-inflammatory and Treg-driven TME. Moreover, ANXA1 could serve as a reliable predictor of response to poly(I:C), with a notable predictive accuracy rate of 92.3%. In light of these notable findings, this study unveils a new perspective of immunotherapy for gliomas.

Aberrant accumulation of Kras-dependent pervasive transcripts during tumor progression renders cancer cells dependent on PAF1 expression.

KRAS is the most commonly mutated oncogene in human cancer, and mutant KRAS is responsible for over 90% of pancreatic ductal adenocarcinoma (PDAC), the most lethal cancer. Here, we show that RNA polymerase II-associated factor 1 complex (PAF1C) is specifically required for survival of PDAC but not normal adult pancreatic cells. We show that PAF1C maintains cancer cell genomic stability by restraining overaccumulation of enhancer RNAs (eRNAs) and promoter upstream transcripts (PROMPTs) driven by mutant Kras. Loss of PAF1C leads to cancer-specific lengthening and accumulation of pervasive transcripts on chromatin and concomitant aberrant R-loop formation and DNA damage, which, in turn, trigger cell death. We go on to demonstrate that the global transcriptional hyperactivation driven by Kras signaling during tumorigenesis underlies the specific demand for PAF1C by cancer cells. Our work provides insights into how enhancer transcription hyperactivation causes general transcription factor addiction during tumorigenesis.

scRNA-seq of gastric tumor shows complex intercellular interaction with an alternative T cell exhaustion trajectory.

The tumor microenvironment (TME) in gastric cancer (GC) has been shown to be important for tumor control but the specific characteristics for GC are not fully appreciated. We generated an atlas of 166,533 cells from 10 GC patients with matched paratumor tissues and blood. Our results show tumor-associated stromal cells (TASCs) have upregulated activity of Wnt signaling and angiogenesis, and are negatively correlated with survival. Tumor-associated macrophages and LAMP3+ DCs are involved in mediating T cell activity and form intercellular interaction hubs with TASCs. Clonotype and trajectory analysis demonstrates that Tc17 (IL-17+CD8+ T cells) originate from tissue-resident memory T cells and can subsequently differentiate into exhausted T cells, suggesting an alternative pathway for T cell exhaustion. Our results indicate that IL17+ cells may promote tumor progression through IL17, IL22, and IL26 signaling, highlighting the possibility of targeting IL17+ cells and associated signaling pathways as a therapeutic strategy to treat GC.

Rapid screening of TCR-pMHC interactions by the YAMTAD system.

Personalized immunotherapy, such as cancer vaccine and TCR-T methods, demands rapid screening of TCR-pMHC interactions. While several screening approaches have been developed, their throughput is limited. Here, the Yeast Agglutination Mediated TCR antigen Discovery system (YAMTAD) was designed and demonstrated to allow fast and unbiased library-on-library screening of TCR-pMHC interactions. Our proof-of-principle study achieved high sensitivity and specificity in identifying antigens for a given TCR and identifying TCRs recognizing a given pMHC for modest library sizes. Finally, the enrichment of high-affinity TCR-pMHC interactions by YAMTAD in library-on-library screening was demonstrated. Given the high throughput (106-108 × 106-108 in theory) and simplicity (identifying TCR-pMHC interactions without purification of TCR and pMHC) of YAMTAD, this study provides a rapid but effective platform for TCR-pMHC interaction screening, with valuable applications in future personalized immunotherapy.

Tumor-derived Jagged1 promotes cancer progression through immune evasion.

Immune checkpoint inhibitor (ICI) therapy is generating remarkable responses in individuals with cancer, but only a small portion of individuals with breast cancer respond well. Here we report that tumor-derived Jagged1 is a key regulator of the tumor immune microenvironment. Jagged1 promotes tumorigenesis in multiple spontaneous mammary tumor models. Through Jagged1-induced Notch activation, tumor cells increase expression and secretion of multiple cytokines to help recruit macrophages into the tumor microenvironment. Educated macrophages crosstalk with tumor-infiltrating T cells to inhibit T cell proliferation and tumoricidal activity. In individuals with triple-negative breast cancer, a high expression level of Jagged1 correlates with increased macrophage infiltration and decreased T cell activity. Co-administration of an ICI PD-1 antibody with a Notch inhibitor significantly inhibits tumor growth in breast cancer models. Our findings establish a distinct signaling cascade by which Jagged1 promotes adaptive immune evasion of tumor cells and provide several possible therapeutic targets.

Knockout of immunotherapy prognostic marker genes eliminates the effect of the anti-PD-1 treatment.

The efficacy of immunotherapy is largely patient-specific due to heterogeneity in tumors. Combining statistic power from a variety of immunotherapies across cancer types, we found four biological pathways significantly correlated with patient survival following immunotherapy. The expression of immunotherapy prognostic marker genes (IPMGs) in these pathways can predict the patient survival with high accuracy not only in the TCGA cohort (89.36%) but also in two other independent cohorts (80.91%), highlighting that the activity of the IPMGs can reflect the sensitivity of the tumor immune microenvironment (TIME) to immunotherapies. Using mouse models, we show that knockout of one of the IPMGs, MALT1, which is critical for the T-cell receptor signaling, can eliminate the antitumor effect of anti-PD-1 treatment completely by impairing the activation of CD8+ T cells. Notably, knockout of another IPMG, CLEC4D, a C-type lectin receptor that expressed on myeloid cells, also reduced the effect of anti-PD-1 treatment potentially through maintaining the immunosuppressive effects of myeloid cells. Our results suggest that priming TIME via activating the IPMGs may increase the response rate and the effect of immune checkpoint blockers.

Mutant Kras co-opts a proto-oncogenic enhancer network in inflammation-induced metaplastic progenitor cells to initiate pancreatic cancer.

Kras-activating mutations display the highest incidence in pancreatic ductal adenocarcinoma. Pancreatic inflammation accelerates mutant Kras-driven tumorigenesis in mice, suggesting high selectivity in the cells that oncogenic Kras transforms, although the mechanisms dictating this specificity are poorly understood. Here we show that pancreatic inflammation is coupled to the emergence of a transient progenitor cell population that is readily transformed in the presence of mutant KrasG12D. These progenitors harbor a proto-oncogenic transcriptional program driven by a transient enhancer network. KrasG12D mutations lock this enhancer network in place, providing a sustained Kras-dependent oncogenic program that drives tumors throughout progression. Enhancer co-option occurs through functional interactions between the Kras-activated transcription factors Junb and Fosl1 and pancreatic lineage transcription factors, potentially accounting for inter-tissue specificity of oncogene transformation. The pancreatic ductal adenocarcinoma cell of origin thus provides an oncogenic transcriptional program that fuels tumor progression beyond initiation, accounting for the intra-tissue selectivity of Kras transformation.

Novel technologies in cfDNA analysis and potential utility in clinic.

The profiling of plasma cell-free DNA (cfDNA) is becoming a valuable tool rapidly for tumor diagnosis, monitoring and prognosis. Diverse plasma cfDNA technologies have been in routine or emerging use, including analyses of mutations, copy number alterations, gene fusions and DNA methylation. Recently, new technologies in cfDNA analysis have been developed in laboratories, and potentially reflect the status of epigenetic modification, the immune microenvironment and the microbiome in tumor tissues. In this review, the authors discuss the principles, methods and effects of the current cfDNA assays and provide an overview of studies that may inform clinical applications in the near future.

An immune-related gene signature for predicting survival and immunotherapy efficacy in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) ranks the fourth in terms of cancer-related mortality globally. Herein, in this research, we attempted to develop a novel immune-related gene signature that could predict survival and efficacy of immunotherapy for HCC patients. METHODS: The transcriptomic and clinical data of HCC samples were downloaded from The Cancer Genome Atlas (TCGA) and GSE14520 datasets, followed by acquiring immune-related genes from the ImmPort database. Afterwards, an immune-related gene-based prognostic index (IRGPI) was constructed using the Least Absolute Shrinkage and Selection Operator (LASSO) regression model. Kaplan-Meier survival curves as well as time-dependent receiver operating characteristic (ROC) curve were performed to evaluate its predictive capability. Besides, both univariate and multivariate analyses on overall survival for the IRGPI and multiple clinicopathologic factors were carried out, followed by the construction of a nomogram. Finally, we explored the possible correlation of IRGPI with immune cell infiltration or immunotherapy efficacy. RESULTS: Analysis of 365 HCC samples identified 11 differentially expressed immune-related genes, which were selected to establish the IRGPI. Notably, it can predict the survival of HCC patients more accurately than published biomarkers. Furthermore, IRGPI can predict the infiltration of immune cells in the tumor microenvironment of HCC, as well as the response of immunotherapy. CONCLUSION: Collectively, the currently established IRGPI can accurately predict survival, reflect the immune microenvironment, and predict the efficacy of immunotherapy among HCC patients.

Guizhi Fuling Decoction inhibiting the PI3K and MAPK pathways in breast cancer cells revealed by HTS2 technology and systems pharmacology.

As one of the classical traditional Chinese medicine (TCM) prescriptions in treating gynecological tumors, Guizhi Fuling Decoction (GFD) has been used to treat breast cancer (BRCA). Nonetheless, the potential molecular mechanism remains unclear so far. Therefore, systems pharmacology was used in combination with high throughput sequencing-based high throughput screening (HTS2) assay and bioinformatic technologies in this study to investigate the molecular mechanisms of GFD in treating BRCA. By computationally analyzing 76 active ingredients in GFD, 38 potential therapeutic targets were predicted and significantly enriched in the "pathways in cancer". Meanwhile, experimental analysis was carried out to examine changes in the expression levels of 308 genes involved in the "pathways in cancer" in BRCA cells treated by five herbs of GFD utilizing HTS2 platform, and 5 key therapeutic targets, including HRAS, EGFR, PTK2, SOS1, and ITGB1, were identified. The binding mode of active compounds to these five targets was analyzed by molecular docking and molecular dynamics simulation. It was found after integrating the computational and experimental data that, GFD possessed the anti-proliferation, pro-apoptosis, and anti-angiogenesis activities mainly through regulating the PI3K and the MAPK signaling pathways to inhibit BRCA. Besides, consistent with the TCM theory about the synergy of Cinnamomi Ramulus (Guizhi) by Cortex Moutan (Mudanpi) in GFD, both of these two herbs acted on the same targets and pathways. Taken together, the combined application of computational systems pharmacology techniques and experimental HTS2 platform provides a practical research strategy to investigate the functional and biological mechanisms of the complicated TCM prescriptions.

p53-mediated control of aspartate-asparagine homeostasis dictates LKB1 activity and modulates cell survival.

Asparagine synthetase (ASNS) catalyses the ATP-dependent conversion of aspartate to asparagine. However, both the regulation and biological functions of asparagine in tumour cells remain largely unknown. Here, we report that p53 suppresses asparagine synthesis through the transcriptional downregulation of ASNS expression and disrupts asparagine-aspartate homeostasis, leading to lymphoma and colon tumour growth inhibition in vivo and in vitro. Moreover, the removal of asparagine from culture medium or the inhibition of ASNS impairs cell proliferation and induces p53/p21-dependent senescence and cell cycle arrest. Mechanistically, asparagine and aspartate regulate AMPK-mediated p53 activation by physically binding to LKB1 and oppositely modulating LKB1 activity. Thus, we found that p53 regulates asparagine metabolism and dictates cell survival by generating an auto-amplification loop via asparagine-aspartate-mediated LKB1-AMPK signalling. Our findings highlight a role for LKB1 in sensing asparagine and aspartate and connect asparagine metabolism to the cellular signalling transduction network that modulates cell survival.

Chromatin-associated APC regulates gene expression in collaboration with canonical WNT signaling and AP-1.

Mutation of the APC gene occurs in a high percentage of colorectal tumors and is a central event driving tumor initiation in the large intestine. The APC protein performs multiple tumor suppressor functions including negative regulation of the canonical WNT signaling pathway by both cytoplasmic and nuclear mechanisms. Published reports that APC interacts with ß-catenin in the chromatin fraction to repress WNT-activated targets have raised the possibility that chromatin-associated APC participates more broadly in mechanisms of transcriptional control. This screening study has used chromatin immunoprecipitation and next-generation sequencing to identify APC-associated genomic regions in colon cancer cell lines. Initial target selection was performed by comparison and statistical analysis of 3,985 genomic regions associated with the APC protein to whole transcriptome sequencing data from APC-deficient and APC-wild-type colon cancer cells, and two types of murine colon adenomas characterized by activated Wnt signaling. 289 transcripts altered in expression following APC loss in human cells were linked to APC-associated genomic regions. High-confidence targets additionally validated in mouse adenomas included 16 increased and 9 decreased in expression following APC loss, indicating that chromatin-associated APC may antagonize canonical WNT signaling at both WNT-activated and WNT-repressed targets. Motif analysis and comparison to ChIP-seq datasets for other transcription factors identified a prevalence of binding sites for the TCF7L2 and AP-1 transcription factors in APC-associated genomic regions. Our results indicate that canonical WNT signaling can collaborate with or antagonize the AP-1 transcription factor to fine-tune the expression of shared target genes in the colorectal epithelium. Future therapeutic strategies for APC-deficient colorectal cancers might be expanded to include agents targeting the AP-1 pathway.