Human endogenous retrovirus onco-exaptation counters cancer cell senescence through calbindin.

Increased levels and diversity of human endogenous retrovirus (HERV) transcription characterize most cancer types and are linked with disease outcomes. However, the underlying processes are incompletely understood. Here, we show that elevated transcription of HERVH proviruses predicted survival of lung squamous cell carcinoma (LUSC) and identified an isoform of CALB1, encoding calbindin, ectopically driven by an upstream HERVH provirus under the control of KLF5, as the mediator of this effect. HERVH-CALB1 expression was initiated in preinvasive lesions and associated with their progression. Calbindin loss in LUSC cell lines impaired in vitro and in vivo growth and triggered senescence, consistent with a protumor effect. However, calbindin also directly controlled the senescence-associated secretory phenotype (SASP), marked by secretion of CXCL8 and other neutrophil chemoattractants. In established carcinomas, CALB1-negative cancer cells became the dominant source of CXCL8, correlating with neutrophil infiltration and worse prognosis. Thus, HERVH-CALB1 expression in LUSC may display antagonistic pleiotropy, whereby the benefits of escaping senescence early during cancer initiation and clonal competition were offset by the prevention of SASP and protumor inflammation at later stages.

Antibodies against endogenous retroviruses promote lung cancer immunotherapy.

B cells are frequently found in the margins of solid tumours as organized follicles in ectopic lymphoid organs called tertiary lymphoid structures (TLS)1,2. Although TLS have been found to correlate with improved patient survival and response to immune checkpoint blockade (ICB), the underlying mechanisms of this association remain elusive1,2. Here we investigate lung-resident B cell responses in patients from the TRACERx 421 (Tracking Non-Small-Cell Lung Cancer Evolution Through Therapy) and other lung cancer cohorts, and in a recently established immunogenic mouse model for lung adenocarcinoma3. We find that both human and mouse lung adenocarcinomas elicit local germinal centre responses and tumour-binding antibodies, and further identify endogenous retrovirus (ERV) envelope glycoproteins as a dominant anti-tumour antibody target. ERV-targeting B cell responses are amplified by ICB in both humans and mice, and by targeted inhibition of KRAS(G12C) in the mouse model. ERV-reactive antibodies exert anti-tumour activity that extends survival in the mouse model, and ERV expression predicts the outcome of ICB in human lung adenocarcinoma. Finally, we find that effective immunotherapy in the mouse model requires CXCL13-dependent TLS formation. Conversely, therapeutic CXCL13 treatment potentiates anti-tumour immunity and synergizes with ICB. Our findings provide a possible mechanistic basis for the association of TLS with immunotherapy response.

An Immunogenic Model of KRAS-Mutant Lung Cancer Enables Evaluation of Targeted Therapy and Immunotherapy Combinations.

Mutations in oncogenes such as KRAS and EGFR cause a high proportion of lung cancers. Drugs targeting these proteins cause tumor regression but ultimately fail to elicit cures. As a result, there is an intense interest in how to best combine targeted therapies with other treatments, such as immunotherapies. However, preclinical systems for studying the interaction of lung tumors with the host immune system are inadequate, in part due to the low tumor mutational burden in genetically engineered mouse models. Here we set out to develop mouse models of mutant KRAS-driven lung cancer with an elevated tumor mutational burden by expressing the human DNA cytosine deaminase, APOBEC3B, to mimic the mutational signature seen in human lung cancer. This failed to substantially increase clonal tumor mutational burden and autochthonous tumors remained refractory to immunotherapy. However, establishing clonal cell lines from these tumors enabled the generation of an immunogenic syngeneic transplantation model of KRAS-mutant lung adenocarcinoma that was sensitive to immunotherapy. Unexpectedly, antitumor immune responses were not directed against neoantigens but instead targeted derepressed endogenous retroviral antigens. The ability of KRASG12C inhibitors to cause regression of KRASG12C -expressing tumors was markedly potentiated by the adaptive immune system, highlighting the importance of using immunocompetent models for evaluating targeted therapies. Overall, this model provides a unique opportunity for the study of combinations of targeted and immunotherapies in immune-hot lung cancer. SIGNIFICANCE: This study develops a mouse model of immunogenic KRAS-mutant lung cancer to facilitate the investigation of optimal combinations of targeted therapies with immunotherapies.

GREM1 is required to maintain cellular heterogeneity in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) shows pronounced epithelial and mesenchymal cancer cell populations1-4. Cellular heterogeneity in PDAC is an important feature in disease subtype specification3-5, but how distinct PDAC subpopulations interact, and the molecular mechanisms that underlie PDAC cell fate decisions, are incompletely understood. Here we identify the BMP inhibitor GREM16,7 as a key regulator of cellular heterogeneity in pancreatic cancer in human and mouse. Grem1 inactivation in established PDAC in mice resulted in a direct conversion of epithelial into mesenchymal PDAC cells within days, suggesting that persistent GREM1 activity is required to maintain the epithelial PDAC subpopulations. By contrast, Grem1 overexpression caused an almost complete 'epithelialization' of highly mesenchymal PDAC, indicating that high GREM1 activity is sufficient to revert the mesenchymal fate of PDAC cells. Mechanistically, Grem1 was highly expressed in mesenchymal PDAC cells and inhibited the expression of the epithelial-mesenchymal transition transcription factors Snai1 (also known as Snail) and Snai2 (also known as Slug) in the epithelial cell compartment, therefore restricting epithelial-mesenchymal plasticity. Thus, constant suppression of BMP activity is essential to maintain epithelial PDAC cells, indicating that the maintenance of the cellular heterogeneity of pancreatic cancer requires continuous paracrine signalling elicited by a single soluble factor.

Functional antibody and T-cell immunity following SARS-CoV-2 infection, including by variants of concern, in patients with cancer: the CAPTURE study.

Patients with cancer have higher COVID-19 morbidity and mortality. Here we present the prospective CAPTURE study (NCT03226886) integrating longitudinal immune profiling with clinical annotation. Of 357 patients with cancer, 118 were SARS-CoV-2-positive, 94 were symptomatic and 2 patients died of COVID-19. In this cohort, 83% patients had S1-reactive antibodies, 82% had neutralizing antibodies against WT, whereas neutralizing antibody titers (NAbT) against the Alpha, Beta, and Delta variants were substantially reduced. Whereas S1-reactive antibody levels decreased in 13% of patients, NAbT remained stable up to 329 days. Patients also had detectable SARS-CoV-2-specific T cells and CD4+ responses correlating with S1-reactive antibody levels, although patients with hematological malignancies had impaired immune responses that were disease and treatment-specific, but presented compensatory cellular responses, further supported by clinical. Overall, these findings advance the understanding of the nature and duration of immune response to SARS-CoV-2 in patients with cancer.

Favorable antibody responses to human coronaviruses in children and adolescents with autoimmune rheumatic diseases.

BACKGROUND: Differences in humoral immunity to coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), between children and adults remain unexplained, and the effect of underlying immune dysfunction or suppression is unknown. Here, we sought to examine the antibody immune competence of children and adolescents with prevalent inflammatory rheumatic diseases, juvenile idiopathic arthritis (JIA), juvenile dermatomyositis (JDM), and juvenile systemic lupus erythematosus (JSLE) against the seasonal human coronavirus (HCoV)-OC43 that frequently infects this age group. METHODS: Sera were collected from JIA (n = 118), JDM (n = 49), and JSLE (n = 30) patients and from healthy control (n = 54) children and adolescents prior to the coronavirus disease 19 (COVID-19) pandemic. We used sensitive flow-cytometry-based assays to determine titers of antibodies that reacted with the spike and nucleoprotein of HCoV-OC43 and cross-reacted with the spike and nucleoprotein of SARS-CoV-2, and we compared them with respective titers in sera from patients with multisystem inflammatory syndrome in children and adolescents (MIS-C). FINDINGS: Despite immune dysfunction and immunosuppressive treatment, JIA, JDM, and JSLE patients maintained comparable or stronger humoral responses than healthier peers, which was dominated by immunoglobulin G (IgG) antibodies to HCoV-OC43 spike, and harbored IgG antibodies that cross-reacted with SARS-CoV-2 spike. In contrast, responses to HCoV-OC43 and SARS-CoV-2 nucleoproteins exhibited delayed age-dependent class-switching and were not elevated in JIA, JDM, and JSLE patients, which argues against increased exposure. CONCLUSIONS: Consequently, autoimmune rheumatic diseases and their treatment were associated with a favorable ratio of spike to nucleoprotein antibodies. FUNDING: This work was supported by a Centre of Excellence Centre for Adolescent Rheumatology Versus Arthritis grant, 21593, UKRI funding reference MR/R013926/1, the Great Ormond Street Children's Charity, Cure JM Foundation, Myositis UK, Lupus UK, and the NIHR Biomedical Research Centres at GOSH and UCLH. This work was supported by the Francis Crick Institute, which receives its core funding from Cancer Research UK, the UK Medical Research Council, and the Wellcome Trust.

The transition from normal lung anatomy to minimal and established fibrosis in idiopathic pulmonary fibrosis (IPF).

BACKGROUND: The transition from normal lung anatomy to minimal and established fibrosis is an important feature of the pathology of idiopathic pulmonary fibrosis (IPF). The purpose of this report is to examine the molecular and cellular mechanisms associated with this transition. METHODS: Pre-operative thoracic Multidetector Computed Tomography (MDCT) scans of patients with severe IPF (n = 9) were used to identify regions of minimal(n = 27) and established fibrosis(n = 27). MDCT, Micro-CT, quantitative histology, and next-generation sequencing were used to compare 24 samples from donor controls (n = 4) to minimal and established fibrosis samples. FINDINGS: The present results extended earlier reports about the transition from normal lung anatomy to minimal and established fibrosis by showing that there are activations of TGFBI, T cell co-stimulatory genes, and the down-regulation of inhibitory immune-checkpoint genes compared to controls. The expression patterns of these genes indicated activation of a field immune response, which is further supported by the increased infiltration of inflammatory immune cells dominated by lymphocytes that are capable of forming lymphoid follicles. Moreover, fibrosis pathways, mucin secretion, surfactant, TLRs, and cytokine storm-related genes also participate in the transitions from normal lung anatomy to minimal and established fibrosis. INTERPRETATION: The transition from normal lung anatomy to minimal and established fibrosis is associated with genes that are involved in the tissue repair processes, the activation of immune responses as well as the increased infiltration of CD4, CD8, B cell lymphocytes, and macrophages. These molecular and cellular events correlate with the development of structural abnormality of IPF and probably contribute to its pathogenesis.

Functional antibody and T cell immunity following SARS-CoV-2 infection, including by variants of concern, in patients with cancer: the CAPTURE study.

Patients with cancer have higher COVID-19 morbidity and mortality. Here we present the prospective CAPTURE study, integrating longitudinal immune profiling with clinical annotation. Of 357 patients with cancer, 118 were SARS-CoV-2 positive, 94 were symptomatic and 2 died of COVID-19. In this cohort, 83% patients had S1-reactive antibodies and 82% had neutralizing antibodies against wild type SARS-CoV-2, whereas neutralizing antibody titers against the Alpha, Beta and Delta variants were substantially reduced. S1-reactive antibody levels decreased in 13% of patients, whereas neutralizing antibody titers remained stable for up to 329 days. Patients also had detectable SARS-CoV-2-specific T cells and CD4+ responses correlating with S1-reactive antibody levels, although patients with hematological malignancies had impaired immune responses that were disease and treatment specific, but presented compensatory cellular responses, further supported by clinical recovery in all but one patient. Overall, these findings advance the understanding of the nature and duration of the immune response to SARS-CoV-2 in patients with cancer.

Assessment of long non-coding RNA expression reveals novel mediators of the lung tumour immune response.

The tumour immune microenvironment is a crucial mediator of lung tumourigenesis, and characterizing the immune landscape of patient tumours may guide immunotherapy treatment regimens and uncover novel intervention points. We sought to identify the landscape of tumour-infiltrating immune cells in the context of long non-coding RNA (lncRNAs), known regulators of gene expression. We examined the lncRNA profiles of lung adenocarcinoma (LUAD) tumours by interrogating RNA sequencing data from microdissected and non-microdissected samples (BCCRC and TCGA). Subsequently, analysis of single-cell RNA sequencing data from lung tumours and flow-sorted healthy peripheral blood mononuclear cells identified lncRNAs in immune cells, highlighting their biological and prognostic relevance. We discovered lncRNA expression patterns indicative of regulatory relationships with immune-related protein-coding genes, including the relationship between AC008750.1 and NKG7 in NK cells. Activation of NK cells in vitro was sufficient to induce AC008750.1 expression. Finally, siRNA-mediated knockdown of AC008750.1 significantly impaired both the expression of NKG7 and the anti-tumour capacity of NK cells. We present an atlas of cancer-cell extrinsic immune cell-expressed lncRNAs, in vitro evidence for a functional role of lncRNAs in anti-tumour immune activity, which upon further exploration may reveal novel clinical utility as markers of immune infiltration.

Soluble PD-L1 generated by endogenous retroelement exaptation is a receptor antagonist.

Immune regulation is a finely balanced process of positive and negative signals. PD-L1 and its receptor PD-1 are critical regulators of autoimmune, antiviral and antitumoural T cell responses. Although the function of its predominant membrane-bound form is well established, the source and biological activity of soluble PD-L1 (sPD-L1) remain incompletely understood. Here, we show that sPD-L1 in human healthy tissues and tumours is produced by exaptation of an intronic LINE-2A (L2A) endogenous retroelement in the CD274 gene, encoding PD-L1, which causes omission of the transmembrane domain and the regulatory sequence in the canonical 3' untranslated region. The alternatively spliced CD274-L2A transcript forms the major source of sPD-L1 and is highly conserved in hominids, but lost in mice and a few related species. Importantly, CD274-L2A-encoded sPD-L1 lacks measurable T cell inhibitory activity. Instead, it functions as a receptor antagonist, blocking the inhibitory activity of PD-L1 bound on cellular or exosomal membranes.

Epithelial tumor suppressor ELF3 is a lineage-specific amplified oncogene in lung adenocarcinoma.

Gene function in cancer is often cell type-specific. The epithelial cell-specific transcription factor ELF3 is a documented tumor suppressor in many epithelial tumors yet displays oncogenic properties in others. Here, we show that ELF3 is an oncogene in the adenocarcinoma subtype of lung cancer (LUAD), providing genetic, functional, and clinical evidence of subtype specificity. We discover a region of focal amplification at chromosome 1q32.1 encompassing the ELF3 locus in LUAD which is absent in the squamous subtype. Gene dosage and promoter hypomethylation affect the locus in up to 80% of LUAD analyzed. ELF3 expression was required for tumor growth and a pan-cancer expression network analysis supports its subtype and tissue specificity. We further show that ELF3 displays strong prognostic value in LUAD but not LUSC. We conclude that, contrary to many other tumors of epithelial origin, ELF3 is an oncogene and putative therapeutic target in LUAD.

Somatic mutation-associated T follicular helper cell elevation in lung adenocarcinoma.

T follicular helper cells (Tfh) play crucial roles in the development of humoral immunity. In the B cell-rich germinal center of lymphoid organs, they select for high-affinity B cells and aid in their maturation. While Tfh have known roles in B cell malignancies and have prognostic value in some epithelial cancers, their role in lung tumour initiation and development is unknown. Through immune cell deconvolution, we observed significantly increased Tfh in tumours from two independent cohorts of lung adenocarcinomas and found that this upregulation occurs early in tumour development. A subset of tumours were stained for T and B cells using multicolour immunohistochemistry, which revealed the presence of tumour-adjacent tertiary lymphoid organs in 17/20 cases each with an average of 16 Tfh observed in the germinal center. Importantly, Tfh levels were correlated with tumour mutational load and immunogenic cancer testis antigens, suggesting their involvement in mounting an active immune response against tumour neoantigens.

Large-scale discovery of previously undetected microRNAs specific to human liver.

MicroRNAs (miRNAs) are crucial regulators of gene expression in normal development and cellular homeostasis. While miRNA repositories contain thousands of unique sequences, they primarily contain molecules that are conserved across several tissues, largely excluding lineage and tissue-specific miRNAs. By analyzing small non-coding RNA sequencing data for abundance and secondary RNA structure, we discovered 103 miRNA candidates previously undescribed in liver tissue. While expression of some of these unannotated sequences is restricted to non-malignant tissue, downregulation of most of the sequences was detected in liver tumors, indicating their importance in the maintenance of liver homeostasis. Furthermore, target prediction revealed the involvement of the unannotated miRNA candidates in fatty-acid metabolism and tissue regeneration, which are key pathways in liver biology. Here, we provide a comprehensive analysis of the undiscovered liver miRNA transcriptome, providing new resources for a deeper exploration of organ-specific biology and disease.

Non-coding RNAs predict recurrence-free survival of patients with hypoxic tumours.

Hypoxia promotes tumour aggressiveness and reduces patient survival. A spectrum of poor outcome among patients with hypoxic tumours suggests that additional factors modulate how tumours respond to hypoxia. PIWI-interacting RNAs (piRNAs) are small non-coding RNAs with a pivotal role in genomic stability and epigenetic regulation of gene expression. We reported that cancer type-specific piRNA signatures vary among patients. However, remarkably homogenous piRNA profiles are detected across patients with renal cell carcinoma, a cancer characterized by constitutive upregulation of hypoxia-related signaling induced by common mutation or loss of von Hippel-Lindau factor (VHL). By investigating >3000 piRNA transcriptomes in hypoxic and non-hypoxic tumors from seven organs, we discovered 40 hypoxia-regulated piRNAs and validated this in cells cultured under hypoxia. Moreover, a subset of these hypoxia-regulated piRNAs are regulated by VHL/HIF signaling in vitro. A hypoxia-regulated piRNA-based score (PiSco) was associated with poor RFS for hypoxic tumours, particularly Stage I lung adenocarcinomas, suggesting that hypoxia-regulated piRNA expression can predict tumour recurrence even in early-stage tumours and thus may be of clinical utility.

cGAS-STING and Cancer: Dichotomous Roles in Tumor Immunity and Development.

cGMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) sensing has emerged as a key regulator of innate immune responses to both exogenous and endogenous DNA. Recent studies reveal critical roles for this pathway in natural antitumor immunity across cancer types as well as in immune checkpoint blockade therapy. However, it is also clear that some tumors evade cGAS-STING-mediated immune responses, and immunomodulatory therapeutics are currently being explored to target this pathway. Finally, we also discuss recent observations that cGAS-STING-mediated inflammation may promote tumor initiation, growth, and metastasis in certain malignancies and how this may complicate the utility of this pathway in therapeutic development.

Small non-coding RNA transcriptome of the NCI-60 cell line panel.

Only 3% of the transcribed human genome is translated into protein, and small non-coding RNAs from these untranslated regions have demonstrated critical roles in transcriptional and translational regulation of proteins. Here, we provide a resource that will facilitate cell line selection for gene expression studies involving sncRNAs in cancer research. As the most accessible and tractable models of tumours, cancer cell lines are widely used to study cancer development and progression. The NCI-60 panel of 59 cancer cell lines was curated to provide common models for drug screening in 9 tissue types; however, its prominence has extended to use in gene regulation, xenograft models, and beyond. Here, we present the complete small non-coding RNA (sncRNA) transcriptomes of these 59 cancer cell lines. Additionally, we examine the abundance and unique sequences of annotated microRNAs (miRNAs), PIWI-interacting RNAs (piRNAs), small nuclear RNAs (snRNAs), and small nucleolar RNAs (snoRNAs), and reveal novel unannotated microRNA sequences.

DLK1-DIO3 imprinted locus deregulation in development, respiratory disease, and cancer.

INTRODUCTION: The imprinted DLK1-DIO3 locus at 14q32.1-32.31 holds biological significance in fetal development, whereby imprinting errors are causal to developmental disorders. Emerging evidence has implicated this locus in other diseases including cancer, highlighting the biological parallels between fetal organ and tumour development. Areas covered: Controlled regulation of gene expression from the imprinted DLK1-DIO3 locus at 14q32.1-32.31 is crucial for proper fetal development. Deregulation of locus gene expression due to imprinting errors has been mechanistically linked to the developmental disorders Kagami-Ogata Syndrome and Temple Syndrome. In adult tissues, deregulation of locus genes has been associated with multiple malignancies although the causal genetic mechanisms remain largely uncharacterised. Here, we summarize the genetic mechanisms underlying the developmental disorders that arise as a result of improper locus imprinting and the resulting developmental phenotypes, emphasizing both the coding and noncoding components of the locus. We further highlight biological parallels common to both fetal development and disease, with a specific focus on lung development, respiratory disease, and lung cancer. Expert commentary: Many commonalities between respiratory and developmental defects have emerged with respect to the 14q32 locus, emphasizing the importance of studying the effects of imprinting on gene regulation patterns at this locus in both biological settings.

Oncogenomic disruptions in arsenic-induced carcinogenesis.

Chronic exposure to arsenic affects more than 200 million people worldwide, and has been associated with many adverse health effects, including cancer in several organs. There is accumulating evidence that arsenic biotransformation, a step in the elimination of arsenic from the human body, can induce changes at a genetic and epigenetic level, leading to carcinogenesis. At the genetic level, arsenic interferes with key cellular processes such as DNA damage-repair and chromosomal structure, leading to genomic instability. At the epigenetic level, arsenic places a high demand on the cellular methyl pool, leading to global hypomethylation and hypermethylation of specific gene promoters. These arsenic-associated DNA alterations result in the deregulation of both oncogenic and tumour-suppressive genes. Furthermore, recent reports have implicated aberrant expression of non-coding RNAs and the consequential disruption of signaling pathways in the context of arsenic-induced carcinogenesis. This article provides an overview of the oncogenomic anomalies associated with arsenic exposure and conveys the importance of non-coding RNAs in the arsenic-induced carcinogenic process.

Emerging roles of T helper 17 and regulatory T cells in lung cancer progression and metastasis.

Lung cancer is a leading cause of cancer-related deaths worldwide. Lung cancer risk factors, including smoking and exposure to environmental carcinogens, have been linked to chronic inflammation. An integral feature of inflammation is the activation, expansion and infiltration of diverse immune cell types, including CD4+ T cells. Within this T cell subset are immunosuppressive regulatory T (Treg) cells and pro-inflammatory T helper 17 (Th17) cells that act in a fine balance to regulate appropriate adaptive immune responses.In the context of lung cancer, evidence suggests that Tregs promote metastasis and metastatic tumor foci development. Additionally, Th17 cells have been shown to be an integral component of the inflammatory milieu in the tumor microenvironment, and potentially involved in promoting distinct lung tumor phenotypes. Studies have shown that the composition of Tregs and Th17 cells are altered in the tumor microenvironment, and that these two CD4+ T cell subsets play active roles in promoting lung cancer progression and metastasis.We review current knowledge on the influence of Treg and Th17 cells on lung cancer tumorigenesis, progression, metastasis and prognosis. Furthermore, we discuss the potential biological and clinical implications of the balance among Treg/Th17 cells in the context of the lung tumor microenvironment and highlight the potential prognostic function and relationship to metastasis in lung cancer.