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Small-cell lung cancer (SCLC) is speculated to harbor complex genomic intratumor heterogeneity (ITH) associated with high recurrence rate and suboptimal response to immunotherapy. Here, using multi-region whole exome/T cell receptor (TCR) sequencing as well as immunohistochemistry, we reveal a rather homogeneous mutational landscape but extremely cold and heterogeneous TCR repertoire in limited-stage SCLC tumors (LS-SCLCs). Compared to localized non-small cell lung cancers, LS-SCLCs have similar predicted neoantigen burden and genomic ITH, but significantly colder and more heterogeneous TCR repertoire associated with higher chromosomal copy number aberration (CNA) burden. Furthermore, copy number loss of IFN-γ pathway genes is frequently observed and positively correlates with CNA burden. Higher mutational burden, higher T cell infiltration and positive PD-L1 expression are associated with longer overall survival (OS), while higher CNA burden is associated with shorter OS in patients with LS-SCLC., Small-cell lung cancer (SCLC) is an aggressive disease with limited therapeutic options. Here the authors perform an immunogenomic analysis of limited-stage SCLC, revealing a homogeneous mutational landscape, but limited T-cell infiltration and a cold and heterogeneous T cell repertoire.

The mechanism by which anti-cancer immunity shapes early carcinogenesis of lung adenocarcinoma (ADC) is unknown. In this study, we characterize the immune contexture of invasive lung ADC and its precursors by transcriptomic immune profiling, T cell receptor (TCR) sequencing and multiplex immunofluorescence (mIF). Our results demonstrate that anti-tumor immunity evolved as a continuum from lung preneoplasia, to preinvasive ADC, minimally-invasive ADC and frankly invasive lung ADC with a gradually less effective and more intensively regulated immune response including down-regulation of immune-activation pathways, up-regulation of immunosuppressive pathways, lower infiltration of cytotoxic T cells (CTLs) and anti-tumor helper T cells (Th), higher infiltration of regulatory T cells (Tregs), decreased T cell clonality, and lower frequencies of top T cell clones in later-stages. Driver mutations, chromosomal copy number aberrations (CNAs) and aberrant DNA methylation may collectively impinge host immune responses and facilitate immune evasion, promoting the outgrowth of fit subclones in preneoplasia into dominant clones in invasive ADC. The evolution of immune landscape in lung adenocarcinoma (ADC) is largely unknown. Here the authors use a cohort of resected invasive lung ADC and its precursors and show a gradual increase of immunosuppression and decrease of anti-tumor response associated with specific genomic and epigenetic features.

Optimal pH conditions for efficient artificial photosynthesis, hydrogen/oxygen evolution reactions, and photoreduction of carbon dioxide are now successfully achievable with catalytic bipolar membranes-integrated water dissociation and in-situ acid-base generations. However, inefficiency and instability are severe issues in state-of-the-art membranes, which need to urgently resolve with systematic membrane designs and innovative, inexpensive junctional catalysts. Here we show a shielding and in-situ formation strategy of fully-interconnected earth-abundant goethite Fe+3O(OH) catalyst, which lowers the activation energy barrier from 5.15 to 1.06 eV per HO − H bond and fabricates energy-efficient, cost-effective, and durable shielded catalytic bipolar membranes. Small water dissociation voltages at limiting current density (ULCD: 0.8 V) and 100 mA cm−2 (U100: 1.1 V), outstanding cyclic stability at 637 mA cm−2, long-time electro-stability, and fast acid-base generations (H2SO4: 3.9 ± 0.19 and NaOH: 4.4 ± 0.21 M m−2 min−1 at 100 mA cm−2) infer confident potential use of the novel bipolar membranes in emerging sustainable technologies., Bipolar membranes integrated water dissociation and acid-base generations have great potential in emerging sustainable technologies but remains inefficient. Here, the authors circumvent this inefficiency and instability of the membranes by developing polyaniline shielded catalytic bipolar membranes.

The evolution of DNA methylome and methylation intra-tumor heterogeneity (ITH) during early carcinogenesis of lung adenocarcinoma has not been systematically studied. We perform reduced representation bisulfite sequencing of invasive lung adenocarcinoma and its precursors, atypical adenomatous hyperplasia, adenocarcinoma in situ and minimally invasive adenocarcinoma. We observe gradual increase of methylation aberrations and significantly higher level of methylation ITH in later-stage lesions. The phylogenetic patterns inferred from methylation aberrations resemble those based on somatic mutations suggesting parallel methylation and genetic evolution. De-convolution reveal higher ratio of T regulatory cells (Tregs) versus CD8 + T cells in later-stage diseases, implying progressive immunosuppression with neoplastic progression. Furthermore, increased global hypomethylation is associated with higher mutation burden, copy number variation burden and AI burden as well as higher Treg/CD8 ratio, highlighting the potential impact of methylation on chromosomal instability, mutagenesis and tumor immune microenvironment during early carcinogenesis of lung adenocarcinomas., It is known that invasive lung adenocarcinomas evolve from pre-cancerous dysplastic lesions. In this study, the authors show that evolution of pre-cancerous lesions is accompanied by DNA methylation alterations, and that global hypomethylation correlates with immune infiltration, mutational burden and copy number alterations.

Acute graft versus host disease (aGVHD) is a rare, but severe complication of liver transplantation (LT). It is caused by the activation of donor immune cells in the graft against the host shortly after transplantation, but the contributing pathogenic factors remain unclear. Here we show that human T cell lymphotropic virus type I (HTLV-1) infection of donor T cells is highly associated with aGVHD following LT. The presence of HTLV-1 in peripheral blood and tissue samples from a discovery cohort of 7 aGVHD patients and 17 control patients is assessed with hybridization probes (TargetSeq), mass cytometry (CyTOF), and multiplex immunohistology (IMC). All 7 of our aGVHD patients display detectable HTLV-1 Tax signals by IMC. We identify donor-derived cells based on a Y chromosome-specific genetic marker, EIF1AY. Thus, we confirm the presence of CD4+Tax+EIF1AY+ T cells and Tax+CD68+EIF1AY+ antigen-presenting cells, indicating HTLV-1 infection of donor immune cells. In an independent cohort of 400 patients, we verify that HTLV-1 prevalence correlates with aGVHD incidence, while none of the control viruses shows significant associations. Our findings thus provide new insights into the aetio-pathology of liver-transplantation-associated aGVHD and raise the possibility of preventing aGVHD prior to transplantation.

Endoplasmic reticulum-associated degradation (ERAD) is a unique mechanism to degrade misfolded proteins via complexes containing several highly-conserved ER-anchored ubiquitin ligases such as HMG-CoA reductase degradation1 (Hrd1). Arabidopsis has a similar Hrd1-containing ERAD machinery; however, our knowledge of this complex is limited. Here we report two closely-related Arabidopsis proteins, Protein Associated With Hrd1-1 (PAWH1) and PAWH2, which share a conserved domain with yeast Altered Inheritance of Mitochondria24. PAWH1 and PAWH2 localize to the ER membrane and associate with Hrd1 via EMS-mutagenized Bri1 Suppressor7 (EBS7), a plant-specific component of the Hrd1 complex. Simultaneously elimination of two PAWHs constitutively activates the unfolded protein response and compromises stress tolerance. Importantly, the pawh1 pawh2 double mutation reduces the protein abundance of EBS7 and Hrd1 and inhibits degradation of several ERAD substrates. Our study not only discovers additional plant-specific components of the Arabidopsis Hrd1 complex but also reveals a distinct mechanism for regulating the Hrd1 stability., Endoplasmic reticulum (ER)-associated degradation (ERAD) removes misfolded proteins from the secretory pathway. Here the authors identify two plant-specific proteins in Arabidopsis, PAWH1 and PAWH2, that bind to and stabilise the ER-anchored ubiquitin ligase Hrd1.

There has been a dramatic increase in the detection of lung nodules, many of which are preneoplasia atypical adenomatous hyperplasia (AAH), adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA) or invasive adenocarcinoma (ADC). The molecular landscape and the evolutionary trajectory of lung preneoplasia have not been well defined. Here, we perform multi-region exome sequencing of 116 resected lung nodules including AAH (n = 22), AIS (n = 27), MIA (n = 54) and synchronous ADC (n = 13). Comparing AAH to AIS, MIA and ADC, we observe progressive genomic evolution at the single nucleotide level and demarcated evolution at the chromosomal level supporting the early lung carcinogenesis model from AAH to AIS, MIA and ADC. Subclonal analyses reveal a higher proportion of clonal mutations in AIS/MIA/ADC than AAH suggesting neoplastic transformation of lung preneoplasia is predominantly associated with a selective sweep of unfit subclones. Analysis of multifocal pulmonary nodules from the same patients reveal evidence of convergent evolution., There has been a drastic increase in detection of lung nodules, many of which are precancers, preinvasive, minimally invasive or sometimes invasive lung cancers. Here, Hu et al. perform multi-region exome sequencing to discern the evolutional trajectory from precancers to invasive lung cancers.

Osteosarcoma is the most frequent primary bone tumor with poor prognosis. Through RNA-sequencing of 100,987 individual cells from 7 primary, 2 recurrent, and 2 lung metastatic osteosarcoma lesions, 11 major cell clusters are identified based on unbiased clustering of gene expression profiles and canonical markers. The transcriptomic properties, regulators and dynamics of osteosarcoma malignant cells together with their tumor microenvironment particularly stromal and immune cells are characterized. The transdifferentiation of malignant osteoblastic cells from malignant chondroblastic cells is revealed by analyses of inferred copy-number variation and trajectory. A proinflammatory FABP4+ macrophages infiltration is noticed in lung metastatic osteosarcoma lesions. Lower osteoclasts infiltration is observed in chondroblastic, recurrent and lung metastatic osteosarcoma lesions compared to primary osteoblastic osteosarcoma lesions. Importantly, TIGIT blockade enhances the cytotoxicity effects of the primary CD3+ T cells with high proportion of TIGIT+ cells against osteosarcoma. These results present a single-cell atlas, explore intratumor heterogeneity, and provide potential therapeutic targets for osteosarcoma., Osteosarcomas are difficult to treat and have a limited response to immunotherapy. Here, the authors analyse osteosarcomas at the single-cell level, and identify both the transdifferentiation of malignant cells and an array of immune cells that could have implications for metastasis and immunotherapy.

Methylthioadenosine phosphorylase, an essential enzyme for the adenine salvage pathway, is often deficient (MTAPdef) in tumors with 9p21 loss and hypothetically renders tumors susceptible to synthetic lethality by antifolates targeting de novo purine synthesis. Here we report our single arm phase II trial (NCT02693717) that assesses pemetrexed in MTAPdef urothelial carcinoma (UC) with the primary endpoint of overall response rate (ORR). Three of 7 enrolled MTAPdef patients show response to pemetrexed (ORR 43%). Furthermore, a historic cohort shows 4 of 4 MTAPdef patients respond to pemetrexed as compared to 1 of 10 MTAP-proficient patients. In vitro and in vivo preclinical data using UC cell lines demonstrate increased sensitivity to pemetrexed by inducing DNA damage, and distorting nucleotide pools. In addition, MTAP-knockdown increases sensitivity to pemetrexed. Furthermore, in a lung adenocarcinoma retrospective cohort (N = 72) from the published BATTLE2 clinical trial (NCT01248247), MTAPdef associates with an improved response rate to pemetrexed. Our data demonstrate a synthetic lethal interaction between MTAPdef and de novo purine inhibition, which represents a promising therapeutic strategy for larger prospective trials.

Subclonal architecture and genomic evolution of small-cell lung cancer (SCLC) under treatment has not been well studied primarily due to lack of tumor specimens, particularly longitudinal samples acquired during treatment. SCLC is characterized by early hematogenous spread, which makes circulating cell-free tumor DNA (ctDNA) sequencing a promising modality for genomic profiling. Here, we perform targeted deep sequencing of 430 cancer genes on pre-treatment tumor biopsies, as well as on plasma samples collected prior to and during treatment from 22 SCLC patients. Similar subclonal architecture is observed between pre-treatment ctDNA and paired tumor DNA. Mean variant allele frequency of clonal mutations from pre-treatment ctDNA is associated with progression-free survival and overall survival. Pre- and post-treatment ctDNA mutational analysis demonstrate that mutations of DNA repair and NOTCH signaling pathways are enriched in post-treatment samples. These data suggest that ctDNA sequencing is promising to delineate genomic landscape, subclonal architecture, and genomic evolution of SCLC., Small cell lung cancer (SCLC) may evolve under treatment. But tumor tissues are often not available to study evolution of SCLC. Here, the authors utilize circulating tumor DNA to investigate the genomic evolution and subclonal architecture of SCLC during therapy.

Previous studies from the Cancer Cell Line Encyclopedia (CCLE) project have adopted commercial pan-cancer cell line models to identify drug sensitivity biomarkers. However, drug sensitivity biomarkers in esophageal squamous cell carcinoma (ESCC) have not been widely explored. Here, eight patient-derived cell lines (PDCs) are successfully established from 123 patients with ESCC. The mutation profiling of PDCs can partially recapture the tumor tissue actionable mutations from 161 patients with ESCC. Based on these mutations and relative pathways in eight PDCs, 46 targeted drugs are selected for screening. Interestingly, some drug and biomarker relationships are established that were not discovered in the CCLE project. For example, CDKN2A or CDKN2B loss is significantly associated with the sensitivity of CDK4/6 inhibitors. Furthermore, both PDC xenografts and patient-derived xenografts confirm CDKN2A/2B loss as a biomarker predictive of CDK4/6 inhibitor sensitivity. Collectively, patient-derived models could predict targeted drug sensitivity associated with actionable mutations in ESCC., Predicting the drug response of patients with cancer is crucial for implementing targeted therapy. Here, Su et al. make patient-derived cell lines and perform targeted sequencing and RNA-seq to identify CDKN2A/2B loss as a predictor of response to CDK4/6 inhibitors in esophageal squamous cell carcinoma.

Immunotherapy targeting T cells is increasingly utilized to treat solid tumors including non-small cell lung cancer (NSCLC). This requires a better understanding of the T cells in the lungs of patients with NSCLC. Here, we report T cell repertoire analysis in a cohort of 236 early-stage NSCLC patients. T cell repertoire attributes are associated with clinicopathologic features, mutational and immune landscape. A considerable proportion of the most prevalent T cells in tumors are also prevalent in the uninvolved tumor-adjacent lungs and appear specific to shared background mutations or viral infections. Patients with higher T cell repertoire homology between the tumor and uninvolved tumor-adjacent lung, suggesting a less tumor-focused T cell response, exhibit inferior survival. These findings indicate that a concise understanding of antigens and T cells in NSCLC is needed to improve therapeutic efficacy and reduce toxicity with immunotherapy, particularly adoptive T cell therapy., Relevant features of T cell repertoire in human cancer remain to be delineated. Here the authors show, by TCR sequencing in a large cohort of lung cancer patients, that while a majority of T cell clones are shared between tumor and adjacent lung tissue, less frequent tumor-unique T cell clones correlate with worse prognosis.

Subclonal architecture and genomic evolution of small-cell lung cancer (SCLC) under treatment has not been well studied primarily due to lack of tumor specimens, particularly longitudinal samples acquired during treatment. SCLC is characterized by early hematogenous spread, which makes circulating cell-free tumor DNA (ctDNA) sequencing a promising modality for genomic profiling. Here, we perform targeted deep sequencing of 430 cancer genes on pre-treatment tumor biopsies, as well as on plasma samples collected prior to and during treatment from 22 SCLC patients. Similar subclonal architecture is observed between pre-treatment ctDNA and paired tumor DNA. Mean variant allele frequency of clonal mutations from pre-treatment ctDNA is associated with progression-free survival and overall survival. Pre- and post-treatment ctDNA mutational analysis demonstrate that mutations of DNA repair and NOTCH signaling pathways are enriched in post-treatment samples. These data suggest that ctDNA sequencing is promising to delineate genomic landscape, subclonal architecture, and genomic evolution of SCLC.

Multiple synchronous lung cancers (MSLCs) present a clinical dilemma as to whether individual tumours represent intrapulmonary metastases or independent tumours. In this study we analyse genomic profiles of 15 lung adenocarcinomas and one regional lymph node metastasis from 6 patients with MSLC. All 15 lung tumours demonstrate distinct genomic profiles, suggesting all are independent primary tumours, which are consistent with comprehensive histopathological assessment in 5 of the 6 patients. Lung tumours of the same individuals are no more similar to each other than are lung adenocarcinomas of different patients from TCGA cohort matched for tumour size and smoking status. Several known cancer-associated genes have different mutations in different tumours from the same patients. These findings suggest that in the context of identical constitutional genetic background and environmental exposure, different lung cancers in the same individual may have distinct genomic profiles and can be driven by distinct molecular events., Some patients present with multiple lung tumours but it is unclear whether these are metastases or individual lesions. Here, the authors use genomics techniques to demonstrate in six patients that multiple tumours have individual genetic profiles and represent separate tumours.