Intratumoral heterogeneity (ITH) is a fundamental property of cancer; however, the origins of ITH remain poorly understood. We performed single-cell transcriptome profiling of peritoneal ...carcinomatosis (PC) from 15 patients with gastric adenocarcinoma (GAC), constructed a map of 45,048 PC cells, profiled the transcriptome states of tumor cell populations, incisively explored ITH of malignant PC cells and identified significant correlates with patient survival. The links between tumor cell lineage/state compositions and ITH were illustrated at transcriptomic, genotypic, molecular and phenotypic levels. We uncovered the diversity in tumor cell lineage/state compositions in PC specimens and defined it as a key contributor to ITH. Single-cell analysis of ITH classified PC specimens into two subtypes that were prognostically independent of clinical variables, and a 12-gene prognostic signature was derived and validated in multiple large-scale GAC cohorts. The prognostic signature appears fundamental to GAC carcinogenesis and progression and could be practical for patient stratification.
The high comorbidity between obesity and mental disorders, such as depression and anxiety, often exacerbates metabolic and neurological symptoms significantly. However, neural mechanisms that ...underlie reciprocal control of feeding and mental states are largely elusive. Here we report that melanocortin 4 receptor (MC4R) neurons located in the dorsal bed nucleus of the stria terminus (dBNST) engage in the regulation of mentally associated weight gain by receiving GABAergic projections from hypothalamic AgRP neurons onto α5-containing GABA
receptors and serotonergic afferents onto 5-HT
receptors. Chronic treatment with a high-fat diet (HFD) significantly blunts the hyperexcitability of AgRP neurons in response to not only hunger but also anxiety and depression-like stimuli. Such HFD-mediated desensitization reduces GABAergic outputs from AgRP neurons to downstream MC4R
neurons, resulting in severe mental dysregulation. Genetic enhancement of the GABA
R-α5 or suppression of the 5-HT
R within the MC4R
neurons not only abolishes HFD-induced anxiety and depression but also robustly reduces body weight by suppression of food intake. To gain further translational insights, we revealed that combined treatment of zonisamide (enhancing the GABA
R-α5 signaling) and granisetron (a selective 5-HT
R antagonist) alleviates mental dysfunction and yields a robust reversal of diet-induced obesity by reducing total calorie intake and altering food preference towards a healthy low-fat diet. Our results unveil a neural mechanism for reciprocal control of appetite and mental states, which culminates in a novel zonisamide-granisetron cocktail therapy for potential tackling the psychosis-obesity comorbidity.
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.
Autologous chimeric antigen receptor (CAR) T cell therapies targeting CD19 have high efficacy in large B cell lymphomas (LBCLs), but long-term remissions are observed in less than half of patients, ...and treatment-associated adverse events, such as immune effector cell-associated neurotoxicity syndrome (ICANS), are a clinical challenge. We performed single-cell RNA sequencing with capture-based cell identification on autologous axicabtagene ciloleucel (axi-cel) anti-CD19 CAR T cell infusion products to identify transcriptomic features associated with efficacy and toxicity in 24 patients with LBCL. Patients who achieved a complete response by positron emission tomography/computed tomography at their 3-month follow-up had three-fold higher frequencies of CD8 T cells expressing memory signatures than patients with partial response or progressive disease. Molecular response measured by cell-free DNA sequencing at day 7 after infusion was significantly associated with clinical response (P = 0.008), and a signature of CD8 T cell exhaustion was associated (q = 2.8 × 10
) with a poor molecular response. Furthermore, a rare cell population with monocyte-like transcriptional features was associated (P = 0.0002) with high-grade ICANS. Our results suggest that heterogeneity in the cellular and molecular features of CAR T cell infusion products contributes to variation in efficacy and toxicity after axi-cel therapy in LBCL, and that day 7 molecular response might serve as an early predictor of CAR T cell efficacy.
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.
Unlike the well-established picture for the entry of enveloped viruses, the mechanism of cellular entry of non-enveloped eukaryotic viruses remains largely mysterious. Picornaviruses are ...representative models for such viruses, and initiate this entry process by their functional receptors. Here we present the structural and functional studies of SCARB2, a functional receptor of the important human enterovirus 71 (EV71). SCARB2 is responsible for attachment as well as uncoating of EV71. Differences in the structures of SCARB2 under neutral and acidic conditions reveal that SCARB2 undergoes a pivotal pH-dependent conformational change which opens a lipid-transfer tunnel to mediate the expulsion of a hydrophobic pocket factor from the virion, a pre-requisite for uncoating. We have also identified the key residues essential for attachment to SCARB2, identifying the canyon region of EV71 as mediating the receptor interaction. Together these results provide a clear understanding of cellular attachment and initiation of uncoating for enteroviruses
Ebolavirus can cause hemorrhagic fever in humans with a mortality rate of 50%-90%. Currently, no approved vaccines and antiviral therapies are available. Human TIM1 is considered as an attachment ...factor for EBOV, enhancing viral infection through interaction with PS located on the viral envelope. However, reasons under- lying the preferable usage of hTIM-1, but not other PS binding receptors by filovirus, remain unknown. We firstly demonstrated a direct interaction between hTIM-1 and EBOV GP in vitro and determined the crystal structures of the Ig V domains of hTIM-1 and hTIM-4. The binding region in hTIM-1 to EBOV GP was mapped by chimeras and mutation assays, which were designed based on structural analysis. Pseudovirion infection assays performed using hTIM-1 and its homologs as well as point mutants verified the location of the GP binding site and the importance of EBOV GP-hTIM-1 interaction in EBOV cellular entry.
BackgroundIn LUAD, KEAP1 is the third most common tumor suppressor and loss-of-function mutations in KEAP1 commonly co-occur with STK11/LKB1 and KRAS mutations. KEAP1 protein that regulates the ...degradation of the antioxidant transcription factor NRF2. The role of STK11/LKB1 mutations in immunotherapy resistance has been characterized, however the mechanistic understanding of KEAP1 deficiency in shaping LUAD phenotype and therapy response is still very limited. Recent clinical data has been reported suggesting that mutations in STK11/LKB1 and KEAP1 are strongly associated with immune checkpoint blockade resistance in LUAD, particularly those with KRAS mutations. Nevertheless, the biology of KEAP1-deficient tumors and the immune suppression mechanisms are to be characterized.MethodsWe have first validated response to anti-PD1 treatment in vivo using subcutaneous murine models, and performed a deep profiling and characterization of tumor microenvironment (TME) heterogeneity of KRAS-mutant (K) and LKB1 (KL), and/or KEAP1 deficient (KK and KLK) tumors using single-cell RNA sequencing (scRNA-seq) and multiplex staining. Data from pre-clinical models has been used to survey the immune genomic data available from the MD Anderson ICON study (a cohort of early stage lung cancer untreated 148 resected tumors) and TCGA lung cohorts to further validate our findings.ResultsWhile K tumors showed significant response to anti-PD1 treatment, KEAP1 loss completely impaired therapeutic response to this immunotherapy. KEAP1-deficient tumors were characterized by low immune infiltration while displayed an enrichment of cancer associated fibroblasts (CAFs) and endothelial cells. scRNA-seq data indicated a significant reduction of T cell infiltration, in particularly, CD8 and NK T cells, pronounced decreased of B cell population and a marked M2 macrophages polarization. Likewise, IHC and multiplex analysis of CD3 and F4/80 markers confirmed these previous findings. In TCGA lung cancer cohort, CD8B expression was dramatically decreased while MIF (macrophage migration inhibitory factor) was upregulated in KK compared to K LUADs tumors, and expression of KEAP1 inversely correlated with CD163, ARG2 and IL10, which are mainly secreted by macrophages. Concordantly, KEAP1-deficient pre-clinical tumors showed a significant upregulation of MIF expression and secretion, and CRISPR-Cas9 deletion of MIF dramatically impaired in vivo tumor growth in KK and KLK but not in K or KL models.ConclusionsThese findings indicate that loss of KEAP1, alone or in combination with STK11/LKB1 alterations, unfavorably reprograms TME. These changes appear to be mediated at least in part through MIF upregulation, providing a potential therapeutic strategy for overcoming KEAP1-dependent resistance to immunotherapy.
BackgroundKRAS-mutant non-small cell lung cancers (NSCLC) have exhibited unique response patterns to immunotherapy based on their co-occurring mutations. Patients harboring KRAS & STK11/LKB1 ...co-mutations (KL) have experienced shorter progression-free and overall survival compared to those with only KRAS mutations (K). Despite their limited responses, KL tumors exhibit a tumor mutational burden comparable to their K counterparts, suggesting the presence of additional mechanisms impairing antigen-specific responses. Accordingly, here we investigated the role of the MHC I antigen processing and presentation pathway in KL tumors.MethodsTCGA lung adenocarcinoma (LUAD) data were investigated for changes in expression of HLA molecules and chaperones involved in antigen processing and presentation. In mice, we performed single cell RNA sequencing of resected LKR13 K and KL tumors to evaluate changes in the tumor microenvironment and intrinsic differences in tumor antigen processing machinery. In vitro experiments were performed using the ovalbumin antigen to evaluate changes in antigen-specific T cell responses.ResultsExpression of HLA-A (p<0.0001), -B (p<0.0001), -C (p<0.0001), and beta2-microglobulin (B2M, p<0.0002) was downregulated in KL tumors from TCGA, as were expression of the TAP1 (p<0.001) and TAP2 (p<0.001) transporter associated with antigen processing subunits. LKR13 KL tumors exhibited similar patterns with lower H2-k1 (p<0.0001), H2-d1 (p<0.0001), B2m (p<0.0001), Tap1 (p<0.0001) and Tap2 (p<0.0001). As a result, LKR13 KL were resistant to recognition (p<0.005) and killing (56.9% K versus 7.8% KL) by OT-I T cells. Decreased expression of IFN-gamma-regulated genes such as PSMB8 (p<0.001), PSMB9 (p<0.0001), PSMB10 (p<001), CIITA (p<0.0001), NLRC5 (p<0.0001), IFNGR1 (p<0.0001), and IFNGR2 (p<0.0001) was also noted in KL tumors. Accordingly, KL tumors were unresponsive to exogenous IFN-gamma stimulation, maintaining repression of surface H2-Kb and resistance to T cell recognition (p<0.05) and killing (12.8% K versus 4% KL). Expression of T cell chemokines and receptors CXCR3 (p<0.0001), CXCL9 (p<0.0001), and CXCL10 (p<0.0001) was also repressed, potentially contributing to the lack of T cell infiltration in KL tumors.ConclusionsKRAS-mutant tumors harboring STK11/LKB1 alterations have an immunosuppressed phenotype and resistance to PD-1/PD-L1 inhibitors. Our findings provide evidence that these alterations are associated with markedly reduced antigen presentation and resistance to T cell killing, responsiveness to IFN-gamma stimulation, and impaired production of T cell chemokines, providing mechanistic insights into this immunosuppressed phenotype that could help guide the development of new therapeutic strategies for enhancing anti-tumor immunity.
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