Screening drugs on patient biopsies from solid tumours has immense potential, but is challenging due to the small amount of available material. To address this, we present here a plug-based ...microfluidics platform for functional screening of drug combinations. Integrated Braille valves allow changing the plug composition on demand and enable collecting >1200 data points (56 different conditions with at least 20 replicates each) per biopsy. After deriving and validating efficient and specific drug combinations for two genetically different pancreatic cancer cell lines and xenograft mouse models, we additionally screen live cells from human solid tumours with no need for ex vivo culturing steps, and obtain highly specific sensitivity profiles. The entire workflow can be completed within 48 h at assay costs of less than US$ 150 per patient. We believe this can pave the way for rapid determination of optimal personalized cancer therapies.
Primary liver tumors and liver metastasis currently represent the leading cause of cancer-related death. Commensal bacteria are important regulators of antitumor immunity, and although the liver is ...exposed to gut bacteria, their role in antitumor surveillance of liver tumors is poorly understood. We found that altering commensal gut bacteria in mice induced a liver-selective antitumor effect, with an increase of hepatic CXCR6
natural killer T (NKT) cells and heightened interferon-γ production upon antigen stimulation. In vivo functional studies showed that NKT cells mediated liver-selective tumor inhibition. NKT cell accumulation was regulated by CXCL16 expression of liver sinusoidal endothelial cells, which was controlled by gut microbiome-mediated primary-to-secondary bile acid conversion. Our study suggests a link between gut bacteria-controlled bile acid metabolism and liver antitumor immunosurveillance.
Oncogene-induced senescence causes hepatocytes to secrete cytokines, which induce their immune-mediated clearance to prevent tumor initiation, a process termed “senescence surveillance.” However, ...senescent hepatocytes give rise to hepatocellular carcinomas (HCCs), if the senescence program is bypassed or if senescent cells are not cleared. Here, we show context-specific roles for CCR2+ myeloid cells in liver cancer. Senescence surveillance requires the recruitment and maturation of CCR2+ myeloid cells, and CCR2 ablation caused outgrowth of HCC. In contrast, HCC cells block the maturation of recruited myeloid precursors, which, through NK cell inhibition, promote growth of murine HCC and worsen the prognosis and survival of human HCC patients. Thus, while senescent hepatocyte-secreted chemokines suppress liver cancer initiation, they may accelerate the growth of fully established HCC.
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•The CCL2-CCR2 axis is necessary for clearance of pre-cancerous senescent hepatocytes•Absence of the CCL2-CCR2 axis leads to HCC outgrowth from senescent hepatocytes•Peritumoral tissue senescence accelerates growth of HCC in mice and humans•Senescence-recruited CCR2+ myeloid cells enhance HCC growth by NK cell inhibition
Eggert et al. show that CCL2 is secreted from oncogene-induced senescent hepatocytes to recruit CCR2+ immature myeloid cells (iMC). These iMC differentiate into macrophages that clear pre-malignant senescent cells, but iMC promote growth of established hepatocellular carcinoma through NK cell inhibition.
Macrophages are key regulators of liver fibrosis progression and regression in nonalcoholic steatohepatitis (NASH). Liver macrophages comprise resident phagocytes, Kupffer cells, and monocyte‐derived ...cells, which are recruited through the chemokine receptor C‐C motif chemokine receptor 2 (CCR2). We aimed at elucidating the therapeutic effects of inhibiting monocyte infiltration in NASH models by using cenicriviroc (CVC), an oral dual chemokine receptor CCR2/CCR5 antagonist that is under clinical evaluation. Human liver tissues from NASH patients were analyzed for CCR2+ macrophages, and administration of CVC was tested in mouse models of steatohepatitis, liver fibrosis progression, and fibrosis regression. In human livers from 17 patients and 4 controls, CCR2+ macrophages increased parallel to NASH severity and fibrosis stage, with a concomitant inflammatory polarization of these cluster of differentiation 68+, portal monocyte‐derived macrophages (MoMF). Similar to human disease, we observed a massive increase of hepatic MoMF in experimental models of steatohepatitis and liver fibrosis. Therapeutic treatment with CVC significantly reduced the recruitment of hepatic Ly‐6C+ MoMF in all models. In experimental steatohepatitis with obesity, therapeutic CVC application significantly improved insulin resistance and hepatic triglyceride levels. In fibrotic steatohepatitis, CVC treatment ameliorated histological NASH activity and hepatic fibrosis. CVC inhibited the infiltration of Ly‐6C+ monocytes, without direct effects on macrophage polarization, hepatocyte fatty acid metabolism, or stellate cell activation. Importantly, CVC did not delay fibrosis resolution after injury cessation. RNA sequencing analysis revealed that MoMF, but not Kupffer cells, specifically up‐regulate multiple growth factors and cytokines associated with fibrosis progression, while Kupffer cells activated pathways related to inflammation initiation and lipid metabolism. Conclusion: Pharmacological inhibition of CCR2+ monocyte recruitment efficiently ameliorates insulin resistance, hepatic inflammation, and fibrosis, corroborating the therapeutic potential of CVC in patients with NASH. (Hepatology 2018;67:1270‐1283)
Oncogene-induced senescence (OIS) is crucial for tumour suppression. Senescent cells implement a complex pro-inflammatory response termed the senescence-associated secretory phenotype (SASP). The ...SASP reinforces senescence, activates immune surveillance and paradoxically also has pro-tumorigenic properties. Here, we present evidence that the SASP can also induce paracrine senescence in normal cells both in culture and in human and mouse models of OIS in vivo. Coupling quantitative proteomics with small-molecule screens, we identified multiple SASP components mediating paracrine senescence, including TGF-β family ligands, VEGF, CCL2 and CCL20. Amongst them, TGF-β ligands play a major role by regulating p15(INK4b) and p21(CIP1). Expression of the SASP is controlled by inflammasome-mediated IL-1 signalling. The inflammasome and IL-1 signalling are activated in senescent cells and IL-1α expression can reproduce SASP activation, resulting in senescence. Our results demonstrate that the SASP can cause paracrine senescence and impact on tumour suppression and senescence in vivo.
Primary liver cancer represents a major health problem. It comprises hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC), which differ markedly with regards to their morphology, ...metastatic potential and responses to therapy. However, the regulatory molecules and tissue context that commit transformed hepatic cells towards HCC or ICC are largely unknown. Here we show that the hepatic microenvironment epigenetically shapes lineage commitment in mosaic mouse models of liver tumorigenesis. Whereas a necroptosis-associated hepatic cytokine microenvironment determines ICC outgrowth from oncogenically transformed hepatocytes, hepatocytes containing identical oncogenic drivers give rise to HCC if they are surrounded by apoptotic hepatocytes. Epigenome and transcriptome profiling of mouse HCC and ICC singled out Tbx3 and Prdm5 as major microenvironment-dependent and epigenetically regulated lineage-commitment factors, a function that is conserved in humans. Together, our results provide insight into lineage commitment in liver tumorigenesis, and explain molecularly why common liver-damaging risk factors can lead to either HCC or ICC.
Gut dysbiosis is commonly observed in patients with cirrhosis and chronic gastrointestinal disorders; however, its effect on antitumor immunity in the liver is largely unknown. Here we studied how ...the gut microbiome affects antitumor immunity in cholangiocarcinoma. Primary sclerosing cholangitis (PSC) or colitis, two known risk factors for cholangiocarcinoma which promote tumor development in mice, caused an accumulation of CXCR2
polymorphonuclear myeloid-derived suppressor cells (PMN-MDSC). A decrease in gut barrier function observed in mice with PSC and colitis allowed gut-derived bacteria and lipopolysaccharide to appear in the liver and induced CXCL1 expression in hepatocytes through a TLR4-dependent mechanism and an accumulation of CXCR2
PMN-MDSCs. In contrast, neomycin treatment blocked CXCL1 expression and PMN-MDSC accumulation and inhibited tumor growth even in the absence of liver disease or colitis. Our study demonstrates that the gut microbiome controls hepatocytes to form an immunosuppressive environment by increasing PMN-MDSCs to promote liver cancer. SIGNIFICANCE: MDSCs have been shown to be induced by tumors and suppress antitumor immunity. Here we show that the gut microbiome can control accumulation of MDSCs in the liver in the context of a benign liver disease or colitis.
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ObjectiveThere is a striking association between human cholestatic liver disease (CLD) and inflammatory bowel disease. However, the functional implications for intestinal microbiota and ...inflammasome-mediated innate immune response in CLD remain elusive. Here we investigated the functional role of gut–liver crosstalk for CLD in the murine Mdr2 knockout (Mdr2−/−) model resembling human primary sclerosing cholangitis (PSC).DesignMale Mdr2 −/−, Mdr2−/− crossed with hepatocyte-specific deletion of caspase-8 (Mdr2−/− /Casp8∆hepa) and wild-type (WT) control mice were housed for 8 or 52 weeks, respectively, to characterise the impact of Mdr2 deletion on liver and gut including bile acid and microbiota profiling. To block caspase activation, a pan-caspase inhibitor (IDN-7314) was administered. Finally, the functional role of Mdr2−/− -associated intestinal dysbiosis was studied by microbiota transfer experiments.Results Mdr2−/− mice displayed an unfavourable intestinal microbiota signature and pronounced NLRP3 inflammasome activation within the gut–liver axis. Intestinal dysbiosis in Mdr2−/− mice prompted intestinal barrier dysfunction and increased bacterial translocation amplifying the hepatic NLRP3-mediated innate immune response. Transfer of Mdr2−/− microbiota into healthy WT control mice induced significant liver injury in recipient mice, highlighting the causal role of intestinal dysbiosis for disease progression. Strikingly, IDN-7314 dampened inflammasome activation, ameliorated liver injury, reversed serum bile acid profile and cholestasis-associated microbiota signature.ConclusionsMDR2-associated cholestasis triggers intestinal dysbiosis. In turn, translocation of endotoxin into the portal vein and subsequent NLRP3 inflammasome activation contribute to higher liver injury. This process does not essentially depend on caspase-8 in hepatocytes, but can be blocked by IDN-7314.
Hepatitis C virus (HCV) is a major causative agent of chronic liver disease in humans. To gain insight into host factor requirements for HCV replication, we performed a siRNA screen of the human ...kinome and identified 13 different kinases, including phosphatidylinositol-4 kinase III alpha (PI4KIIIα), as being required for HCV replication. Consistent with elevated levels of the PI4KIIIα product phosphatidylinositol-4-phosphate (PI4P) detected in HCV-infected cultured hepatocytes and liver tissue from chronic hepatitis C patients, the enzymatic activity of PI4KIIIα was critical for HCV replication. Viral nonstructural protein 5A (NS5A) was found to interact with PI4KIIIα and stimulate its kinase activity. The absence of PI4KIIIα activity induced a dramatic change in the ultrastructural morphology of the membranous HCV replication complex. Our analysis suggests that the direct activation of a lipid kinase by HCV NS5A contributes critically to the integrity of the membranous viral replication complex.
Anaplastic thyroid cancer (ATC) is one of the most aggressive malignancies, representing less than 5% of all thyroid carcinomas. Τhe median survival is limited to months due to the resistance of ATC ...to surgery, radioiodine therapy, radiotherapy and chemotherapy. This review will cover novel agents involving several cellular signaling pathways including the BRAF pathway. The BRAF inhibitor vemurafenib improves survival among patients with metastatic melanoma, hairy-cell leukemia and intracranial neoplasms with BRAF gene mutations. The frequency of a BRAF (V600E) mutation in ATC is about 25%.
We report the first case of a marked partial response to adjuvant first line monotherapy with vemurafenib in BRAF V600E-mutated ATC. The 78-year-old man showed a sustained response for 7 months, thereafter scans revealed progressive disease and the patient died 10 months after first diagnosis. This case report is accompanied by a comprehensive review of current strategies and tools for ATC treatment.
This case and the review of current data confirm the benefit of BRAF inhibition in BRAF-mutated ATC, limited by acquired resistance to targeted therapy.
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