Hepatic stellate cells limit hepatocellular carcinoma progression through the orphan receptor endosialin

Abstract Hepatocellular carcinoma (HCC) is among the most common and deadliest cancers worldwide. A major contributor to HCC progression is the cross talk between tumor cells and the surrounding stroma including activated hepatic stellate cells (HSC). Activation of HSC during liver damage leads to upregulation of the orphan receptor endosialin (CD248), which contributes to regulating the balance of liver regeneration and fibrosis. Based on the established role of endosialin in regulating HSC/hepatocyte cross talk, we hypothesized that HSC‐expressed endosialin might similarly affect cell proliferation during hepatocarcinogenesis. Indeed, the histological analysis of human HCC samples revealed an inverse correlation between tumor cell proliferation and stromal endosialin expression. Correspondingly, global genetic inactivation of endosialin resulted in accelerated tumor growth in an inducible mouse HCC model. A candidate‐based screen of tumor lysates and differential protein arrays of cultured HSC identified several established hepatotropic cytokines, including IGF2, RBP4, DKK1, and CCL5 as being negatively regulated by endosialin. Taken together, the experiments identify endosialin‐expressing HSC as a negative regulator of HCC progression

Hepatic stellate cells limit hepatocellular carcinoma progression through the orphan receptor endosialin

Abstract Hepatocellular carcinoma (HCC) is among the most common and deadliest cancers worldwide. A major contributor to HCC progression is the cross talk between tumor cells and the surrounding stroma including activated hepatic stellate cells (HSC). Activation of HSC during liver damage leads to upregulation of the orphan receptor endosialin (CD248), which contributes to regulating the balance of liver regeneration and fibrosis. Based on the established role of endosialin in regulating HSC/hepatocyte cross talk, we hypothesized that HSC‐expressed endosialin might similarly affect cell proliferation during hepatocarcinogenesis. Indeed, the histological analysis of human HCC samples revealed an inverse correlation between tumor cell proliferation and stromal endosialin expression. Correspondingly, global genetic inactivation of endosialin resulted in accelerated tumor growth in an inducible mouse HCC model. A candidate‐based screen of tumor lysates and differential protein arrays of cultured HSC identified several established hepatotropic cytokines, including IGF2, RBP4, DKK1, and CCL5 as being negatively regulated by endosialin. Taken together, the experiments identify endosialin‐expressing HSC as a negative regulator of HCC progression

Postsurgical Adjuvant Tumor Therapy by Combining Anti-Angiopoietin-2 and Metronomic Chemotherapy Limits Metastatic Growth

Antiangiogenic tumor therapy has failed in the adjuvant setting. Here we show that inhibition of the Tie2 ligand angiopoietin-2 (Ang2) effectively blocks metastatic growth in preclinical mouse models of postsurgical adjuvant therapy. Ang2 antibody treatment combines well with low-dose metronomic chemotherapy (LDMC) in settings in which maximum-dose chemotherapy does not prove effective. Mechanistically, Ang2 blockade could be linked to quenching the inflammatory and angiogenic response of endothelial cells (ECs) in the metastatic niche. Reduced EC adhesion molecule and chemokine expression inhibits the recruitment of tumor-promoting CCR2(+)Tie2(-) metastasis-associated macrophages. Moreover, LDMC contributes to therapeutic efficacy by inhibiting the recruitment of protumorigenic bone marrow-derived myeloid cells. Collectively, these data provide a rationale for mechanism-guided adjuvant tumor therapies

Preclinical validation of a novel metastasis‐inhibiting Tie1 function‐blocking antibody

Abstract The angiopoietin (Ang)–Tie pathway has been intensely pursued as candidate second‐generation anti‐angiogenic target. While much of the translational work has focused on the ligand Ang2, the clinical efficacy of Ang2‐targeting drugs is limited and failed to improve patient survival. In turn, the orphan receptor Tie1 remains therapeutically unexplored, although its endothelial‐specific genetic deletion has previously been shown to result in a strong reduction in metastatic growth. Here, we report a novel Tie1 function‐blocking antibody (AB‐Tie1‐39), which suppressed postnatal retinal angiogenesis. During primary tumor growth, neoadjuvant administration of AB‐Tie1‐39 strongly impeded systemic metastasis. Furthermore, the administration of AB‐Tie1‐39 in a perioperative therapeutic window led to a significant survival advantage as compared to control‐IgG‐treated mice. Additional in vivo experimental metastasis and in vitro transmigration assays concurrently revealed that AB‐Tie1‐39 treatment suppressed tumor cell extravasation at secondary sites. Taken together, the data phenocopy previous genetic work in endothelial Tie1 KO mice and thereby validate AB‐Tie1‐39 as a Tie1 function‐blocking antibody. The study establishes Tie1 as a therapeutic target for metastasis in a perioperative or neoadjuvant setting

Hepatic stellate cell‐expressed endosialin balances fibrogenesis and hepatocyte proliferation during liver damage

Abstract Liver fibrosis is a reversible wound‐healing response to injury reflecting the critical balance between liver repair and scar formation. Chronic damage leads to progressive substitution of liver parenchyma by scar tissue and ultimately results in liver cirrhosis. Stromal cells (hepatic stellate cells [HSC] and endothelial cells) have been proposed to control the balance between liver fibrosis and regeneration. Here, we show that endosialin, a C‐type lectin, expressed in the liver exclusively by HSC and portal fibroblasts, is upregulated in liver fibrosis in mouse and man. Chronic chemically induced liver damage resulted in reduced fibrosis and enhanced hepatocyte proliferation in endosialin‐deficient (ENKO) mice. Correspondingly, acute‐liver‐damage‐induced hepatocyte proliferation (partial hepatectomy) was increased in ENKO mice. A candidate‐based screen of known regulators of hepatocyte proliferation identified insulin‐like growth factor 2 (IGF2) as selectively endosialin‐dependent hepatocyte mitogen. Collectively, the study establishes a critical role of HSC in the reciprocal regulation of fibrogenesis vs. hepatocyte proliferation and identifies endosialin as a therapeutic target in non‐neoplastic settings

Endosialin (Tem1) Is a Marker of Tumor-Associated Myofibroblasts and Tumor Vessel-Associated Mural Cells

Endosialin (Tem1) has been identified by two independent experimental approaches as an antigen of tumor-associated endothelial cells, and it has been claimed to be the most abundantly expressed tumor endothelial antigen, making it a prime candidate for vascular targeting purposes. Recent experiments have challenged the endothelial expression of endosialin and suggested an expression by activated fibroblasts and pericytes. Thus, clarification of the controversial cellular expression of endosialin is critically important for an understanding of its role during tumor progression and its validation as a potential therapeutic target. We have therefore performed extensive expression profiling analyses of endosialin. The experiments unambiguously demonstrate that endosialin is expressed by tumor-associated myofibroblasts and mural cells and not by endothelial cells. Endosialin expression is barely detectable in normal human tissues with moderate expression only detectable in the stroma of the colon and the prostate. Corresponding cellular experiments confirmed endosialin expression by mesenchymal cells and indicated that it may in fact be a marker of mesenchymal stem cells. Silencing endosialin expression in fibroblasts strongly inhibited migration and proliferation. Collectively, the experiments validate endosialin as a marker of tumor-associated myofibroblasts and tumor vessel-associated mural cells. The data warrant further functional analysis of endosialin during tumor progression and its exploitation as marker of tumor vessel-associated mural cells, expression of which may reflect the non-normalized phenotype of the tumor vasculature

Angiopoietin-2 differentially regulates angiogenesis through TIE2 and integrin signaling

Angiopoietin-2 (ANG-2) is a key regulator of angiogenesis that exerts context-dependent effects on ECs. ANG-2 binds the endothelial-specific receptor tyrosine kinase 2 (TIE2) and acts as a negative regulator of ANG-1/TIE2 signaling during angiogenesis, thereby controlling the responsiveness of ECs to exogenous cytokines. Recent data from tumors indicate that under certain conditions ANG-2 can also promote angiogenesis. However, the molecular mechanisms of dual ANG-2 functions are poorly understood. Here, we identify a model for the opposing roles of ANG-2 in angiogenesis. We found that angiogenesis-activated endothelium harbored a subpopulation of TIE2-negative ECs (TIE2lo). TIE2 expression was downregulated in angiogenic ECs, which abundantly expressed several integrins. ANG-2 bound to these integrins in TIE2lo ECs, subsequently inducing, in a TIE2-independent manner, phosphorylation of the integrin adaptor protein FAK, resulting in RAC1 activation, migration, and sprouting angiogenesis. Correspondingly, in vivo ANG-2 blockade interfered with integrin signaling and inhibited FAK phosphorylation and sprouting angiogenesis of TIE2lo ECs. These data establish a contextual model whereby differential TIE2 and integrin expression, binding, and activation control the role of ANG-2 in angiogenesis. The results of this study have immediate translational implications for the therapeutic exploitation of angiopoietin signaling