Iris Helfrich

Publications of Iris Helfrich

• A wound size-dependent effect of myeloid cell-derived vascular endothelial growth factor on wound healing.

  Authors: Christian Stockmann, Santina Kirmse, Iris Helfrich, Alexander Weidemann, Norihiko Takeda, Andrew Doedens, Randall S Johnson

  The Journal of investigative dermatology. 03/2011; 131(3):797-801.

• Blood vessel maturation, vascular phenotype and angiogenic potential in malignant melanoma: one step forward for overcoming anti-angiogenic drug resistance?

  Authors: Iris Helfrich, Dirk Schadendorf

  Molecular oncology. 02/2011; 5(2):137-49.

  Angiogenesis is a pivotal process for growth, invasion and spread of the majority of solid tumors including melanoma. Anti-angiogenic agents have not been systematically tested in patients withAngiogenesis is a pivotal process for growth, invasion and spread of the majority of solid tumors including melanoma. Anti-angiogenic agents have not been systematically tested in patients with advanced melanoma. Clinical efficacy of angiogenesis inhibitors targeting endothelial cells has not been as affirmative as initially hoped and improved clinical outcomes have been observed in combination with chemotherapy or additional drugs for many types of human cancer. However, angiogenesis is not only dependent on endothelial cell invasion and proliferation, it also requires pericyte coverage of vascular sprouts for stabilization and maturation of vascular walls. Recent data suggest that pericytes might be able to confer resistance to anti-vascular endothelial growth factor (VEGF) therapy. This review will focus on the significance of the vascular phenotype but also on the impact of pericyte-mediated vessel maturation for the susceptibility to anti-angiogenic therapy, including malignant melanoma, which we identified as crucial factor regarding therapeutic efficacy.

• The VEGF-regulated transcription factor HLX controls the expression of guidance cues and negatively regulates sprouting of endothelial cells.

  Authors: Julia Testori, Bernhard Schweighofer, Iris Helfrich, Caterina Sturtzel, Karoline Lipnik, Sabine Gesierich, Patrick Nasarre, Renate Hofer-Warbinek, Martin Bilban, Hellmut G Augustin, Erhard Hofer

  Blood. 01/2011; 117(9):2735-44.

  The HLX gene encoding a diverged homeobox transcription factor has been found to be up-regulated by vascular endothelial growth factor-A (VEGF-A) in endothelial cells. We have now investigated theThe HLX gene encoding a diverged homeobox transcription factor has been found to be up-regulated by vascular endothelial growth factor-A (VEGF-A) in endothelial cells. We have now investigated the gene repertoire induced by HLX and its potential biologic function. HLX strongly increased the transcripts for several repulsive cell-guidance proteins including UNC5B, plexin-A1, and semaphorin-3G. In addition, genes for transcriptional repressors such as HES-1 were up-regulated. In line with these findings, adenoviral overexpression of HLX inhibited endothelial cell migration, sprouting, and vessel formation in vitro and in vivo, whereas proliferation was unaffected. This inhibition of sprouting was caused to a significant part by HLX-mediated up-regulation of UNC5B as shown by short hairpin RNA (shRNA)-mediated down-modulation of the respective mRNA. VEGF-A stimulation of endothelial cells induced elevated levels of HLX over longer time periods resulting in especially high up-regulation of UNC5B mRNA as well as an increase in cells displaying UNC5B at their surface. However, induction of HLX was strongly reduced and UNC5B up-regulation completely abrogated when cells were exposed to hypoxic conditions. These data suggest that HLX may function to balance attractive with repulsive vessel guidance by up-regulating UNC5B and to down-modulate sprouting under normoxic conditions.

• Resistance to antiangiogenic therapy is directed by vascular phenotype, vessel stabilization, and maturation in malignant melanoma.

  Authors: Iris Helfrich, Inka Scheffrahn, Sönke Bartling, Joachim Weis, Verena von Felbert, Mark Middleton, Masahi Kato, Süleyman Ergün, Dirk Schadendorf

  The Journal of experimental medicine. 03/2010; 207(3):491-503.

  Angiogenesis is not only dependent on endothelial cell invasion and proliferation, it also requires pericyte coverage of vascular sprouts for stabilization of vascular walls. Clinical efficacy ofAngiogenesis is not only dependent on endothelial cell invasion and proliferation, it also requires pericyte coverage of vascular sprouts for stabilization of vascular walls. Clinical efficacy of angiogenesis inhibitors targeting the vascular endothelial growth factor (VEGF) signaling pathway is still limited to date. We hypothesized that the level of vessel maturation is critically involved in the response to antiangiogenic therapies. To test this hypothesis, we evaluated the vascular network in spontaneously developing melanomas of MT/ret transgenic mice after using PTK787/ZK222584 for anti-VEGF therapy but also analyzed human melanoma metastases taken at clinical relapse in patients undergoing adjuvant treatment using bevacizumab. Both experimental settings showed that tumor vessels, which are resistant to anti-VEGF therapy, are characterized by enhanced vessel diameter and normalization of the vascular bed by coverage of mature pericytes and immunoreactivity for desmin, NG-2, platelet-derived growth factor receptor beta, and the late-stage maturity marker alpha smooth muscle actin. Our findings emphasize that the level of mural cell differentiation and stabilization of the vascular wall significantly contribute to the response toward antiangiogenic therapy in melanoma. This study may be useful in paving the way toward a more rational development of second generation antiangiogenic combination therapies and in providing, for the first time, a murine model to study this.

• Host-Derived Angiopoietin-2 Affects Early Stages of Tumor Development and Vessel Maturation but Is Dispensable for Later Stages of Tumor Growth.

  Authors: Patrick Nasarre, Markus Thomas, Karoline Kruse, Iris Helfrich, Vivien Wolter, Carleen Deppermann, Dirk Schadendorf, Gavin Thurston, Ulrike Fiedler, Hellmut G Augustin

  Cancer research. 03/2009;

  The angiopoietin/Tie2 system has been identified as the second vascular-specific receptor tyrosine kinase system controlling vessel assembly, maturation, and quiescence. Angiopoietin-2 (Ang-2) isThe angiopoietin/Tie2 system has been identified as the second vascular-specific receptor tyrosine kinase system controlling vessel assembly, maturation, and quiescence. Angiopoietin-2 (Ang-2) is prominently up-regulated in the host-derived vasculature of most tumors, making it an attractive candidate for antiangiogenic intervention. Yet, the net outcome of Ang-2 functions on tumor angiogenesis is believed to be contextual depending on the local cytokine milieu. Correspondingly, Ang-2 manipulatory therapies have been shown to exert protumorigenic as well as antitumorigenic effects. To clarify the role of Ang-2 for angiogenesis and tumor growth in a definite genetic experimental setting, the present study was aimed at comparatively studying the growth of different tumors in wild-type and Ang-2-deficient mice. Lewis lung carcinomas, MT-ret melanomas, and B16F10 melanomas all grew slower in Ang-2-deficient mice. Yet, tumor growth in wild-type and Ang-2-deficient mice dissociated during early stages of tumor development, whereas tumor growth rates during later stages of primary tumor progression were similar. Analysis of the intratumoral vascular architecture revealed no major differences in microvessel density and perfusion characteristics. However, diameters of intratumoral microvessels were smaller in tumors grown in Ang-2-deficient mice, and the vasculature had an altered pattern of pericyte recruitment and maturation. Ang-2-deficient tumor vessels had higher pericyte coverage indices. Recruited pericytes were desmin and NG2 positive and predominately alpha-smooth muscle actin negative, indicative of a more mature pericyte phenotype. Collectively, the experiments define the role of Ang-2 during tumor angiogenesis and establish a better rationale for combination therapies involving Ang-2 manipulatory therapies. [Cancer Res 2009;69(4):1324-33].

• Angiopoietin-2 levels are associated with disease progression in metastatic malignant melanoma.

  Authors: Iris Helfrich, Lutz Edler, Antje Sucker, Markus Thomas, Sven Christian, Dirk Schadendorf, Hellmut G Augustin

  Clinical cancer research : an official journal of the American Association for Cancer Research. 03/2009; 15(4):1384-92.

  PURPOSE: The blood vessel-destabilizing Tie2 ligand angiopoietin-2 (Ang-2) acts in concert with the vascular endothelial growth factor/vascular endothelial growth factor receptor system to controlPURPOSE: The blood vessel-destabilizing Tie2 ligand angiopoietin-2 (Ang-2) acts in concert with the vascular endothelial growth factor/vascular endothelial growth factor receptor system to control vessel assembly during tumor progression. We hypothesized that circulating soluble Ang-2 (sAng-2) may be involved in melanoma progression. EXPERIMENTAL DESIGN: Serum samples (n=98) from melanoma patients (American Joint Committee on Cancer stages I-IV), biopsies of corresponding patients, and human melanoma cell lines were analyzed for expression of Ang-2 and S100beta. Multiple sera of a subcohort of 33 patients were tested during progression from stage III to IV. Small interfering RNA-based loss-of-function experiments were done to assess effects of Ang-2 on melanoma cells. RESULTS: Circulating levels of sAng-2 correlate with tumor progression in melanoma patients (P<0.0001) and patient survival (P=0.007). Analysis of serum samples during the transition from stage III to IV identified an increase of sAng-2 up to 400%. Comparative analyses revealed a 56% superiority of sAng-2 as predictive marker over the established marker S100beta. Immunohistochemistry and reverse transcription-PCR confirmed the prominent expression of Ang-2 by tumor-associated endothelial cells but identified Ang-2 also as a secreted product of melanoma cells themselves. Corresponding cellular experiments revealed that human melanoma-isolated tumor cells were Tie2 positive and that Ang-2 acted as an autocrine regulator of melanoma cell migration and invasion. CONCLUSIONS: The experiments establish sAng-2 as a biomarker of melanoma progression and metastasis correlating with tumor load and overall survival. The identification of an autocrine angiopoietin/Tie loop controlling melanoma migration and invasion warrants further functional experiments and validate the angiopoietin/Tie system as a promising therapeutic target for human melanomas.

• Endosialin (Tem1) is a marker of tumor-associated myofibroblasts and tumor vessel-associated mural cells.

  Authors: Sven Christian, Renate Winkler, Iris Helfrich, Anja M Boos, Eva Besemfelder, Dirk Schadendorf, Hellmut G Augustin

  The American journal of pathology. 03/2008; 172(2):486-94.

  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 tumorEndosialin (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.

• Role of aPKC isoforms and their binding partners Par3 and Par6 in epidermal barrier formation.

  Authors: Iris Helfrich, Annika Schmitz, Paola Zigrino, Christian Michels, Ingo Haase, André Le Bivic, Michael Leitges, Carien M Niessen

  The Journal of investigative dermatology. 05/2007; 127(4):782-91.

  The skin water barrier, essential for terrestrial life, is formed by a multilayered stratifying epithelium, which shows a polarized distribution of both differentiation and intercellular junctionThe skin water barrier, essential for terrestrial life, is formed by a multilayered stratifying epithelium, which shows a polarized distribution of both differentiation and intercellular junction markers. Recently, several reports showed the crucial importance of tight junctions for the in vivo water barrier function of the skin. In simple epithelial cells, intercellular junction formation is closely coupled to the establishment of polarity. However, if and how polarity proteins contribute to epidermal differentiation and junction formation is not yet known. Here, we have characterized the localization and isoform expression of the polarity protein atypical PKC (aPKC) and its binding partners Par3 and Par6 in epidermis and primary keratinocytes of mice. Their distribution is only partially overlapping in the granular layer, the site of functional tight junctions, suggesting that next to a common Par3/Par6/aPKC function they also may have functions independent of each other. Both aPKCzeta and aPKCiota/lambda, are expressed in the epidermis but only aPKCiota/lambda showed a strong enrichment in the junctions, suggesting that this aPKC isoform is important for epidermal tight junction function. Indeed, inhibition of aPKC function showed that endogenous aPKC is crucial for in vitro barrier function and this required the presence of both the Par3 and Par6 binding sites.

• E-cadherin is essential for in vivo epidermal barrier function by regulating tight junctions.

  Authors: Judith A Tunggal, Iris Helfrich, Annika Schmitz, Heinz Schwarz, Dorothee Günzel, Michael Fromm, Rolf Kemler, Thomas Krieg, Carien M Niessen

  The EMBO journal. 04/2005; 24(6):1146-56.

  Cadherin adhesion molecules are key determinants of morphogenesis and tissue architecture. Nevertheless, the molecular mechanisms responsible for the morphogenetic contributions of cadherins remainCadherin adhesion molecules are key determinants of morphogenesis and tissue architecture. Nevertheless, the molecular mechanisms responsible for the morphogenetic contributions of cadherins remain poorly understood in vivo. Besides supporting cell-cell adhesion, cadherins can affect a wide range of cellular functions that include activation of cell signalling pathways, regulation of the cytoskeleton and control of cell polarity. To determine the role of E-cadherin in stratified epithelium of the epidermis, we have conditionally inactivated its gene in mice. Here we show that loss of E-cadherin in the epidermis in vivo results in perinatal death of mice due to the inability to retain a functional epidermal water barrier. Absence of E-cadherin leads to improper localization of key tight junctional proteins, resulting in permeable tight junctions and thus altered epidermal resistance. In addition, both Rac and activated atypical PKC, crucial for tight junction formation, are mislocalized. Surprisingly, our results indicate that E-cadherin is specifically required for tight junction, but not desmosome, formation and this appears to involve signalling rather than cell contact formation.

• Increased incidence of squamous cell carcinomas in Mastomys natalensis papillomavirus E6 transgenic mice during two-stage skin carcinogenesis.

  Authors: Iris Helfrich, Min Chen, Rainer Schmidt, Gerhard Fürstenberger, Annette Kopp-Schneider, David Trick, Hermann-Josef Gröne, Harald zur Hausen, Frank Rösl

  Journal of virology. 05/2004; 78(9):4797-805.

  Papillomaviruses cause certain forms of human cancers, most notably carcinomas of the uterine cervix. In contrast to the well-established involvement of papillomavirus infection in the etiology ofPapillomaviruses cause certain forms of human cancers, most notably carcinomas of the uterine cervix. In contrast to the well-established involvement of papillomavirus infection in the etiology of cervical carcinomas and in carcinomas of a rare hereditary condition, epidermodysplasia verruciformis, a causative role for cutaneous human papillomavirus types in the development of nonmelanoma skin cancer has not been proven. In order to better understand the functions of individual genes of cutaneous papillomavirus types, we generated transgenic mice carrying oncogene E6 of the Mastomys natalensis papillomavirus (MnPV), which causes keratoacanthomas of the skin in its natural host. In the present study, we demonstrate that under conditions of experimental two-stage skin carcinogenesis, fast-paced squamous cell carcinomas develop in nearly 100% of MnPV E6 transgenic mice in comparison to 10% in their nontransgenic littermates (log rank test; P < 0.0001). Therefore, we conclude that the MnPV E6 transgene favors the malignant progression of chemically induced tumors. Whereas an activating H-ras mutation is a consistent feature in benign and malignant tumors in wild-type mice, the majority of papillomas and keratoacanthomas and all squamous cell carcinomas obtained in MnPV E6 transgenic mice contain nonmutated ras alleles. These results indicate that the development of squamous cell carcinomas in MnPV E6 transgenic mice does not depend on an activated H-ras oncogene.