Rosenkranz, Stephan - CAP 7

Role of PI3Kd in atherosclerosis

The main research focus is the importance of receptor tyrosine kinases (RTK), non-receptor tyrosine kinases and other growth factors in vascular remodeling. Recent projects utilizing genetic and pharmacological approaches have identified critical growth factor-dependent signaling mechanisms in atherosclerosis, neointima formation, pulmonary hypertension and heart failure, which gain insight into the pathogenic mechanisms of these diseases and serve as novel therapeutic targets.

Introduction

Although numerous studies indicate that receptor tyrosine kinase (RTK) mediated signals are critically involved in the pathobiology of vascular disease, they are not readily targeted at present, and the precise underlying signaling mechanisms remain largely elusive. We thus focus on the importance of signal relay initiated by RTKs, non-receptor TKs and other growth factors in vascular diseases such as atherosclerosis, neointima formation, and pulmonary hypertension (PH). We initially characterized PDGF-dependent signaling pathways by generating and utilizing panels of PDGF receptor mutants in vitro and in vivo. These investigations identified PI 3-kinase (PI3K) as the critical signal relay molecule driving pathogenic vascular remodeling downstream of the PDGFR. Based on the results of this systematic approach, and in order to specifically target PI3K signaling, we more recently focussed on the importance of RTK-activated class IA PI3K isoforms, particularly p110α and p110δ. The rationale of this approach is to identify specific but central targets downstream of multiple RTKs. Furthermore, we focused on protease-activated receptors (PARs) and their role in pulmonary vascular remodeling.

Class IA PI3K subunit p110α is critical for vascular remodeling

Neointima formation following coronary interventions remains a significant problem in clinical cardiology. Conditional deletion of the RTK downstream target p110α in smooth muscle-specific knockout mice blunted the mitogenic and chemotactic signal of vascular smooth muscle cells (VSMCs) upon stimulation with multiple growth factors, and led to a complete abrogation of neointima formation after vascular injury in vivo, thus identifying p110α as a promising target for the prevention of neointima formation after vascular interventions (Vantler et al., ATVB 2015). In addition, we identified the role of the βPDGFR and PDGF-dependent downstream pathways via PI3K and PLCγ for pulmonary vascular remodeling and PH (ten Freyhaus, ATVB 2015), a devastating disease with unacceptably high mortality, and targeted deletion of p110α in VSMCs prevented the onset and progression of PH in vivo (Berghausen, submitted). Our genetic models provide direct proof for a role of βPDGFR/RTK-induced p110α signaling in the pathobiology of PH. Moreover, pharmacological inhibition of p110α was effective in reversing established PH in both the MCT and Su/Hx models of PH. Hence, p110α appears as a promising central therapeutic target in PH.

Class IA PI3K subunit p110α is critical for vascular remodeling

Neointima formation following coronary interventions remains a significant problem in clinical cardiology. Conditional deletion of the RTK downstream target p110α in smooth muscle-specific knockout mice blunted the mitogenic and chemotactic signal of vascular smooth muscle cells (VSMCs) upon stimulation with multiple growth factors, and led to a complete abrogation of neointima formation after vascular injury in vivo, thus identifying p110α as a promising target for the prevention of neointima formation after vascular interventions (Vantler et al., ATVB 2015). In addition, we identified the role of the βPDGFR and PDGF-dependent downstream pathways via PI3K and PLCγ for pulmonary vascular remodeling and PH (ten Freyhaus, ATVB 2015), a devastating disease with unacceptably high mortality, and targeted deletion of p110α in VSMCs prevented the onset and progression of PH in vivo (Berghausen, submitted). Our genetic models provide direct proof for a role of βPDGFR/RTK-induced p110α signaling in the pathobiology of PH. Moreover, pharmacological inhibition of p110α was effective in reversing established PH in both the MCT and Su/Hx models of PH. Hence, p110α appears as a promising central therapeutic target in PH.

An unexpected role for p110δ in atherosclerosis

The onset of acute coronary syndromes due to vul-nerability and rupture of atherosclerotic plaques is of major importance, yet the mechanisms of plaque progression and vulnerability are poorly understood. TK signaling and involvement of leukocytes appear to be of major importance. In this context, we hypothesized that hematopoietic p110δ activity is an important contributor to plaque formation and vulnerability. Surprisingly, we found that p110δ exerts anti-atherogenic functions, as hematopoietic PI3Kδ deficiency led to aggravated atherosclerosis in LDLR-/- mice (Fig. 1). This was associated with profound reductions of T cells, CD4+ T-cell activation, B lymphocytes and Tregs. PI3Kδ-deficient Tregs exhibited impaired immune-suppressive capabilities. Consequently, adoptive transfer of PI3Kδ+/+ Tregs into PI3Kδ-/- transplanted LDLR-/- mice rescued the atherosclerotic phenotype of PI3Kδ-/- transplanted LDLR-/- mice (Zierden et al., submitted).

Deficiency of protease-activated receptor 1 and 2 protect against pulmonary hypertension

We recently found that protease-activated receptors (PARs) 1 and 2, which are activated via coagulation Factor Xa (FXa) and thrombin, mediate VSMC migration and proliferation and thus promote vascular remodeling. In mice, both PAR-1 and PAR-2 deficiency substantially decreased hypoxia-induced PH and RV hypertrophy when compared to WT controls (Fig. 2). Analysis of PASMC from these mice revealed that thrombin and FXa induced proliferation via PAR-1 and PAR-2, respectively (Joseph et al., unpublished). These results demonstrate a crucial role for PARs in vascular remodeling, indicating a link between coagulation proteases and PARs, which may be relevant for anticoagulative drugs.

Perspectives

Our recent and ongoing projects provide the molecular basis for the development of new therapeutic concepts in the context of RTK signaling and vascular disease that are currently further explored. Specific aspects of ongoing work include testing of orally available compounds inhibiting p110α, the development of strategies for localized / cell type-specific targeting, evaluation of anti-inflammatory approaches (STAMP-2, IL-6, p110δ), and mechanistic insights into p110δ signaling in PH and atherosclerosis. The results will further increase our understanding of vascular remodeling, and will thus help to improve diagnostic, preventive, and therapeutic strategies in various vascular diseases.

Selected publications

Opitz CF, Hoeper MM, Gibbs JSR, Kaemmerer H, Pepke-Zaba J, Coghlan JG, Scelsi L, D’Alto M, Olsson KM, Ulrich S, Scholtz W, Schulz U, Grünig E, Vizza CD, Staehler G, Bruch L, Huscher D, Pittrow D, Rosenkranz S (2016) Pre-capillary, combined, and post-capillary pulmonary hypertension: A pathophysiological continuum. J Am Coll Cardiol 68: 368-378.

Rosenkranz S, Gibbs JSR, Wachter R, De Marco T, Vonk-Noordegraaf A, Vachiéry JL (2016) Left ventricular heart failure and pulmonary hypertension. Eur Heart J 2016; 37: 942-954.

Behringer A, Trappiel M, Berghausen EM, ten Freyhaus H, Wellnhofer E, Odenthal M, Blaschke F, Er F, Gassanov N, Rosenkranz S, Baldus S, Kappert K, Caglayan E (2016) Pioglita-zone alleviates cardiac and vascular remodelling and improves survival in monocrotaline induced pulmonary arterial hyper-tension. Naunyn Schmiedebergs Arch Pharmacol 389: 369-379.

Vantler M, Jesus J, Leppänen O, Scherner M, Berghausen EM, Mustafov L, Chen X, Kramer T, Zierden M, Gerhardt F, ten Freyhaus H, Blaschke F, Sterner-Kock A, Baldus S, Zhao JJ, Rosenkranz S (2015) Class IA phosphatidylinositol 3-kinase isoform p110 mediates vascular remodeling. Arterioscler Thromb Vasc Biol 35: 1434-1444.

Ten Freyhaus H, Berghausen EM, Janssen W, Leuchs M, Zierden M, Murmann K, Klinke A, Vantler M, Caglayan E, Kramer T, Baldus S, Schermuly RT, Tallquist MD, Rosenkranz S (2015) Genetic ablation of PDGF-dependent signaling pathways abolishes vascular remodeling and experimental pulmonary hypertension. Arterioscler Thromb Vasc Biol 35: 1236-1245.

Klinke A, Möller A, Pekarova M, Ravekes T, Friedrichs K, Berlin M, Scheu KM, Kubala L, Kolarova H, Ambrozova G, Schermuly RT, Woodcock SR, Freeman BA, Rosenkranz S, Baldus S, Rudolph V, Rudolph TK (2014) Protective effects of 10-Nitro-Oleic acid in a hypoxia-induced murine model of pulmonary hypertension. Am J Respir Cell Mol Biol 51: 155-162.

Kukat A, Dogan SA, Edgar D, Mourier A, Jacoby C,  Maiti P, Senft K, Wibom R, Kudin A, Hultenby K, Flögel U, Rosenkranz S, Ric-quier D, Wolfram S, Kunz WS, Trifunovic A (2014)  Loss of UCP2 attenuates mitochondrial dysfunction without altering ROS pro-duction and uncoupling activity. PLoS Genetics 10: e1004385.

Former Funding Period 01/2014 - 12/2016

Information from this funding period will not be updated anymore. New research related information is available here.


Prof. Dr. med. Stephan Rosenkranz

Dept. III of Internal Medicine

Prof. Dr. med. Stephan Rosenkranz

Principal Investigator CAP 7

Stephan.Rosenkranz@uk-koeln.de,

Work +49 221 478 32401

Heart Center University Hospital Cologne
Kerpener Str. 62
50937 Cologne

http://herzzentrum.uk-koeln.de/de/kardiologie/forschung/ag_rosenkranz

Publications

Link to PubMed

Group Members

Marius Vantler, PhD (PostDoc)
Eva M. Berghausen, PhD (PostDoc)
Mario Zierden, PhD (PostDoc)
Christine Joseph (PhD doctoral student)
Mehreen Batool (PhD doctoral student)
Frank Oberhäuser (technician)
Max Becker (technician)
Jana Thomas (doctoral student)
Maximilian Gerhardt (doctoral student)
Jonas Alfitian (doctoral student)
Fabian Nienhaus (doctoral student)
Christopher Millarg (doctoral student)
Christopher Skoruppa (doctoral student)

Further clinical members of the group, related to basic research / CMMC:

Henrik ten Freyhaus (MD)
Samuel Lee (MD)
Tilman Kramer (MD)
Joana Jesus (MD)
Felix Gerhardt (MD)
Jessika Schnitker (MD)

Figure 1

Figure 1: Hematopoietic deficiency of PI3Kδ aggravates athero-sclerosis in LDLR-/- mice. Bone marrows from PI3Kδ+/+ or PI3Kδ-/- mice were transplanted into lethally irradiated LDLR-/- mice. Representative oil red O-stained aortic root sections and quantification of lesion size in aortic roots (n = 10-20; ***p<0.001).

Figure 2

Figure 2: Reduced hypoxia-induced PH in PAR-1-/- and PAR2-/- deficient mice compared to WT. Shown are right ventricular systolic pressure and RV hypertrophy (n=10-12; ***p<0.001)