Alexander Schultze

Axl, a prognostic and therapeutic target in acute myeloid leukemia mediates paracrine crosstalk of leukemia cells with bone marrow stroma

Acute myeloid leukemia (AML) represents a clonal disease of hematopoietic progenitors characterized by acquired heterogenous genetic changes that alter normal mechanisms of proliferation, self-renewal, and differentiation.(1) Although 40% to 45% of patients younger than 65 years of age can be cured with current therapies, only 10% of older patients reach long-term survival.(1) Because only very few novel AML drugs were approved in the past 2 decades, there is an urgent need to identify novel targets and therapeutic strategies to treat underserved AML patients. We report here that Axl, a member of the Tyro3, Axl, Mer receptor tyrosine kinase family,(2-4) represents an independent prognostic marker and therapeutic target in AML. AML cells induce expression and secretion of the Axl ligand growth arrest-specific gene 6 (Gas6) by bone marrow-derived stromal cells (BMDSCs). Gas6 in turn mediates proliferation, survival, and chemoresistance of Axl-expressing AML cells. This Gas6-Axl paracrine axis between AML cells and BMDSCs establishes a chemoprotective tumor cell niche that can be abrogated by Axl-targeting approaches. Axl inhibition is active in FLT3-mutated and FLT3 wild-type AML, improves clinically relevant end points, and its efficacy depends on presence of Gas6 and Axl. Axl inhibition alone or in combination with chemotherapy might represent a novel therapeutic avenue for AML.

Therapeutic potential and limitations of new FAK inhibitors in the treatment of cancer

Importance of the field: Activation of the non-receptor tyrosine kinase focal adhesion kinase (FAK) has been implicated in progression of multiple mesenchymal and epithelial malignant tumors. FAK plays an important role in regulation of proliferation, migration and apoptosis of neoplastic cells. Areas covered in this review: We review the importance of FAK expression as a prognostic marker in cancer patients, discuss the available small-molecule inhibitors of FAK, summarize the available data from early-phase clinical trials with FAK inhibitors and cover the antiangiogenic properties of FAK inhibitors, as well as their potential to overcome chemoresistance. What the reader will gain: This review enables the reader to overview current knowledge about FAK inhibition in cancer therapy and its role in the clinical setting. The reader will be able to consider FAK inhibitors not only as direct antitumor but also as antineoangiogenic agents and drugs that can overcome the problem of chemoresistance. Take home message: Emerging data from early-phase clinical trials with orally available small-molecule inhibitors of FAK are promising. There are early indicators of clinical efficacy. In the future, combination therapy with cytotoxic or antiangiogenic drugs may help to overcome chemoresistance and enhance efficacy of antivascular therapy.

Clinical Importance and Potential Use of Small Molecule Inhibitors of Focal Adhesion Kinase

Since its first description Focal Adhesion Kinase (FAK), a cytoplasmatic tyrosine kinase, has been implicated in the formation and progression of solid and liquid malignant tumors. Therefore orally available selective small molecule inhibitors of FAK have been developed, three of them (PF-562-271, PF-04554878 and GSK2256098) are already in clinical testing. This review discusses the recent data obtained from these Phase 1 trials. We also discuss available data on the mechanisms of action of these inhibitors in carcinogenesis and demonstrate that FAK plays an important role in neoangiogenesis which is a crucial step in cancer growth.

TAE226-mediated inhibition of focal adhesion kinase interferes with tumor angiogenesis and vasculogenesis

SummaryNeoangiogenesis plays an important role in tumor growth and metastasis. Evaluation of new anti-angiogenic targets may broaden the armament for future therapeutic concepts. Focal adhesion kinase (FAK), expressed in endothelial and tumor cells, is essential for adhesion and mobility of adherent cells. In the current study we analyzed the anti-angiogenic properties of the FAK inhibitor TAE226 on the proliferation of blood outgrowth endothelial cell (OEC) and differentiation of endothelial progenitor cells (EPC), derived from peripheral blood CD133+ cells, tube formation and on neovascularization in a HT29 xenotransplant model. The effects of TAE226 were compared to those of the rapamycin analogue RAD001. The combination of both drugs was also studied. We showed that HT29 tumor cells and OEC were most sensitive to the action of TAE226 compared to EPC in vitro. In contrast, RAD001 affected the proliferation of both types of endothelial cells stronger than that of HT29 cells. Furthermore we could show that TAE226 inhibited tube formation in a dose dependent manner. In a HT29 subcutaneous tumor model TAE226 and RAD001 diminished MVD at commonly employed doses to a similar degree. Combination of both compounds did not show synergy in vitro or in vivo. Since TAE226 has been shown to inhibit the PI3 kinase, Akt kinase, mTor pathway, addition of RAD001 may not increase this effect. In conclusion, we have shown that treatment with TAE leads to a reduction of neoangiogenesis in vitro and in a mouse model. The effects are mediated by inhibition of angiogenesis and vasculogenic OEC and EPC.

Macrophage–tumor crosstalk: role of TAMR tyrosine kinase receptors and of their ligands

Ample clinical and preclinical evidence indicates that macrophages interact with tumor cells as well as with virtually all populations of host cells present in the tumor microenvironment. This crosstalk can strongly promote malignancy, but also has in principle the potential to inhibit tumor growth. Thus, it is of the utmost importance to improve our understanding of the mechanisms driving the pro- and antimalignant behavior of tumor-associated macrophages (TAMs) in order to develop better anticancer therapies. In this review, we discuss the biological consequences of reciprocal interactions between TAMs, cancer cells, endothelial cells, fibroblasts and other leukocyte subfractions within tumors. It was recently elucidated that tumors specifically educate macrophages to secrete growth arrest-specific gene 6 (Gas6), the common ligand of the Tyro3, Axl, Mer receptor (TAMR) family. In turn, Gas6 fosters tumor growth by promoting cancer cell proliferation. Therefore, the Gas6–TAMR axis might represent a novel target for disrupting tumor–macrophage crosstalk. We summarize here what is known about TAMR and their ligands in (human) cancer biology. In order to shed more light on the role of macrophages in human cancer, we additionally provide an overview of what is currently known about the prognostic impact of TAMs in human cancer.

ErbB2 signaling activates the Hedgehog pathway via PI3K–Akt in human esophageal adenocarcinoma: Identification of novel targets for concerted therapy concepts

The Hedgehog pathway plays an important role in the pathogenesis of several tumor types, including esophageal cancer. In our study, we show an expression of the ligand Indian hedgehog (Ihh) and its downstream mediator Gli-1 in primary resected adenocarcinoma tissue by immunohistochemistry and quantitative PCR in fifty percent of the cases, while matching healthy esophagus mucosa was negative for both proteins. Moreover, a functionally important regulation of Gli-1 by ErbB2-PI3K-mTORC signaling as well as a Gli-1-dependent regulation of Ihh in the ErbB2 amplified esophageal adenocarcinoma cell line OE19 was observed. Treatment of OE19 cells with the Her2 antibody trastuzumab, the PI3K-mTORC1 inhibitor NVP BEZ235 (BEZ235) or the knockdown of Akt1 resulted in a downregulation of Gli-1 and Ihh as well as in a reduction of viable OE19 cells in vitro. Interestingly, the Hedgehog receptor Smo, which acts upstream of Gli-1, was not expressed in OE19 cells and in the majority of primary human esophageal adenocarcinoma, suggesting a non-canonical upregulation of Gli-1 expression by the ErbB2-PI3K axis. To translate our findings into a therapeutic concept, we targeted ErbB2-PI3K-mTORC1 by trastuzumab and BEZ235, combining both compounds with the Gli-1/2 inhibitor GANT61. The triple combination led to significantly stronger reduction of tumor cell viability than cisplatinum or each biological alone. Therefore, concomitant blockage of the ErbB2-PI3K pathway and the Hedgehog downstream mediator Gli-1 may provide a new therapeutic strategy for esophageal cancer.

Blood outgrowth endothelial cells from chronic myeloid leukaemia patients are BCR/ABL1 negative

The existence of adult haemangioblasts with dual haematopoietic and endothelial developmental potential was confirmed after detection of Ph+ vascular endothelial cells in chronic myeloid leukaemia (CML) patients. Blood outgrowth endothelial cells (OECs) from CML patients were found not to harbour the Philadelphia translocation and were thus not clonally related to BRC/ABL1+ hematopoietic progenitors, but comprised a distinct subfraction of endothelial cells. Remarkably, the frequency of CML‐derived OECs was 9‐fold higher as compared to healthy donors (n = 19 and n = 300, respectively; P < 0·0001) and these cells showed increased proliferative potential, possibly reflecting the mobilisation of OEC progenitors by pro‐angiogenic cytokines.

Quantitative MR imaging of targeted SPIO particles on the cell surface and comparison to flow cytometry

PURPOSE To detect anti-CEACAM5 targeted superparamagnetic iron oxide (SPIO) particles in vitro on the cell surface by quantitative magnetic resonance (MR) imaging and to compare with flow cytometry. MATERIALS AND METHODS The monoclonal mouse antibody T84.1 and an appropriate IgG isotype antibody were conjugated to dextran-coated SPIO particles. HT29 cells expressing carcinoembryonic antigen (CEACAM5) were treated with antibody-conjugated SPIO particles. Purified cell samples were examined on a 3.0-T MR scanner using a multi-echo spin-echo sequence for MR relaxometry. Aliquots of the cell samples were further treated with a fluorescein isothiocyanate (FITC) anti-dextran antibody and an Alexa Fluor 488 anti-mouse antibody for the corresponding flow cytometry. RESULTS MR relaxometry revealed a dose-dependent binding of T84.1-conjugated SPIO particles with a positive correlation between R(2) relaxation rate of cell samples and SPIO particle concentration during incubation (r=0.993, P<.01). Positive correlations were also observed between R(2) relaxation rate and flow cytometry (geometric mean) with both fluorescent antibodies (r=0.972 and r=0.953, both P<.01), respectively. CONCLUSION The study revealed the feasibility of quantitative MR imaging of targeted SPIO particles on the cell surface comparable to flow cytometry.

AXL BLOCKADE BY BGB324 INHIBITS BCR-ABL TYROSINE KINASE INHIBITOR-SENSITIVE AND -RESISTANT CHRONIC MYELOID LEUKEMIA

Purpose: BCR-ABL kinase inhibitors are employed successfully for chronic myeloid leukemia (CML) treatment. However, resistant disease and persistence of BCR-ABL1–independent leukemia stem and progenitor cells (LSPC) remain clinical challenges. The receptor tyrosine kinase Axl can mediate survival and therapy resistance of different cancer cells. We investigated the therapeutic potential of Axl inhibition in CML. Experimental Design: We used primary cells from patients with CML and TKI-sensitive and -resistant BCR-ABL1+ CML cell lines and a novel ponatinib-resistant cell line KCL-22 PonR. We analyzed the effects of genetic and pharmacologic Axl blockade by the small-molecule Axl inhibitor BGB324 in vitro and in vivo. In BCR-ABL1–unmutated cells, we also investigated BGB324 in combination with imatinib. Results: We demonstrate overexpression of Axl receptor tyrosine kinase in primary cells of patients with CML compared with healthy individuals and a further increase of Axl expression in BCR-ABL TKI-resistant patients. We show that Axl blockage decreased growth of BCR-ABL TKI-sensitive CML cells including CD34+ cells and exerts additive effects with imatinib via inhibition of Stat5 activation. BGB324 also inhibits BCR-ABL TKI-resistant cells, including T315I-mutated and ponatinib-resistant primary cells. BGB324 exerted therapeutic effects in BCR-ABL1 T315I-mutated and ponatinib-resistant preclinical mouse models. Notably, BGB324 does not inhibit BCR-ABL1 and consequently inhibits CML independent of BCR-ABL1 mutational status. Conclusions: Our data show that Axl inhibition has therapeutic potential in BCR-ABL TKI-sensitive as well as -resistant CML and support the need for clinical trials. Clin Cancer Res; 23(9); 2289–300. ©2016 AACR.

Targeting the Gremlin–VEGFR2 axis – a promising strategy for multiple diseases?†

Published recently in the Journal of Pathology, Lavoz et al. show that Gremlin promotes renal inflammation directly via VEGFR2. As Gremlin has been implicated in many other diseases, such as heart, lung and liver fibrosis, osteogenesis, angiogenesis and cancer, the new findings provide a rationale for novel concepts to investigate and potentially treat several pathologies. Copyright