Receptor tyrosine kinase inhibitors: Are they real tumor killers?

Inhibiting tumor growth by targeting the tumor vasculature was first proposed by Judah Folkman almost 40 years ago. Since then, different approaches and numerous drugs and agents have been developed to achieve this goal, either with the aim of inhibiting tumor neoangiogenesis or normalizing the tumor vasculature. Among the most promising therapeutic targets are receptor tyrosine kinases (RTKs), some of which are predominantly expressed on tumor endothelial cells, although they are sometimes also present on tumor cells. The majority of RTK inhibitors investigated over the past two decades competes with ATP at the active site of the kinase and therefore block the phosphorylation of intracellular targets. Some of these drugs have been approved for therapy, whereas others are still in clinical trials. Here, we discuss the scientific basis, current status, problems and future prospects of RTK inhibition in anti-tumor therapy.

The inhibition of tyrosine kinase receptor signalling in leiomyosarcoma cells using the small molecule kinase inhibitor PTK787/ZK222584 (Vatalanib®).

Leiomyosarcomas remain challenging tumors to manage and novel therapy strategies besides radiation and conventional chemotherapy are needed. Targeting angiogenesis by inhibition of vascular endothelial growth factor (VEGF) receptor tyrosine kinases (RTKs) of the tumor vasculature with small molecules is a promising new therapy. It has been shown recently that these receptors are not only expressed on tumor endothelium but also on tumor cells themselves. Thus, we investigated the expression of members of the VEGF receptor (VEGFR) family and corresponding growth factors in leiomyosarcoma tissue specimens and in the leiomyosarcoma cell lines SK-LMS-1 and SK-UT-1. We evaluated the influence of the VEGFR inhibitor PTK787/ZK222584 (PTK787) on cell growth, migration, apoptosis and phosphorylation of intracellular signalling molecules. In human leiomyosarcoma tissue specimens VEGFR­1/-2 and platelet-derived growth factor receptor (PDGFR-ß) were strongly expressed. Both leiomyosarcoma cell lines expressed VEGFR­1/-3 and PDGFR-ß but VEGFR-2 protein expression was positive only in SK-UT-1. SK-LMS-1 and SK-UT-1 cells secreted high and low amounts of VEGF-A, respectively, whereas PDGF-BB secretion was similar in both cell lines. Application of PTK787 led to partial inhibition of PDGF-BB-activated AKT/p90RSK and ERK1/2 signalling pathways. In contrast, protein phosphorylation was not affected by PTK787 in VEGF-A-treated cells. PTK787 turned out to inhibit cell migration even though no effects were observed upon stimulation with VEGF-A or PDGF-BB. In line, cell growth in leiomyosarcoma cell lines remained unchanged upon PTK787 treatment alone and with subsequent VEGF-A- or PDGF-BB-stimulation. However, VEGF-A, but not PDGF-BB-treated cells showed increased cell death upon PTK787 treatment. VEGFR family members are expressed in leiomyosarcomas in vivo and in vitro. Upon receptor stimulation, PTK787 is able to inhibit subsequent phosphorylation events and influences cell survival but not metabolic activity and migration. Thus, the inhibitor is possibly an additional option in the treatment of leiomyosarcomas.

Increased angiogenesis and FGFR protein expression indicate a favourable prognosis in bladder cancer.

Compared to other members of the fibroblast growth factor receptor (FGFR) family, only few studies investigate FGFR3 in tumour angiogenesis. We investigated the connection between angiogenesis and FGF/FGFR expression including FGFR3 mutation status in urothelial carcinomas. Immunohistochemistry was performed in invasive and non-invasive urothelial cancers of 61 patients. Protein expression of CD31, factor VIII (FVIII), FGF-1/2, FGFR1, FGFR3 and FGFR4 and FGFR3 mutation status were evaluated. Morphometric assessment of angiogenesis including microvessel count (MVC) and vascular surface area (VSA) was analysed. Correlation and survival analyses (overall survival (OS) and disease-free survival (DFS)) with univariate and multivariate analyses were performed. CD31 values (MVC and VSA) significantly correlated with OS and DFS. OS and DFS were significantly better in patients with FGFR3 overexpression. Multivariate analysis revealed FGFR3 protein expression and tumour grading (WHO classification 2004) as independent prognostic factors of OS and VSA of CD31 and FGFR3 protein expression of DFS. FGFR3 mutation status was correlated with VSA measured by FVIII. FGFR3 may be able to induce a pro-angiogenic phenotype in urothelial carcinomas and significantly influence prognosis. Consequently, FGFR3 is a potential therapeutic target also from the angiogenesis perspective.

Microvessel density and VEGF/VEGF receptor status and their role in sarcomas of the pulmonary artery.

Neoangiogenesis, driven by a variety of angiogenic factors, plays an essential role during development and progression of malignant tumors. Vascular endothelial growth factor (VEGF) and its receptors have been designated a central part in the angiogenic process during malignancy. We studied the vascular parameters by means of morphology and morphometry in 7 sarcomas of the pulmonary artery (SPA) and 10 poorly differentiated leiomyosarcomas of soft tissue. Immunohistochemical analysis of VEGF and VEGFR was related to survival and prognosis. The microvessel density (MVD) and intervascular distances (IVD) differed significantly only at sites of necrosis compared to non-necrotic areas in SPA but not for soft tissue leiomyosarcomas. MVD, IVD and vascular surface area (VSA) revealed no difference between SPA and leiomyosarcomas of different origin. We found a more pronounced expression of VEGF in most tumors at sites of necrosis. The receptors were present in a subset of tumor vessels mostly at the tumor border. VEGFR-2 expression was also seen in a subset of tumor cells whereas VEGFR-1 showed only weak expression in some tumors. Local hypoxia seems to induce a higher MVD and a lower IVD at sites of necrosis compared to those areas without necrosis. The presence of necrosis in both sarcoma groups was correlated with the presence of VEGF due to local tumor hypoxia and subsequent up-regulation of VEGFR-2 and VEGFR-1 in tumor vessels as well as tumor cells. Overall and relapse-free survival showed no difference concerning all examined parameters. Thus, microvessel density does not seem to be a prognostic factor in SPA and other sarcomas.