Johann Schoenberger

Blockade of Neoangiogenesis, a New and Promising Technique to Control the Growth of Malignant Tumors and their Metastases

Neoangiogenesis, the development of new blood vessels from a pre-existing vasculature involves the migration behavior, proliferation and differentiation of endothelial cells. This process is an important aspect to both the growth and metastasis of solid tumors. In growing tumors, the development of a sufficient vascular supply is a highly complex event, which is controlled by many factors. An interaction between tumor cells, endothelial cells, macrophages, fibroblasts, and the extracellular matrix, all of which are capable of releasing factors influencing the angiogenic mechanisms is necessary for the building of new sufficient vascular structures. The increased understanding of the balance of angiogenesis and mechanisms of vascular control allowed the development of new therapeutic substances that influence that process in different ways. For example, work done over the last decade has elucidated the central role of vascular endothelial growth factor (VEGF) in the regulation of normal and pathological angiogenesis. Therefore, the blockade of VEGF signaling is currently a major part of strategies for the therapy of malignant tumors. Several studies tested neutralizing antibodies against mature VEGF protein or its isoforms, blockade of VEGF receptors by VEGF receptor antibodies, soluble VEGF receptor mutants or fusion-proteins and finally, intracellular interference with VEGF mRNA or signaling in the target cell. In this review, we present strategies and substances for the blockade of neoangiogenesis that are already in clinical use or in pre-clinical trials. Also the combination of these approaches with conventional chemotherapy or radiotherapy will be discussed.

Innovative Strategies in In Vivo Apoptosis Imaging

Apoptosis (programmed cell death) plays a key role in the pathogenesis of many disorders including cerebral and myocardial ischemia, autoimmune and neurodegenerative diseases, infections, organ and bone marrow transplant rejection, and tumor response to chemotherapy and/or radiotherapy. Apoptosis in itself represents a complex mechanism where numerous (pro-apoptotic and anti-apoptotic) molecules interact in an elaborate manner. Since the original description by Kerr et al. in 1972, clinical assessment of apoptosis has always required biopsies or aspirated material for in vitro investigations. Several well-established methods are available for in vitro tests using tissue specimens. However, a non-invasive detection of apoptosis would be of great benefit for many patients in various situations. Today, non-invasive techniques for direct in vivo detection of apoptotic cells are rare and urgently need improvement. The early in vivo detection of apoptotic cells can provide the physician with important information to develop further therapeutic strategies in chemotherapy or radiotherapy of tumors, in transplantation of organs, or in healing of infarct areas. In some preliminary publications, several authors reported on the in vivo use of caspase-inhibitors and annexin V, labeled with indium-111, technetium-99m, iodine-123, iodine-124 or fluoride-18. In the present paper, we review the current applicability of both techniques for in vivo apoptosis imaging, and discuss the methodical problems.

Radiolabeled annexin V for imaging apoptosis in radiated human follicular thyroid carcinomas — is an individualized protocol necessary?

INTRODUCTION Induction of apoptosis is a widely used strategy for cancer therapy, but evaluating the degree and success of this therapy still poses a problem. Radiolabeled annexin V has been proposed to be a promising candidate for detecting apoptotic cells in tumors following chemotherapy in vivo. In order to see whether radiolabeled annexin V could be a suitable substance for the noninvasive in vivo detection of apoptosis in thyroid tissue and to establish an optimized study protocol, we investigated two poorly differentiated thyroid carcinoma cell lines: ML-1 and FTC-133. METHODS Apoptosis was evaluated before as well as 2 and 4 days after in vitro irradiation with 30 Gy X-rays. In this study, binding of FITC- and of (125)I-labeled annexin V was measured in comparison to other apoptosis markers such as Bax, caspase-3 and Fas, which were determined by flow cytometry and Western blot analysis with densitometric evaluation. RESULTS ML-1 and FTC-133 cells showed a significant increase in annexin V binding 48 h after irradiation. Ninety-six hours after irradiation, the annexin V absorption capability of ML-1 cells was still maximal, while the living fraction of FTC-133 increased significantly. The amount of caspase-3 and Bax was clearly increased 48 h after irradiation and had normalized after 96 h in both cell lines. Fas protein concentrations remained unchanged in ML-1 cells but were significantly enhanced in FTC-133 cells. CONCLUSION The binding of FITC- and (125)I-labeled annexin V showed a significant accordance. A reliable evaluation of apoptosis induced by radiotherapy in thyroid tumors was possible 48 h after irradiation, when binding of radiolabeled annexin V is most significantly enhanced. Using two poorly differentiated cell lines of thyroid carcinoma, one may expect to find a nearly similar response to external irradiation. In contrast, the cell lines showed a completely contrary response. However, an individualized study protocol for each type of tumor and probably within each type is necessary.

Evaluation of the clinical value of bone metabolic parameters for the screening of osseous metastases compared to bone scintigraphy

BackgroundBone metastases are common in many types of cancer. As screening methods different imaging modalities are available. A new approach for the screening of osseous metastases represents the measurement of bone metabolic markers. Therefore aim of this study was to evaluate the usefulness of the determination of bone metabolic markers aminoterminal propeptide of type I procollagen (PINP, osteoblastic activity) and the carboxyterminal pyridinoline cross-linked telopeptide of type I collagen (ICTP, osteoclastic activity) for the detection of bone metastases associated with other malignancies.Methods88 patients aged 21 – 82 years with malignant tumors were prospectively studied. The serum concentrations of PINP and ICTP were measured and compared to the results of bone scintigraphy, radiological bone series, CT, MRI and clinical follow-up.ResultsOsseous metastases were found in 21 patients. 19 of them were correctly identified by bone scintigraphy (sensitivity: 90%). For bone metabolic markers results were as follows: ICTP sensitivity: 71%, specificity: 42%; PINP sensitivity: 24%, specificity: 96%.ConclusionsAs markers of bone metabolism PINP and ICTP showed low sensitivity and/or specificity for the detection of osseous metastases. The presented markers did not seem to be sufficient enough to identify patients with bone metastases or to replace established screening methods.

Changes of Apoptosis, p53, and bcl-2 by Irradiation in Poorly Differentiated Thyroid Carcinoma Cell Lines: A Prognostic Marker for the Prospect of Therapeutic Success?

BACKGROUND Poorly differentiated thyroid carcinoma (PDTC) has an unfavorable prognosis. Surgical management is the principal treatment approach. In addition, radioiodine treatment and external beam radiotherapy (EBRT) are given to reduce the risk of local relapse. Despite aggressive therapy, the response to treatment tends to become increasingly poorer over time. The objective of this study was to investigate the induction of apoptosis by EBRT as a function of p53 and bcl-2 protein levels in PDTC. The predictive value of these molecules with respect to treatment efficacy was evaluated. MATERIALS AND METHODS Two different cell lines of PDTC (FTC-133 and ML-1) were irradiated with a dose of 30 Gy. Apoptotic cells were quantified using terminal deoxynucleotidyltransferase-dUTP nick-end labeling staining without irradiation, 48 and 96 hours after irradiation. The protein levels of p53 and bcl-2 were measured simultaneously using flow cytometry and western blotting. The cell cycle distribution was determined. RESULTS Untreated FTC-133 cells showed a high rate of apoptosis, a high protein level of p53, and a low bcl-2 protein level. Forty-eight hours after irradiation, a slight reduction in apoptotic cells was observed in conjunction with an increase in bcl-2 and p53 protein levels. The slightly reduced fraction of apoptotic cells remained at the same level up to 96 hours after irradiation, whereas the p53 protein level was further downregulated. The cell cycle distribution showed a significant G2/M arrest after 48 hours and recovery 96 hours after irradiation. ML-1 cells did not show any detectable p53 levels and revealed a low rate of apoptosis which significantly increased 48 hours after irradiation. Ninety-six hours after irradiation, a decrease in apoptosis was detectable. The protein level of bcl-2 increased significantly within 48 hours and decreased 96 hours after irradiation. The cell cycle distribution showed a G2/M arrest after 48 hours without a recovery 96 hours after irradiation. CONCLUSIONS The p53 and bcl-2 expression profiles and the observed apoptotic rates of FTC-133 and ML-1 under irradiation are consistent with a more aggressive FTC-133 phenotype. Alterations in p53- and bcl-2 protein levels yield predictive information for EBRT efficacy.