The performance of three oncogeriatric screening tools - G8, optimised G8 and CARG - in predicting chemotherapy-related toxicity in older patients with cancer. A prospective clinical study.

BACKGROUND: Older patients are vulnerable to chemotherapy-related toxicity (CRT). Therefore we evaluated screening tools in their power to predict CRT. METHODS: Patients with cancer aged ≥65 years completed three screening questionnaires (G8, optimised G8 and Cancer and Ageing Research Group (CARG). Additionally, Comprehensive geriatric assessment (CGA) for verification of supportive care needs was undertaken on patients with impaired G8 scores. During chemotherapy treatment patients were assessed, capturing grade 0-5 CRT as defined by NCI CTCAE 4. RESULTS: 104 patients with non-haematological cancers were included at three study sites. Median age was 73 years (range 65-85). Onco-geriatric screening detected 74% as impaired using G8 and optimised G8 questionnaires and 86% using CARG screening. Grade 3-5 toxicity affected 64.4% of all patients. G8 (OR 0.3 95% CI [0.1;1.0]) and optimised G8 (OR 0.4 95% CI [0.1; 1.5]) did not reliably predict CRT, whereas screening with CARG demonstrated a strong prediction of severe CRT: OR 4.2, 95% CI [1.1, 15.9]. CGA was undertaken on 66 patients, revealing deficiencies in nutritional (83%) and functional-status (54%) and occurrence of relevant comorbidity (53%). CONCLUSION: The CARG tool could be useful for predicting CRT. CGA showed clinically relevant supportive care needs in patients with a positive G8 screening.

Tie2 as a novel key factor of microangiopathy in systemic sclerosis.

BACKGROUND: The angiopoietin(Ang)/Tie2 system is a key regulator of vascular biology. The expression of membrane bound (mb) Tie2 and Ang-1 ensures vessel stability, whereas Ang-2, inducible by vascular endothelial growth factor (VEGF), hypoxia, and inflammation, acts as an antagonist. Tie2 signalling is also attenuated by soluble Tie2 (sTie2), the extracellular domain of the receptor, which is shed upon stimulation with VEGF. Herein, we investigate the role of Ang/Tie2 in the peripheral vasculopathy in systemic sclerosis (SSc) including animal models. METHODS: The expression of Ang-1/-2 and Tie2 in skin/serum of SSc patients was compared with healthy controls by immunohistochemistry (IHC)/ELISA. Expression of Ang/Tie2 was analysed in different animal models: VEGF transgenic (tg) mice, hypoxia model, bleomycin-induced skin fibrosis, and tight skin 1 (TSK1) mice. RESULTS: In SSc, dermal microvessels abundantly expressed Ang-2, but not Ang-1 compared with healthy controls. The percentage of mbTie2+ microvessels was profoundly decreased whereas the levels of sTie2 were increased already in early disease. Both in skin and sera of SSc patients, the Ang1/2 ratio was reduced, being lowest in patients with digital ulcers indicating vessel destabilizing conditions. We next studied potential influencing factors in animal models. The VEGF tg mouse model, the hypoxia, and the inflammation-dependent bleomycin model all showed a similar dysregulation of Ang/Tie2 as in SSc, which did not apply for the non-inflammatory TSK1 model. CONCLUSION: Peripheral microvasculopathy in SSc results from a complex dysregulation of angiogenic signalling networks including the VEGF and the Ang/Tie2 system. The profoundly disturbed Ang-/Tie-2 balance might represent an important target for vascular therapeutic approaches in SSc.

Trichostatin A prevents the accumulation of extracellular matrix in a mouse model of bleomycin-induced skin fibrosis.

OBJECTIVE: Tissue fibrosis is a hallmark compromising feature of many disorders. In this study, we investigated the antifibrogenic effects of the histone deacetylase inhibitor trichostatin A (TSA) on cytokine-driven fibrotic responses in vitro and in vivo. METHODS: Skin fibroblasts from patients with systemic sclerosis (SSc) and normal healthy control subjects were stimulated with profibrotic cytokines in combination with TSA. Human Colalpha1(I) and fibronectin were measured using real-time polymerase chain reaction, and levels of soluble collagen were estimated using the SirCol collagen assay. Electromobility shift assay and confocal fluorescence microscopy were used to investigate the intracellular distribution of Smad transcription factors. For in vivo analysis, skin fibrosis was quantified by histologic assessment of mouse skin tissue in a model of bleomycin-induced fibrosis. RESULTS: Reductions in the cytokine-induced transcription of Colalpha1(I) and fibronectin were observed in both normal and SSc skin fibroblasts following the addition of TSA. Similarly, the expression of total collagen protein in TSA-stimulated SSc skin fibroblasts was reduced to basal levels. The mechanism of action of TSA included inhibition of the nuclear translocation and DNA binding of profibrotic Smad transcription factors. Western blot analysis revealed an up-regulation of the cell cycle inhibitor p21 by TSA, leading to reduced proliferation of fibroblasts. In addition, in bleomycin-induced fibrosis in mice, TSA prevented dermal accumulation of extracellular matrix in vivo. CONCLUSION: These findings provide novel insights into the epigenetic regulation of fibrosis. TSA and similar inhibitory compounds appear to represent early therapeutic strategies for achieving reversal of the cytokine-driven induction of matrix synthesis that leads to fibrosis.

Histone deacetylase/acetylase activity in total synovial tissue derived from rheumatoid arthritis and osteoarthritis patients.

OBJECTIVE: Rheumatoid arthritis (RA) is a chronic inflammatory disorder of unknown origin. Histone deacetylase (HDA) activity is considered to play a major role in the transcriptional regulation of proinflammatory genes. We undertook this study to investigate the balance of histone acetylase and HDA activity in synovial tissue from RA patients compared with that from patients with osteoarthritis (OA) and normal controls. METHODS: Activity of histone acetylases and HDAs was measured in nuclear extracts of total synovial tissue samples, which were obtained from RA and OA patients undergoing surgical joint replacement, and compared with the activity in synovial tissues from patients without arthritis. Tissue expression of HDAs 1 and 2 was quantified by Western blotting. In addition, immunohistochemistry was performed for HDA-2. RESULTS: Mean+/-SEM HDA activity in synovial tissue samples derived from patients with RA was measured as 1.5+/-0.3 micromoles/microg, whereas the activity levels in OA (3.2+/-0.7 micromoles/microg) and normal (7.1+/-4.2 micromoles/microg) synovial tissue samples were significantly higher. Histone acetylase activity reached similar levels in RA and OA tissues and in normal tissues. The ratio of HDA activity to histone acetylase activity in RA synovial tissue was significantly reduced (12+/-2%) compared with that in OA synovial tissue (26+/-3%). The activity ratio in normal control samples was arbitrarily set at 100+/-40%. In addition, the tissue expression of HDA-1 and HDA-2 proteins was clearly lower in RA samples than in OA samples. CONCLUSION: The balance of histone acetylase/HDA activities is strongly shifted toward histone hyperacetylation in patients with RA. These results offer novel molecular insights into the pathogenesis of the disease that might be relevant to the development of future therapeutic approaches.

The role of membrane lipids in the induction of macrophage apoptosis by microparticles.

Microparticles are membrane-derived vesicles that are released from cells during activation or cell death. These particles can serve as mediators of intercellular cross-talk and induce a variety of cellular responses. Previous studies have shown that macrophages undergo apoptosis after phagocytosing microparticles. Here, we have addressed the hypothesis that microparticles trigger this process via lipid pathways. In these experiments, microparticles induced apoptosis in primary macrophage cells or cell lines (RAW 264.7 or U937) with up to a 5-fold increase. Preincubation of macrophages with phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)BP) reduced the microparticle-induced apoptosis in a dose-dependent manner. PtdIns(3,5)BP is a specific inhibitor of the acid sphingomyelinase and thus can block the generation of pro-apoptotic ceramides. Similarly, the pre-incubation of macrophages with PtdIns(3,5)BP prevented microparticle-induced upregulation of caspase 8, which is a major target molecule of ceramide action in the apoptosis pathway. PtdIns(3,5)BP, however, had no effect on the spontaneous rate of apoptosis. To evaluate further signaling pathways induced by microparticles, the extracellular signal regulated kinase (ERK-) 1 was investigated. This kinase plays a role in activating phospholipases A2 which cleaves membrane phospholipids into arachidonic acid; microparticles have been suggested to be a preferred substrate for phospholipases A2. As shown in our experiments, microparticles strongly increased the amount of phosphorylated ERK1/2 in RAW 264.7 macrophages in a time-dependent manner, peaking 15 min after co-incubation. Addition of PD98059, a specific inhibitor of ERK1, prevented the increase in apoptosis of RAW 264.7 macrophages. Together, these data suggest that microparticles perturb lipid homeostasis of macrophages and thereby induce apoptosis. These results emphasize the importance of biolipids in the cellular cross-talk of immune cells. Based on the fact that in clinical situations with excessive cell death such as malignancies, autoimmune diseases and following chemotherapies high levels of circulating microparticles might modulate phagocytosing cells, a suppression of the immune response might occur due to loss of macrophages.

Technology insight: gene transfer and the design of novel treatments for rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by systemic inflammation and joint destruction. Novel therapies have emerged during the past decade, marking a new era in the treatment of RA. Meanwhile, in vivo and in vitro gene-transfer studies have provided valuable insights into mechanisms of disease pathogenesis. Advanced gene-delivery techniques and animal models promise further progress in RA research and the development of novel therapeutic strategies for this disease. In this article we provide an overview of the wide spectrum of potential targets that have been identified so far, discuss currently available gene-transfer methods, and outline the barriers that need to be overcome for these approaches to be successfully applied in daily practice.

Ex vivo homeostatic proliferation of CD4+ T cells in rheumatoid arthritis is dysregulated and driven by membrane-anchored TNFalpha.

The systemic CD4(+) T cell compartment in patients with rheumatoid arthritis (RA) is characterized by TCR repertoire contraction, shortened telomere lengths, and decreased numbers of recent thymic emigrants, suggesting a disturbed CD4(+) T cell homeostasis. In mice, homeostatic proliferation of peripheral CD4(+) T cells is regulated by TCR interaction with self peptide-MHC complexes (pMHC) and can be reproduced in vitro. We have established an ex vivo model of homeostatic proliferation, in which self-replication of human CD4(+) T cells is induced by cell-cell contact with autologous monocytes. In healthy individuals, blockade of TCR-pMHC class II contact resulted in decreased CD4(+) T cell division. In contrast, homeostatic proliferation in RA patients was not inhibited by pMHC blockade, but increased during the initial culture period. The anti-TNF-alpha Ab cA2 inhibited homeostasis-driven ex vivo proliferation in healthy controls and in RA patients. In addition, treatment of RA patients with infliximab decreased the ex vivo rate of homeostatic proliferation of CD4(+) T cells. Our results suggest a disturbed regulation of CD4(+) T cell homeostasis leading to the repertoire aberrations reported in RA. Membrane-anchored TNF-alpha appears to be a cell-cell contact-dependent stimulus of homeostatic proliferation of CD4(+) T cells, possibly favoring self-replication of autoreactive CD4(+) T cells in patients with RA.