Dovitinib in patients with gastrointestinal stromal tumour refractory and/or intolerant to imatinib.

BACKGROUND: This multicentre phase II trial (DOVIGIST) evaluated the antitumour activity of dovitinib as second-line treatment of patients with gastrointestinal stromal tumour (GIST) refractory to imatinib or who do not tolerate imatinib. METHODS: Patients received oral dovitinib 500 mg day-1, 5 days on/2 days off, until GIST progression or unacceptable toxicity, with an objective to evaluate efficacy, assessed as the disease control rate (DCR) at 12 weeks. Tumour assessment and response to dovitinib therapy were evaluated by Response Evaluation Criteria In Solid Tumours (RECIST v1.1) and the Choi criteria. Secondary objectives included assessment of progression-free survival (PFS), safety and tolerability, and DCR at the end of treatment. RESULTS: Thirty-eight of the 39 patients enrolled had histologically confirmed GIST. The DCR at 12 weeks was 52.6% (90% confidence interval (CI), 38.2-66.7%) meeting the preset efficacy criterion for the primary end point. The objective response rate (complete response+partial response) was 2.6% (1 of 38; 90% CI, 0.1-11.9%), and 5.3% (n=2; 90% CI, 0.9-15.7%) at the end of the study. The median PFS was 4.6 months (90% CI, 2.8-7.4 months). Dose interruption was required in 26 patients (66.7%), of which 18 (69.2%) were due to adverse events. The most frequently observed grade 3 adverse events included hypertension (n=7), fatigue (n=5), vomiting (n=4), hypertriglyceridaemia (n=4), and γ-glutamyltransferase increase (n=4). CONCLUSIONS: Dovitinib is an active treatment for patients with GIST who are intolerant to imatinib or whose GIST progresses on imatinib.

A pro-inflammatory signature mediates FGF2-induced angiogenesis.

Fibroblast growth factor-2 (FGF2) is a potent angiogenic growth factor. Here, gene expression profiling of FGF2-stimulated microvascular endothelial cells revealed, together with a prominent pro-angiogenic profile, a pro-inflammatory signature characterized by the upregulation of pro-inflammatory cytokine/chemokines and their receptors, endothelial cell adhesion molecules and members of the eicosanoid pathway. Real-time quantitative PCR demonstrated early induction of most of the FGF2-induced, inflammation-related genes. Accordingly, chick embryo chorioallantoic membrane (CAM) and murine Matrigel plug angiogenesis assays demonstrated a significant monocyte/macrophage infiltrate in the areas of FGF2-driven neovascularization. Similar results were obtained when the conditioned medium (CM) of FGF2-stimulated endothelial cells was delivered onto the CAM, suggesting that FGF2-upregulated chemoattractants mediate the inflammatory response. Importantly, FGF2-triggered new blood vessel formation was significantly reduced in phosphatidylinositol 3-kinase-gamma null mice exhibiting defective leucocyte migration or in clodronate liposome-treated, macrophage-depleted mice. Furthermore, the viral pan-chemokine antagonist M3 inhibited the angiogenic and inflammatory responses induced by the CM of FGF2-stimulated endothelial cells and impaired FGF2-driven neovascularization in the CAM assay. These findings point to inflammatory chemokines as early mediators of FGF2-driven angiogenesis and indicate a non-redundant role for inflammatory cells in the neovascularization process elicited by the growth factor.

Anti-FGF2 approaches as a strategy to compensate resistance to anti-VEGF therapy: long-pentraxin 3 as a novel antiangiogenic FGF2-antagonist.

Angiogenesis, the formation of new blood vessels from the endothelium of the existing vasculature, plays a pivotal role in tumor growth, progression and metastasis. Over the last 30 years, numerous pro- and antiangiogenic molecules, their ligands, and intracellular signaling pathways have been identified, and significant efforts have been undertaken to develop antiangiogenic strategies for cancer therapy. Agents that selectively target vascular endothelial growth factor (VEGF) and its receptors have shown promising activity in clinical trials and have been approved for use in selected cancer indications. However, patients may ultimately develop resistance to these drugs. One proposed mechanism of tumor escape from anti-VEGF therapy is the up-regulation of fibroblast growth factor-2 (FGF2). FGF2 is a pleiotropic, angiogenesis inducer belonging to the family of the heparin-binding FGF growth factors. FGF2 is expressed by numerous tumor types and exerts its proangiogenic activity by interacting with tyrosine kinase receptors, heparan-sulfate proteoglycans, and integrins expressed on the endothelial cell surface. Experimental evidence suggests that targeting FGF2, in addition to VEGF, might provide synergistic effects in the treatment of angiogenesis-related diseases, including cancer. Several FGF2 inhibitors, with different chemical structure and mechanism of action, have been identified. Recent observations have shown the ability of the soluble pattern recognition receptor long-pentraxin-3 (PTX3) to bind FGF2, thus acting as a FGF2 antagonist. PTX3 binds FGF2 with high affinity and specificity. This interaction prevents the binding of FGF2 to its cognate tyrosine kinase receptors, leading to inhibition of the angiogenic activity of the growth factor. Further, preliminary observations support the hypothesis that PTX3 may inhibit FGF2-mediated tumor angiogenesis and growth. The identification of the FGF2-binding domain in the unique N-terminal extension of PTX3 has allowed the design of PTX3-derived synthetic peptides endowed with significant antiangiogenic activity in vitro and in vivo. These findings may provide the basis for the development of novel antiangiogenic FGF2 antagonists, with potential implications for cancer therapy.

Role of the soluble pattern recognition receptor PTX3 in vascular biology.

Pentraxins act as soluble pattern recognition receptors with a wide range of functions in various pathophysiological conditions. The long-pentraxin PTX3 shares the C-terminal pentraxin-domain with short-pentraxins C-reactive protein and serum amyloid P component and possesses an unique N-terminal domain. These structural features suggest that PTX3 may have both overlapping and distinct biological/ligand recognition properties when compared to short-pentraxins. PTX3 serves as a mechanism of amplification of inflammation and innate immunity. Indeed, vessel wall elements produce high amounts of PTX3 during inflammation and the levels of circulating PTX3 increase in several pathological conditions affecting the cardiovascular system. PTX3 exists as a free or extracellular matrix-associated molecule and it binds the complement fraction C1q. PTX3 binds also apoptotic cells and selected pathogens, playing a role in innate immunity processes. In endothelial cells and macrophages, PTX3 upregulates tissue factor expression, suggesting its action as a regulator of endothelium during thrombogenesis and ischaemic vascular disease. Finally, PTX3 binds the angiogenic fibroblast growth factor-2, thus inhibiting its biological activity. Taken together, these properties point to a role for PTX3 during vascular damage, angiogenesis, atherosclerosis, and restenosis.

Identification of an antiangiogenic FGF2-binding site in the N terminus of the soluble pattern recognition receptor PTX3.

Long-pentraxin 3 (PTX3) is a soluble pattern recognition receptor with non-redundant functions in inflammation and innate immunity. PTX3 comprises a pentraxin-like C-terminal domain involved in complement activation via C1q interaction and an N-terminal extension with unknown functions. PTX3 binds fibroblast growth factor-2 (FGF2), inhibiting its pro-angiogenic and pro-restenotic activity. Here, retroviral transduced endothelial cells (ECs) overexpressing the N-terminal fragment PTX3-(1-178) showed reduced mitogenic activity in response to FGF2. Accordingly, purified recombinant PTX3-(1-178) binds FGF2, prevents PTX3/FGF2 interaction, and inhibits FGF2 mitogenic activity in ECs. Also, the monoclonal antibody mAb-MNB4, which recognizes the PTX3-(87-99) epitope, prevents FGF2/PTX3 interaction and abolishes the FGF2 antagonist activity of PTX3. Consistently, the synthetic peptides PTX3-(82-110) and PTX3-(97-110) bind FGF2 and inhibit the interaction of FGF2 with PTX3 immobilized to a BIAcore sensor chip, FGF2-dependent EC proliferation, and angiogenesis in vivo. Thus, the data identify a FGF2-binding domain in the N-terminal extension of PTX3 spanning the PTX3-(97-110) region, pointing to a novel function for the N-terminal extension of PTX3 and underlining the complexity of the PTX3 molecule for modular humoral pattern recognition.

Pentraxin 3 inhibits fibroblast growth factor 2-dependent activation of smooth muscle cells in vitro and neointima formation in vivo.

OBJECTIVE: The fibroblast growth factor (FGF)/FGF receptor system plays an important role in smooth muscle cell (SMC) activation. Long-pentraxin 3 (PTX3) is a soluble pattern recognition receptor with non-redundant functions in inflammation and innate immunity. PTX3 is produced by different cell types of the vessel wall, including SMCs. PTX3 binds FGF2 and inhibits its angiogenic activity on endothelial cells. We investigated the capacity of PTX3 to affect FGF2-dependent SMC activation in vitro and in vivo. METHODS AND RESULTS: When added to human coronary artery SMCs, human PTX3 inhibits cell proliferation driven by endogenous FGF2 and the mitogenic and chemotactic activity exerted by exogenous recombinant FGF2. Accordingly, PTX3 prevents (125)I-FGF2 interaction with FGF receptors on the same cells. Also, PTX3 overexpression after recombinant adeno-associated virus-PTX3 gene transfer inhibits human coronary artery SMC proliferation and survival promoted by FGF2 in vitro. Consistently, a single local endovascular injection of recombinant adeno-associated virus-PTX3 gene inhibits intimal thickening after balloon injury in rat carotid arteries. CONCLUSIONS: PTX3 is a potent inhibitor of the autocrine and paracrine stimulation exerted by FGF2 on SMCs. Local PTX3 upregulation may modulate SMC activation after arterial injury.

Antiangiogenic activity of semisynthetic biotechnological heparins: low-molecular-weight-sulfated Escherichia coli K5 polysaccharide derivatives as fibroblast growth factor antagonists.

OBJECTIVE: Low-molecular-weight heparin (LMWH) exerts antitumor activity in clinical trials. The K5 polysaccharide from Escherichia coli has the same structure as the heparin precursor. Chemical and enzymatic modifications of K5 polysaccharide lead to the production of biotechnological heparin-like compounds. We investigated the fibroblast growth factor-2 (FGF2) antagonist and antiangiogenic activity of a series of LMW N,O-sulfated K5 derivatives. METHODS AND RESULTS: Surface plasmon resonance analysis showed that LMW-K5 derivatives bind FGF2, thus inhibiting its interaction with heparin immobilized to a BIAcore sensor chip. Interaction of FGF2 with tyrosine-kinase receptors (FGFRs), heparan sulfate proteoglycans (HSPGs), and alpha(v)beta3 integrin is required for biological response in endothelial cells. Similar to LMWH, LMW-K5 derivatives abrogate the formation of HSPG/FGF2/FGFR ternary complexes by preventing FGF2-mediated attachment of FGFR1-overexpressing cells to HSPG-bearing cells and inhibit FGF2-mediated endothelial cell proliferation. However, LMW-K5 derivatives, but not LMWH, also inhibit FGF2/alpha(v)beta3 integrin interaction and consequent FGF2-mediated endothelial cell sprouting in vitro and angiogenesis in vivo in the chick embryo chorioallantoic membrane. CONCLUSIONS: LMW N,O-sulfated K5 derivatives affect both HSPG/FGF2/FGFR and FGF2/alpha(v)beta3 interactions and are endowed with FGF2 antagonist and antiangiogenic activity. These compounds may provide the basis for the design of novel LMW heparin-like angiostatic compounds.

Selective recognition of fibroblast growth factor-2 by the long pentraxin PTX3 inhibits angiogenesis.

The long pentraxin PTX3 is a soluble pattern recognition receptor produced by monocytes and endothelial cells that plays a nonredundant role in inflammation. Several pathologic conditions are characterized by local production of both PTX3 and the angiogenic fibroblast growth factor-2 (FGF2). Here, solid-phase binding assays demonstrated that PTX3 binds with high affinity to FGF2 but not to a panel of cytokines and growth factors, including FGF1, FGF4, and FGF8. Accordingly, PTX3 prevented (125)I-FGF2 binding to endothelial cell receptors, leading to specific inhibition of FGF2-induced proliferation. PTX3 hampered also the motogenic activity exerted by endogenous FGF2 on a wounded endothelial cell monolayer. Moreover, PTX3 cDNA transduction in FGF2-transformed endothelial cells inhibited their autocrine FGF2-dependent proliferation and morphogenesis in vitro and their capacity to generate vascular lesions when injected in nude mice. Finally, PTX3 suppressed neovascularization triggered by FGF2 in the chick embryo chorioallantoic membrane with no effect on physiologic angiogenesis. In contrast, the short pentraxin C-reactive protein was a poor FGF2 ligand/antagonist. These results establish the selective binding of a member of the pentraxin superfamily to a growth factor. PTX3/FGF2 interaction may modulate angiogenesis in various physiopathologic conditions driven by inflammation, innate immunity, and/or neoplastic transformation.

Inhibition of intra- and extra-cellular Tat function and HIV expression by pertussis toxin B-oligomer.

HIV-1-transactivating factor Tat contributes to virus replication and to the onset of AIDS-associated pathologies by targeting different infected and uninfected cell types. We previously demonstrated that the B-oligomer of pertussis toxin (PTX-B) inhibits HIV infection and replication in primary T cells and macrophages and Tat-dependent HIV-1 long terminal repeat (LTR) transactivation inT lymphoid Jurkat cells. Here we demonstrate that PTX-B inhibits Tat-dependent NF-kappaB activation and HIV-1 LTR-transactivation in non-permissive epithelial HL3T1 cells in a phosphatidylinositol 3'-kinase-dependent way. PTX-B exerts its inhibition both when Tat is produced endogenously in transfected cells and in cells incubated with the extracellular Tat protein. In this latter case, PTX-B does not interfere with extracellular Tat uptake by cells. PTX-B inhibited also interleukin-8 secretion and virus expression stimulated in chronically infected U1 promonocytic cells by intra- and/or extracellular Tat. The genetically modified holotoxin PT-9 K/129G retains the capacity to inhibit Tat transactivating activity and HIV replication in both HIV-permissive and non-permissive cells. Inconclusion, PTX-B acts as a "pleiotropic" inhibitor of Tat, and this may significantly contribute to the broad spectrum of anti-HIV-1 effects exerted by PTX-B in different cell types, and suggests PTX-B and its derivatives as prototypic for the development of anti-Tat drugs.