Antonino Spartà

Contribution of α4β1 integrin to the antiallergic effect of levocabastine

Levocabastine is an antiallergic drug acting as a histamine H1-receptor antagonist. In allergic conjunctivitis (AC), it may also antagonize up-regulation of the intercellular adhesion molecule-1 (ICAM-1) expressed on epithelial conjunctival cells. However, little is known about its effects on eosinophils, important effector cells in AC. The adhesion molecule integrin alpha(4)beta(1) is expressed in eosinophils; it interacts with the vascular cell adhesion molecule-1 (VCAM-1) and fibronectin (FN) in vascular endothelial cells and contributes to eosinophil activation and infiltration in AC. This study provides evidence that in a scintillation proximity assay levocabastine (IC(50) 406 microM), but not the first-generation antihistamine chlorpheniramine, displaced (125)I-FN binding to human integrin alpha(4)beta(1) and, in flow cytometry analysis, levocabastine antagonized the binding of a primary antibody to integrin alpha(4) expressed on the Jurkat cell surface. Levocabastine, but not chlorpheniramine, binds the alpha(4)beta(1) integrin and prevents eosinophil adhesion to VCAM-1, FN or human umbilical vascular endothelial cells (HUVEC) in vitro. Similarly, levocabastine affects alpha(L)beta(2)/ICAM-1-mediated adhesion of Jurkat cells. In a model of AC levocabastine eye drops reduced the clinical aspects of the late-phase reaction and the conjunctival expression of alpha(4)beta(1) integrin by reducing infiltrated eosinophils. We propose that blockade of integrin-mediated cell adhesion might be a target of the antiallergic action of levocabastine and may play a role in preventing eosinophil adhesion and infiltration in AC.

Therapeutic targeting of Polo-like kinase-1 and Aurora kinases in T-cell acute lymphoblastic leukemia

Polo-like kinases (PLKs) and Aurora kinases (AKs) act as key cell cycle regulators in healthy human cells. In cancer, these protein kinases are often overexpressed and dysregulated, thus contributing to uncontrolled cell proliferation and growth. T-cell acute lymphoblastic leukemia (T-ALL) is a heterogeneous malignancy arising in the thymus from T-cell progenitors. Primary chemoresistant and relapsed T-ALL patients have yet a poor outcome, therefore novel therapies, targeting signaling pathways important for leukemic cell proliferation, are required. Here, we demonstrate the potential therapeutic effects of BI6727, MK-5108, and GSK1070916, three selective inhibitors of PLK1, AK-A, and AK-B/C, respectively, in a panel of T-ALL cell lines and primary cells from T-ALL patients. The drugs were both cytostatic and cytotoxic to T-ALL cells by inducing G2/M-phase arrest and apoptosis. The drugs retained part of their pro-apoptotic activity in the presence of MS-5 bone marrow stromal cells. Moreover, we document for the first time that BI6727 perturbed both the PI3K/Akt/mTORC2 and the MEK/ERK/mTORC1 signaling pathways, and that a combination of BI6727 with specific inhibitors of the aforementioned pathways (MK-2206, CCI-779) displayed significantly synergistic cytotoxic effects. Taken together, our findings indicate that PLK1 and AK inhibitors display the potential for being employed in innovative therapeutic strategies for improving T-ALL patient outcome.

Synthesis and Conformational Analysis of Cyclotetrapeptide Mimetic β-Turn Templates and Validation as 3D Scaffolds

The conformations of all stereoisomers of PMRI cyclotetrapeptide mimetics 1–8 are essentially determined by the predisposition of the diamine to stabilize β‐turns. The peptide mimetics can be regarded as 3D scaffolds for designing molecules with a predictable display of the pharmacophores. We used the models for testing novel RGD analogues as αvβ3‐integrin receptor antagonists.

Targeting phosphatidylinositol 3-kinase signaling in acute myelogenous leukemia.

Introduction: Despite continuous advances in our knowledge of the biology of acute myelogenous leukemia (AML), the prognosis of AML patients treated with standard chemotherapy is still poor, especially in the elderly. Therefore, there is a need for novel targeted and less toxic therapies, particularly for patients who develop resistance to traditional chemotherapeutic drugs. Constitutively active phosphatidylinositol 3-kinase (PI3K) signaling characterizes many types of tumors, including AML, where it negatively influences response to therapeutic treatments. Areas covered: The literature data showed that small inhibitor molecules targeting PI3K signaling induced cell cycle arrest, apoptosis and decreased drug-resistance in AML cells. PI3K inhibitors were also capable of targeting leukemic initiating cells (LICs), the most relevant target for leukemia eradication, whereas they tended to spare healthy hematopoietic stem cells. Expert opinion: Data emerging from pre-clinical settings suggest that the PI3K pathway is critically involved in regulating proliferation, survival and drug-resistance of AML cells. Therefore, we propose that novel drugs targeting this signaling pathway may offer a novel and less toxic treatment option for AML patients, most likely in combination with a lower dosage of traditional chemotherapeutic agents or other innovative therapeutic agents.

New coumarin‐based anti‐inflammatory drug: putative antagonist of the integrins αLβ2 and αMβ2

This study was conducted to investigate putative antagonism of integrin receptors αMβ2 and αLβ2 by a novel coumarin derivative (BOL‐303225‐A), its efficacy in‐vivo after retinal ischaemia‐reperfusion injury, and its bioavailability in rat plasma. A cellular adhesion assay in Jurkat and U937 cells, and a flow cytometry assay with an antibody against the β2 subunit were conducted. BOL‐303225‐A bioavailability in rat plasma and the retinal levels of myeloperoxidase (MPO) after ischaemia‐ reperfusion injury were evaluated after oral administration (10 mg kg−1). In‐vitro cell viability assays revealed no cytotoxicity for BOL‐303225‐A over a wide dose range, and IC50 values of 32.3 ± 1.5 μM and 84.95 ± 2.3 μM were found for Jurkat and U937 cells, respectively. The drug showed specific binding to the αMβ2 and αLβ2 integrin receptors expressed by U937 and Jurkat cells, respectively, producing a fluorescence shift towards lower values in a concentration‐dependent manner. The pharmacokinetic profile of BOL‐303225‐A exhibited rapid absorption following oral administration in the rat. A significant reduction of retinal MPO levels was observed in drug‐treated rats. This study demonstrated that BOL‐303225‐A acts as an antagonist of the integrin αLβ2 and αMβ2 receptors, suggesting that this drug could be used for ocular diseases such as diabetic retinopathy.

β-Arrestin 2-mediated heterologous desensitization of IGF-IR by prolonged exposure of SH-SY5Y neuroblastoma cells to a mu opioid agonist

Prolonged (12 h) exposure of SH‐SY5Y neuroblastoma cells to the mu‐opioid receptor (MOPr) agonist [D‐Ala2,N‐Me‐Phe4,Gly5‐ol]‐enkephalin (DAMGO) causes homologous desensitization as well as heterologous desensitization of the extracellular signal‐regulated kinase 1/2 (ERK 1/2) phosphorylation induced by insulin‐like growth factor (IGF)‐I. Brief (15 min) but not prolonged exposure to DAMGO transregulates the insulin‐like growth factor‐I (IGF‐I) receptor, as evidenced by its phosphorylation in the absence of IGF‐I. Silencing of β‐arrestin 2 uncouples the crosstalk between the two receptors, thus maintaining IGF‐I‐mediated receptor phosphorylation and ERK 1/2 activation even after prolonged DAMGO exposure. Furthermore, MOPr‐induced activation of IGF‐I receptor requires the tyrosine kinase c‐Src.