Annamaria Riccio

Randomised controlled trial of local allergoid immunotherapy on allergic inflammation in mite-induced rhinoconjunctivitis

BACKGROUND Non-injective routes of immunotherapy are thought to be valuable therapeutic options for respiratory allergy. We investigated the clinical efficacy and the effects of sublingual/oral immunotherapy on conjunctival allergic inflammation in patients with mite-induced respiratory allergy. METHODS We used a double-blind placebo-controlled design. 20 patients with mite-induced rhinoconjunctivitis (six of whom also had mild asthma) were randomly assigned sublingual/oral immunotherapy (n=10) or placebo (n=10) for 2 years. We assessed symptom score by diary cards and inflammatory-cell infiltrate, and expression of intercellular adhesion molecule 1 (ICAM-1) in the conjunctiva after specific allergen challenge at enrollment and after 12 and 24 months of treatment. FINDINGS We found significantly lower symptom scores in the immunotherapy group than in the placebo group in most of the winter months (p=0.05). Compared with the placebo group, inflammatory-cell infiltration after conjunctival challenge, and ICAM-1 expression on conjunctival epithelium decreased significantly in the first year of treatment in the immunotherapy group (p=0.04 and p=0.02, respectively). These effects were also seen for the minimum persistent inflammation, in symptom-free patients exposed constantly to allergens (p=0.02). Serum concentrations of eosinophil cationic protein decreased significantly (p=0.04). Immunotherapy was well tolerated and compliance was good. INTERPRETATION Our results suggest that this immunotherapy is clinically effective in rhinoconjunctivitis and that it decreases the immune-mediated inflammatory responses to the allergen.

Clinical and immunologic effects of a rush sublingual immunotherapy to Parietaria species: A double-blind, placebo-controlled trial.

BACKGROUND The local (noninjection) routes of immunotherapy are presently regarded as viable therapeutic options for respiratory allergy, and their mechanisms of action are currently undergoing investigation. OBJECTIVE We evaluated the clinical efficacy of a preseasonal rush sublingual-swallow immunotherapy and its effects on allergic inflammation in patients with seasonal rhinoconjunctivitis caused by Parietaria species. METHODS Thirty patients with Parietaria species-induced rhinoconjunctivitis (13 with mild intermittent asthma) were randomly assigned sublingual-swallow immunotherapy or placebo in a rush preseasonal course. We assessed the seasonal symptom-drug intake score by diary card and the inflammatory infiltration and the intercellular adhesion molecule 1 expression on nasal epithelium after specific allergenic challenge before and after treatment. RESULTS The investigated immunotherapy was well tolerated, and no side effects were recorded. A significant reduction of the symptom score (P =.016) and drug intake score (P =. 008) after immunotherapy was observed only in the active group. A decrease of the cumulative score was observed also in the placebo group (P =.046), but the significance was clearly higher (P =.006) in the active group. In the active group a reduction of neutrophils (P =.001), eosinophils (P =.01), and intercellular adhesion molecule 1 expression (P =.04) after specific nasal challenge was also detected. CONCLUSION The present results suggest that this sublingual-swallow immunotherapy administered through a rush schedule is clinically effective and safe and that it decreases the immune-mediated inflammatory responses to the allergen.

Omalizumab Modulates Bronchial Reticular Basement Membrane Thickness and Eosinophil Infiltration in Severe Persistent Allergic Asthma Patients

Severe persistent asthma causes a substantial morbidity and mortality burden and is frequently not well controlled, despite intensive guideline-based therapy. The unique monoclonal antibody approved for patients with severe allergic asthma is omalizumab: a recombinant humanised murine against IgE antibodies. The aim of the present study is to investigate the effect of long-term anti-IgE on the thickening of the reticular basement membrane (RBM) and eosinophil infiltration in bronchial biopsies from patients with severe persistent allergic asthma. Biopsies were obtained from 11 patients with severe persistent allergic asthma before and after (12 months) treatment with omalizumab. RBM thickness and eosinophils were measured by using light microscope image analysis. A significant mean reduction in RBM thickness and eosinophil infiltration were measured after one-year omalizumab treatment. No correlation between eosinophil reduction and RBM thickness reduction was found. No correlation between each of the previous two parameters and clinical parameters was detected. In conclusion, our study showed that a substantial proportion of severe asthmatics reduced the original bronchial RBM thickness and eosinophil infiltration after one-year treatment with anti-IgE, thus emphasizing the possible role of omalizumab in affecting airway remodeling in severe persistent allergic asthma.

Bone mineral density, bone turnover markers and fractures in patients with indolent systemic mastocytosis

OBJECTIVE We systematically assessed bone mineral density (BMD), bone turnover markers (BTM), and fractures in a large cohort of patients with Indolent Systemic Mastocytosis (ISM). METHODS Eighty-two patients (mean age 48 years, 37 women) with ISM were studied. BMD was measured by dual X-ray absorptiometry at the lumbar spine and proximal hip. The serum markers of bone turnover included bone-specific alkaline phosphatase, C-telopeptides of type I collagen, and serum osteocalcin. Previous clinical fractures were registered and spine X-ray was obtained from all patients. RESULTS Three women were excluded for concomitant diseases associated to osteoporosis. Osteoporosis according with the WHO classification (T-score<-2.5) was found in 16 patients (20.0%) (7 females and 9 men). Mastocytosis-related low BMD (Z-score at either the spine or the hip<-2) was found in 3 women (9%) and 13 men (28%). The BMD was generally lower at the spine than at the hip. No significant correlation was observed between serum tryptase levels and T or Z-score BMD. One or more moderate or severe vertebral fractures were found in 17 patients (12 men); in 11 of them Z-score values were>-2 or not valuable at the spine. No significant difference was found in the prevalence of mastocytosis-related low BMD and/or vertebral fractures between patients with or without skin involvement. Two patients had radiographic and densitometric osteosclerosis-like characteristics. In osteoporotic patients higher, normal or lower serum BTM were found, without correlations with serum tryptase levels, while in patients with osteosclerosis both BTM and serum tryptase values were particularly increased. CONCLUSIONS Vertebral osteoporosis and fractures are frequent in patients with ISM. Spine X-ray and densitometric examination are warranted in all patients, also without skin involvement and particularly in males; Z-score other than T-score BMD must be evaluated. Patients with idiopathic osteoporosis should be evaluated for mast cell disease. Both high than low BTM can be observed in patients with osteoporosis while osteosclerosis is characterized by high bone turnover and serum tryptase levels.

Venom Immunotherapy in Patients with Clonal Mast CELL Disorders: Efficacy, Safety and Practical Considerations

BACKGROUND A preferential association between systemic mastocytosis (SM) and hymenoptera allergy (HVA) has been observed. Patients with both diseases are at risk for more severe reactions, and venom immunotherapy (VIT) may represent a life-saving treatment, but the use of VIT in such patients raised concerns about its safety. OBJECTIVE We evaluated a large population of patients with SM and HVA who received VIT. METHODS This prospective study was performed in Italy and Spain. A diagnosis of SM and HVA and a VIT prescription were made according to international recommendations. The patients were carefully followed up during VIT, with special attention to field stings. RESULTS A total of 84 patients (70 men, 14 women; mean age 52.1 years) were included, 81% with grade IV reaction, 91% with indolent SM. No difference was seen between the Italian and Spanish patients. There were 10 adverse reactions during the induction phase: 3 with the conventional induction and 7 with the rush-modified induction, none resulted in epinephrine administration and/or hospitalization. Fifty patients had one or more field re-sting (95 episodes), none during induction. The time elapsed from starting VIT and first re-sting was 2 months to 7 years, and the number of re-stings per patient was 1-6. Of the 50 patients who were re-stung, 43 (86%) resulted in being fully protected. Seven patients had reactions, and the maintenance dose was safely increased to 200 mcg. The maintenance dose interval was not different between patients with and those without reactions at re-stings. CONCLUSION VIT is well tolerated, safe, and effective in patients with SM.

IL-13 and idiopathic pulmonary fibrosis: Possible links and new therapeutic strategies

The recent advances in the knowledge of immunological aspects of many pulmonary diseases, allowed to identify cells, biological functions, cytokines, and receptors that are preferentially involved in each disease. This is the case of asthma, where IL-13 (together with IL-4) is recognized as a central mediator. The role of IL-13 is strictly related, via complex signaling pathways, to eosinophil recruitment and activation, to mucus secretion, periostin generation and to fibrogenic processes (which are part of the remodeling process). These peculiar roles of IL-13 have suggested the hypothesis of its role in Idiopathic Pulmonary Fibrosis, and consequently of its antagonists in the treatment of such disease. We review herein the immunological roles of IL-13 in asthma and IPF, and the currently ongoing attempts to treat IPF by IL-13 antagonism strategies.

COPD treatment: Real life and experimental effects on peripheral NK cells, their receptors expression and their IFN-γ secretion

A role in pulmonary immunity has been ascribed to Natural Killer (NK) cells and several in vitro studies have shown a corticosteroid-induced inhibition of NK cells mediated cytotoxicity. Several clinical trials on chronic obstructive pulmonary disease (COPD) have suggested a relationship between COPD treatment and occurrence of respiratory infections. Aims of our study were to investigate if real life COPD treatment affects peripheral blood NK cells total count and their receptors expression and to assess if different doses of formoterol and budesonide, administered alone or in combination, are able to modulate the surface expression of activating (NKp30, NKp44, NKp46 and NKG2D) and inhibitory (KIR2DL2/L3, KIR3DL1 and NKG2A) receptors on peripheral blood NK cells of COPD patients. Moreover, we evaluated the potential effect of treatment with budesonide and/or formoterol on IFN-γ secretion in vitro. NK cells were isolated from peripheral blood of 7 healthy volunteers, 9 chronic bronchitis (CB) and 11 COPD patients. Total NK cells count and activating and inhibitory receptors expression were evaluated. NK cells were cultured for 20h in 96-well plates with IL-2 (100IU/ml)+IL-12 (2.5ng/ml), with or without budesonide (Bud; 1 and 0.01μM) and formoterol (For; 30 and 0.3nM) alone or in combination. Cells were analyzed by flow cytometry and IFN-γ was measured in cell supernatants by ELISA test. No difference between real life treated COPD, CB and healthy subjects was found concerning NK total count and NK cell receptors expression. When cells were stimulated over night with cytokines and treated with drugs, only NKG2D receptor was modulated. Its expression was significantly downregulated by budesonide alone and in combination with formoterol in COPD patients. IFN-γ production induced by stimulation with IL-2+IL-12 was decreased in a highly significant way (p<0.01) by all treatments in all groups. Even if in vitro experiments with budesonide, alone or in combination with formoterol, showed a modulation of NKG2D receptor expression and IFN-γ production, our ex vivo results show that real life LABA and ICS treatment does not influence peripheral NK cells count and their receptors phenotype.

Salbutamol: how does it enter smooth muscle cells?

Polyspecific organic cation transporters (OCTs) in human cell membranes are involved in the uptake, distribution and excretion of cationic compounds. Although their relevance to drug disposition in the liver, small intestine and kidney has been investigated previously, less is known about the influence of these transporters on the pharmacokinetics and pharmacodynamics of inhaled drugs. Drugs that are commonly administered by inhalation for the treatment of respiratory diseases, such as glucocorticoids and cationic β2-agonists, might interact with several of these transporters, which are strongly expressed on the surfaces of airway epithelial cells. We evaluated the expression of OCT3 and measured the in vitro uptake of the short-acting β2-agonist salbutamol (SALB), alone or in combination with corticosterone (CS) and beclomethasone dipropionate (BDP), by bronchial smooth muscle cells. Our results showed that these cells express the OCT3 transporter and that SALB enters the cell in a transporter-independent fashion. Moreover, CS and BDP have different activities on SALB transport inside the cell. CS increases SALB transport and BDP decreases SALB transport, although neither of these effects are statistically significant. A better understanding of these mechanisms might lead to the improved treatment of airway diseases.

Poster 1002: Galectin-3 as predictive biomarker of airways remodeling modulation in omalizumab treated severe asthma patients

Background A significant effect of Omalizumab treatment on bronchial remodeling modulation(reduction of Reticular Basal Membrane-RBMthickening) by means of histological evaluation of bronchial biopsies was documented after therapy (Riccio et al. 2012). But we surprisingly found two groups , Responders-R (reduction of tickening of RBM) and Non Responders-NR (increasing or stable RBM). We then applied the proteomic analysis to the bronchial specimens. We used MudPIT (Multidimensional Protein Identification Technology) proteomic approach, a highthroughput methodology that allows the identification of hundreds/thousands of proteins for a single complex sample, evaluation of differential abundance, characterization of involved molecular pathways and sub-typing diseases (Brambilla et al., 2012).