TY - JOUR T1 - Causes of Treatment Failure in Children With Inflammatory Bowel Disease Treated With Infliximab: A Pharmacokinetic Study. JF - J Pediatr Gastroenterol Nutr Y1 - 2019 A1 - Naviglio, Samuele A1 - Lacorte, Doriana A1 - Lucafò, Marianna A1 - Cifù, Adriana A1 - Favretto, Diego A1 - Cuzzoni, Eva A1 - Silvestri, Tania A1 - Pozzi Mucelli, Martina A1 - Radillo, Oriano A1 - Decorti, Giuliana A1 - Fabris, Martina A1 - Bramuzzo, Matteo A1 - Taddio, Andrea A1 - Stocco, Gabriele A1 - Alvisi, Patrizia A1 - Ventura, Alessandro A1 - Martelossi, Stefano AB -

OBJECTIVES: Anti-tumor necrosis factor antibodies have led to a revolution in the treatment of inflammatory bowel diseases (IBD); however, a sizable proportion of patients does not respond to therapy. There is increasing evidence suggesting that treatment failure may be classified as mechanistic (pharmacodynamic), pharmacokinetic, or immune-mediated. Data regarding the contribution of these factors in children with IBD treated with infliximab (IFX) are still incomplete. The aim was to assess the causes of treatment failure in a prospective cohort of pediatric patients treated with IFX.

METHODS: This observational study considered 49 pediatric (median age 14.4) IBD patients (34 Crohn disease, 15 ulcerative colitis) treated with IFX. Serum samples were collected at 6, 14, 22 and 54 weeks, before IFX infusions. IFX and anti-infliximab antibodies (AIA) were measured using enzyme linked immunosorbent assays. Disease activity was determined by Pediatric Crohn's Disease Activity Index or Pediatric Ulcerative Colitis Activity Index.

RESULTS: Clinical remission, defined as a clinical score <10, was obtained by 76.3% of patients at week 14 and by 73.9% at week 54. Median trough IFX concentration was higher at all time points in patients achieving sustained clinical remission. IFX levels during maintenance correlated also with C-reactive protein, albumin, and fecal calprotectin. After multivariate analysis, IFX concentration at week 14 >3.11 μg/mL emerged as the strongest predictor of sustained clinical remission. AIA concentrations were correlated inversely with IFX concentrations and directly with adverse reactions.

CONCLUSIONS: Most cases of therapeutic failure were associated with low serum drug levels. IFX trough levels at the end of induction are associated with sustained long-term response.

VL - 68 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/30211845?dopt=Abstract ER - TY - JOUR T1 - High-Throughput Sequencing of microRNAs in Glucocorticoid Sensitive Paediatric Inflammatory Bowel Disease Patients. JF - Int J Mol Sci Y1 - 2018 A1 - De Iudicibus, Sara A1 - Lucafò, Marianna A1 - Vitulo, Nicola A1 - Martelossi, Stefano A1 - Zimbello, Rosanna A1 - De Pascale, Fabio A1 - Forcato, Claudio A1 - Naviglio, Samuele A1 - Di Silvestre, Alessia A1 - Gerdol, Marco A1 - Stocco, Gabriele A1 - Valle, Giorgio A1 - Ventura, Alessandro A1 - Bramuzzo, Matteo A1 - Decorti, Giuliana KW - Adolescent KW - Biomarkers KW - Child KW - Female KW - Gene Expression Regulation KW - Glucocorticoids KW - High-Throughput Nucleotide Sequencing KW - Humans KW - Inflammatory Bowel Diseases KW - Male KW - MicroRNAs KW - Receptors, Glucocorticoid KW - Transcriptome AB -

The aim of this research was the identification of novel pharmacogenomic biomarkers for better understanding the complex gene regulation mechanisms underpinning glucocorticoid (GC) action in paediatric inflammatory bowel disease (IBD). This goal was achieved by evaluating high-throughput microRNA (miRNA) profiles during GC treatment, integrated with the assessment of expression changes in GC receptor (GR) heterocomplex genes. Furthermore, we tested the hypothesis that differentially expressed miRNAs could be directly regulated by GCs through investigating the presence of GC responsive elements (GREs) in their gene promoters. Ten IBD paediatric patients responding to GCs were enrolled. Peripheral blood was obtained at diagnosis (T0) and after four weeks of steroid treatment (T4). MicroRNA profiles were analyzed using next generation sequencing, and selected significantly differentially expressed miRNAs were validated by quantitative reverse transcription-polymerase chain reaction. In detail, 18 miRNAs were differentially expressed from T0 to T4, 16 of which were upregulated and 2 of which were downregulated. Out of these, three miRNAs (miR-144, miR-142, and miR-96) could putatively recognize the 3’UTR of the GR gene and three miRNAs (miR-363, miR-96, miR-142) contained GREs sequences, thereby potentially enabling direct regulation by the GR. In conclusion, we identified miRNAs differently expressed during GC treatment and miRNAs which could be directly regulated by GCs in blood cells of young IBD patients. These results could represent a first step towards their translation as pharmacogenomic biomarkers.

VL - 19 IS - 5 U1 - http://www.ncbi.nlm.nih.gov/pubmed/29738455?dopt=Abstract ER - TY - JOUR T1 - Pharmacogenetics of treatments for inflammatory bowel disease. JF - Expert Opin Drug Metab Toxicol Y1 - 2018 A1 - Lucafò, Marianna A1 - Franca, Raffaella A1 - Selvestrel, Davide A1 - Curci, Debora A1 - Pugnetti, Letizia A1 - Decorti, Giuliana A1 - Stocco, Gabriele KW - Dose-Response Relationship, Drug KW - Epigenesis, Genetic KW - Gastrointestinal Agents KW - Genetic Markers KW - Genetic Predisposition to Disease KW - Humans KW - Inflammatory Bowel Diseases KW - Pharmacogenetics AB -

INTRODUCTION: Inflammatory bowel disease is a chronic inflammation of the gut whose pathogenesis is still unclear. Although no curative therapy is currently available, a number of drugs are used in induction and maintenance therapy; however, for most of these drugs, a high inter-individual variability in response is observed. Among the factors of this variability, genetics plays an important role. Areas covered: This review summarizes the results of pharmacogenetic studies, considering the most important drugs used and in particular aminosalycilates, glucocorticoids, thiopurines, monoclonal antibodies and thalidomide. Most studies used a candidate gene approach, even if significant breakthroughs have been obtained recently from applying genome-wide studies. When available, also investigations considering epigenetics and pharmacogenetic dosing guidelines have been included. Expert opinion: Only for thiopurines, genetic markers identified as predictors of efficacy or adverse events have allowed the development of dosing guidelines. For the other drugs, encouraging results are available and great expectations rely on the study of epigenetics and integration with pharmacokinetic information, especially useful for biologics. However, to improve therapy of IBD patients with these drugs, for implementation in the clinics of pharmacogenetics, informatic clinical decision support systems and training about pharmacogenetics of health providers are needed.

VL - 14 IS - 12 U1 - http://www.ncbi.nlm.nih.gov/pubmed/30465611?dopt=Abstract ER - TY - JOUR T1 - Role of the Long Non-Coding RNA Growth Arrest-Specific 5 in Glucocorticoid Response in Children with Inflammatory Bowel Disease. JF - Basic Clin Pharmacol Toxicol Y1 - 2018 A1 - Lucafò, Marianna A1 - Di Silvestre, Alessia A1 - Romano, Maurizio A1 - Avian, Alice A1 - Antonelli, Roberta A1 - Martelossi, Stefano A1 - Naviglio, Samuele A1 - Tommasini, Alberto A1 - Stocco, Gabriele A1 - Ventura, Alessandro A1 - Decorti, Giuliana A1 - De Iudicibus, Sara KW - Biomarkers KW - Cell Line, Tumor KW - Cell Proliferation KW - Child KW - Drug Resistance KW - Female KW - Gene Knockdown Techniques KW - Glucocorticoids KW - Humans KW - Inflammatory Bowel Diseases KW - Male KW - Patient Selection KW - Pharmacogenomic Testing KW - Precision Medicine KW - RNA, Long Noncoding KW - RNA, Small Interfering KW - Treatment Outcome KW - Up-Regulation AB -

Glucocorticoids (GCs) are widely employed in inflammatory, autoimmune and neoplastic diseases, and, despite the introduction of novel therapies, remain the first-line treatment for inducing remission in inflammatory bowel disease (IBD). Given the high incidence of suboptimal response, associated with a significant number of side-effects, that are particularly severe in paediatric patients, the identification of subjects that are most likely to respond poorly to GCs is extremely important. Recent evidence suggests that the long non-coding RNA (lncRNA) GAS5 could be a potential marker of GC resistance. To address this issue, we evaluated the association between the lncRNA GAS5 and the efficacy of steroids, in terms of inhibition of proliferation, in two cell lines derived from colon and ovarian cancers, to confirm the sensitivity and specificity of these lncRNAs. These cells showed a different sensitivity to GCs and revealed differential expression of GAS5 after treatment. GAS5 was up-regulated in GC-resistant cells and accumulated more in the cytoplasm compared to the nucleus in response to the drug. The functions of GAS5 were assessed by silencing, and we found that GAS5 knock-down reduced the proliferation during GC treatment. Furthermore, for the first time, we measured GAS5 levels in 19 paediatric IBD patients at diagnosis and after the first cycle of GCs, and we demonstrated an up-regulation of the lncRNA in patients with unfavourable steroid response. Our preliminary results indicate that GAS5 could be considered a novel pharmacogenomic marker useful for the personalization of GC therapy.

VL - 122 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/28722800?dopt=Abstract ER - TY - JOUR T1 - Epratuzumab and Blinatumomab as Therapeutic Antibodies for Treatment of Pediatric Acute Lymphoblastic Leukemia: Current Status and Future Perspectives. JF - Curr Med Chem Y1 - 2017 A1 - Franca, Raffaella A1 - Favretto, Diego A1 - Granzotto, Marilena A1 - Decorti, Giuliana A1 - Rabusin, Marco A1 - Stocco, Gabriele KW - Antibodies, Bispecific KW - Antibodies, Monoclonal KW - Antibodies, Monoclonal, Humanized KW - Antineoplastic Agents KW - Child KW - Clinical Trials as Topic KW - Half-Life KW - Humans KW - Precursor Cell Lymphoblastic Leukemia-Lymphoma KW - Sialic Acid Binding Ig-like Lectin 2 AB -

BACKGROUND: More than 85% of children affected by acute lymphoblastic leukemia (ALL) are successfully treated; however relapse remains a remarkable clinical concern, with 50-60% of relapsing patients facing a fatal outcome. Management of relapsed patients includes standardized intensive risk-adapted regimens based on conventional drugs, and hematopoietic stem cells transplantation for patients with unfavourable features. Biological drugs, in particular the monoclonal antibody epratuzumab and the bi-functional recombinant single chain peptide blinatumomab, have been recently recognized as novel potential agents to be integrated in salvage ALL therapy to further improve rescue outcome.

METHODS: A systematic search of peer-reviewed scientific literature and clinical trials in public databases has been carried out. Both clinical and pre-clinical studies have been included to summarize recent evidence on epratuzumab and blinatumomab for salvage ALL therapy.

RESULTS: Sixty-two papers and 25 clinical trials were included. Although not all patients responded properly to these agents, their use in relapsed and refractory pediatric ALL seems promising.

CONCLUSION: Phase 3 studies have recently begun and more consistent results about epratuzumab and blinatumomab safety and efficacy in comparison to conventional therapies are expected in the next years. Epratuzumab seems safe in the dosing scheme proposed in ALL, but its efficacy over the conventional chemotherapy is still questionable. Blinatumomab has shown promising results in high risk cases such as elder adult patients and conclusive studies on pediatric ALL are needed. Patient inter-individual variability to these agents has not been investigated in depth, but this issue needs to be addressed, in particular for blinatumomab.

VL - 24 IS - 11 U1 - http://www.ncbi.nlm.nih.gov/pubmed/28088906?dopt=Abstract ER - TY - JOUR T1 - Multicentric Case-Control Study on Azathioprine Dose and Pharmacokinetics in Early-onset Pediatric Inflammatory Bowel Disease. JF - Inflamm Bowel Dis Y1 - 2017 A1 - Stocco, Gabriele A1 - Martelossi, Stefano A1 - Arrigo, Serena A1 - Barabino, Arrigo A1 - Aloi, Marina A1 - Martinelli, Massimo A1 - Miele, Erasmo A1 - Knafelz, Daniela A1 - Romano, Claudio A1 - Naviglio, Samuele A1 - Favretto, Diego A1 - Cuzzoni, Eva A1 - Franca, Raffaella A1 - Decorti, Giuliana A1 - Ventura, Alessandro KW - Adolescent KW - Age of Onset KW - Antimetabolites KW - Azathioprine KW - Case-Control Studies KW - Child KW - Child, Preschool KW - Chromatography, High Pressure Liquid KW - Dose-Response Relationship, Drug KW - Erythrocytes KW - Female KW - Guanine Nucleotides KW - Humans KW - Inflammatory Bowel Diseases KW - Male KW - Mercaptopurine KW - Methyltransferases KW - Thioguanine AB -

BACKGROUND: Early-onset inflammatory bowel disease (IBD) is generally aggressive, with a high probability of complications and need of surgery. Despite the introduction of highly effective biological drugs, treatment with azathioprine continues to be important even for early-onset IBD; however, in these patients azathioprine response seems to be reduced. This study evaluated azathioprine doses, metabolite concentrations, and their associations with patients' age in children with IBD treated at 6 tertiary pediatric referral centers.

METHODS: Azathioprine doses, metabolites, and clinical effects were assessed after at least 3 months of therapy in 17 early-onset (age < 6 yr, cases) and 51 nonearly-onset (aged > 12 and <18 yrs, controls) patients with IBD. Azathioprine dose was titrated on therapeutic efficacy (response and adverse effects). Azathioprine metabolites and thiopurine methyltransferase activity were determined by high-performance liquid chromatography with ultra violet-vis detection (HPLC-UV) methods.

RESULTS: Frequency of patients in remission was similar among early-onset and control groups, respectively (82% and 84%, P value = 0.72). Early-onset patients required higher doses of azathioprine (median 2.7 versus 2.0 mg·kg·d, P value = 1.1 × 10). Different doses resulted in comparable azathioprine active thioguanine nucleotide metabolite concentrations (median 263 versus 366 pmol/8 × 10 erythrocytes, P value = 0.41) and methylmercaptopurine nucleotide concentrations (median 1455 versus 1532 pmol/8 × 10 erythrocytes, P value = 0.60). Lower ratios between thioguanine nucleotide metabolites and azathioprine doses were found in early-onset patients (median 98 versus 184 pmol/8 × 10 erythrocytes·mg·kg·d, P value = 0.017). Interestingly, early-onset patients presented also higher thiopurine methyltransferase activity (median 476 versus 350 nmol methylmercaptopurine/mg hemoglobin/h, P-value = 0.046).

CONCLUSIONS: This study demonstrated that patients with early-onset IBD present increased inactivating azathioprine metabolism, likely because of elevated activity of the enzyme thiopurine methyltransferase.

VL - 23 IS - 4 U1 - http://www.ncbi.nlm.nih.gov/pubmed/28296824?dopt=Abstract ER - TY - JOUR T1 - Action of methotrexate and tofacitinib on directly stimulated and bystander-activated lymphocytes. JF - Mol Med Rep Y1 - 2016 A1 - Piscianz, Elisa A1 - Candilera, Vanessa A1 - Valencic, Erica A1 - Loganes, Claudia A1 - Paron, Greta A1 - De Iudicibus, Sara A1 - Decorti, Giuliana A1 - Tommasini, Alberto AB -

Chronic inflammation associated with autoimmune activation is characteristic of rheumatic diseases from childhood to adulthood. In recent decades, significant improvements in the treatment of these types of disease have been achieved using disease modifying anti-rheumatic drugs (DMARDs), such as methotrexate (MTX) and, more recently, using biologic inhibitors. The recent introduction of kinase inhibitors (for example, tofacitinib; Tofa) further increases the available ARDs. However, there are patients that do not respond to any treatment strategies, for whom combination therapies are proposed. The data regarding the combined action of different drugs is lacking and the knowledge of the mechanisms of ARDs and their actions upon pathogenic lymphocytes, which are hypothesized to sustain disease, is poor. An in vitro model of inflammation was developed in the current study, in which stimulated and unstimulated lymphocytes were cultured together, but tracked separately, to investigate the action of MTX and Tofa on the two populations. By analysing lymphocyte proliferation and activation, and cytokine secretion in the culture supernatants, it was established that, due to the presence of activated cells, unstimulated cells underwent a bystander activation that was modulated by the ARDs. Additionally, varying administration schedules were demonstrated to affect lymphocytes differently in vitro, either directly or via bystander activation. Furthermore, MTX and Tofa exerted different effects; while MTX showed an antiproliferative effect, Tofa marginally effected activation, although only a slight antiproliferative action, which could be potentiated by sequential treatment with MTX. Thus, it was hypothesized that these differences may be exploited in sequential therapeutic strategies, to maximize the anti‑rheumatic effect. These findings are notable and must be accounted for, as bystander‑activated cells in vivo could contribute to the spread of autoimmune activation and disease progression.

VL - 14 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27175898?dopt=Abstract ER - TY - JOUR T1 - Carbamazepine-induced thrombocytopenic purpura in a child: Insights from a genomic analysis. JF - Blood Cells Mol Dis Y1 - 2016 A1 - Abate, Maria Valentina A1 - Stocco, Gabriele A1 - Devescovi, Raffaella A1 - Carrozzi, Marco A1 - Pierobon, Chiara A1 - Valencic, Erica A1 - Lucafò, Marianna A1 - Di Silvestre, Alessia A1 - d'Adamo, Pio A1 - Tommasini, Alberto A1 - Decorti, Giuliana A1 - Ventura, Alessandro VL - 59 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27282575?dopt=Abstract ER - TY - JOUR T1 - Differential expression of GAS5 in rapamycin-induced reversion of glucocorticoid resistance. JF - Clin Exp Pharmacol Physiol Y1 - 2016 A1 - Lucafò, Marianna A1 - Bravin, Vanessa A1 - Tommasini, Alberto A1 - Martelossi, Stefano A1 - Rabach, Ingrid A1 - Ventura, Alessandro A1 - Decorti, Giuliana A1 - De Iudicibus, Sara AB -

This study evaluates the association between the long noncoding RNA GAS5 levels and the anti-proliferative effect of the glucocorticoid (GC) methylprednisolone (MP) alone and in combination with rapamycin in peripheral blood mononuclear cells (PBMCs) obtained from healthy donors. The effect of MP, rapamycin, and MP plus rapamycin was determined in 17 healthy donors by labelling metabolically active cells with [methyl-3H] thymidine and the expression levels of GAS5 gene were evaluated by real-time RT-PCR TaqMan analysis. We confirmed a role for GAS5 in modulating GC response: poor responders presented higher levels of GAS5 in comparison with good responders. Interestingly, when PBMCs were treated with the combination of rapamycin plus MP, the high levels of GAS5 observed for each drug in the MP poor responders group decreased in comparison with rapamycin (P value = 0.0134) or MP alone (P value = 0.0193). GAS5 is involved in GC resistance and co-treatment of rapamycin with GCs restores GC effectiveness in poor responders through the downregulation of the long noncoding RNA. GAS5 could be considered a biomarker to personalize therapy and a novel therapeutic target useful for the development of new pharmacological approaches to restore GC sensitivity.

VL - 43 IS - 6 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27001230?dopt=Abstract ER - TY - JOUR T1 - Thiopurine Biotransformation and Pharmacological Effects: Contribution of Oxidative Stress. JF - Curr Drug Metab Y1 - 2016 A1 - Pelin, Marco A1 - De Iudicibus, Sara A1 - Londero, Margherita A1 - Spizzo, Riccardo A1 - Dei Rossi, Sveva A1 - Martelossi, Stefano A1 - Ventura, Alessandro A1 - Decorti, Giuliana A1 - Stocco, Gabriele AB -

BACKGROUND: Thiopurine antimetabolites are important agents for the treatment of severe diseases, such as acute lymphoblastic leukemia and inflammatory bowel disease. Their pharmacological actions depend on biotransformation into active thioguanine-nucleotides; intracellular metabolism is mediated by enzymes of the salvage pathway of nucleotide synthesis and relies on polymorphic enzymes involved in thiopurines' catabolism such as thiopurine-S-methyl transferase. Given the enzymes involved in thiopurines' metabolism, it is reasonable to hypothesize that these drugs are able to induce significant oxidative stress conditions, possibly altering their pharmacological activity.

METHODS: A systemic search of peer-reviewed scientific literature in bibliographic databases has been carried out. Both clinical and preclinical studies as well as mechanistic studies have been included to shed light on the role of oxidative stress in thiopurines' pharmacological effects.

RESULTS: Sixty-nine papers were included in our review, allowing us to review the contribution of oxidative stress in the pharmacological action of thiopurines. Thiopurines are catabolized in the liver by xanthine oxidase, with potential production of reactive oxidative species and azathioprine is converted into mercaptopurine by a reaction with reduced glutathione, that, in some tissues, may be facilitated by glutathione- S-transferase (GST). A clear role of GSTM1 in modulating azathioprine cytotoxicity, with a close dependency on superoxide anion production, has been recently demonstrated. Interestingly, recent genome-wide association studies have shown that, for both azathioprine in inflammatory bowel disease and mercaptopurine in acute lymphoblastic leukemia, treatment effects on patients' white blood cells are related to variants of a gene, NUDT15, involved in biotransformation of oxidated nucleotides.

CONCLUSIONS: Basing on previous evidences published in literature, oxidative stress may contribute to thiopurine effects in significant ways that, however, are still not completely elucidated.

VL - 17 IS - 6 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26935390?dopt=Abstract ER - TY - JOUR T1 - 5-Aminoimidazole-4-carboxamide ribonucleotide-transformylase and inosine-triphosphate-pyrophosphatase genes variants predict remission rate during methotrexate therapy in patients with juvenile idiopathic arthritis. JF - Rheumatol Int Y1 - 2015 A1 - Pastore, Serena A1 - Stocco, Gabriele A1 - Moressa, Valentina A1 - Zandonà, Luigi A1 - Favretto, Diego A1 - Malusà, Noelia A1 - Decorti, Giuliana A1 - Lepore, Loredana A1 - Ventura, Alessandro AB -

For children with juvenile idiopathic arthritis (JIA) who fail to respond to methotrexate, the delay in identifying the optimal treatment at an early stage of disease can lead to long-term joint damage. Recent studies indicate that relevant variants to predict methotrexate response in JIA are those in 5-aminoimidazole-4-carboxamide ribonucleotide-transformylase (ATIC), inosine-triphosphate-pyrophosphatase (ITPA) and solute-liquid-carrier-19A1 genes. The purpose of the study was, therefore, to explore the role of these candidate genetic factors on methotrexate response in an Italian cohort of children with JIA. Clinical response to methotrexate was evaluated as clinical remission stable for a 6-month period, as ACRPed score and as change in Juvenile Arthritis Disease score. The most relevant SNPs for each gene considered were assayed on patients' DNA. ITPA activity was measured in patients' erythrocytes. Sixty-nine patients with JIA were analyzed: 52.2 % responded to therapy (ACRPed70 score), while 37.7 % reached clinical remission stable for 6 months. ATIC rs2372536 GG genotype was associated with improved clinical remission (adjusted p value = 0.0090). For ITPA, rs1127354 A variant was associated with reduced clinical remission: (adjusted p value = 0.028); this association was present even for patients with wild-type ITPA and low ITPA activity. These preliminary results indicate that genotyping of ATIC rs2372536 and ITPA rs1127354 variants or measuring ITPA activity could be useful to predict methotrexate response in children with JIA after validation by further prospective studies on a larger patient cohort.

VL - 35 IS - 4 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25240429?dopt=Abstract ER - TY - JOUR T1 - Effect of Thalidomide on Clinical Remission in Children and Adolescents with Ulcerative Colitis Refractory to Other Immunosuppressives: Pilot Randomized Clinical Trial. JF - Inflamm Bowel Dis Y1 - 2015 A1 - Lazzerini, Marzia A1 - Martelossi, Stefano A1 - Magazzù, Giuseppe A1 - Pellegrino, Salvatore A1 - Lucanto, Maria Cristina A1 - Barabino, Arrigo A1 - Calvi, Angela A1 - Arrigo, Serena A1 - Lionetti, Paolo A1 - Lorusso, Monica A1 - Mangiantini, Francesca A1 - Fontana, Massimo A1 - Zuin, Giovanna A1 - Palla, Gabriella A1 - Maggiore, Giuseppe A1 - Bramuzzo, Matteo A1 - Pellegrin, Maria Chiara A1 - Maschio, Massimo A1 - Villanacci, Vincenzo A1 - Manenti, Stefania A1 - Decorti, Giuliana A1 - De Iudicibus, Sara A1 - Paparazzo, Rossella A1 - Montico, Marcella A1 - Ventura, Alessandro AB -

BACKGROUND: In a randomized controlled trial, thalidomide has shown to be effective in refractory Crohn's disease in children. This pilot study aimed at evaluating thalidomide in refractory pediatric ulcerative colitis (UC).

METHODS: Double-blind, placebo-controlled randomized clinical trial on thalidomide 1.5 to 2.5 mg/kg/day in children with active UC despite multiple immunosuppressive treatments. In an open-label extension, nonresponders to placebo received thalidomide for an additional 8 weeks; all responders were followed up for a minimum of 52 weeks.

RESULTS: Twenty-six children with refractory UC were randomized to thalidomide or placebo. Clinical remission at week 8 was achieved by significantly more children treated with thalidomide {10/12 (83.3%) versus 2/11 (18.8%); risk ratio, 4.5 (95% confidence interval [CI], 1.2-16.4); P = 0.005; number needed to treat, 1.5}. Of the nonresponders to placebo who were switched to thalidomide, 8 of 11 (72.7%) subsequently reached remission at week 8 (risk ratio, 4.0 [95% CI, 1.1-14.7]; number needed to treat, 2.45; P = 0.01). Clinical remission in the thalidomide group was 135.0 weeks (95% CI, 32-238), compared with 8.0 weeks (95% CI, 2.4-13.6) in the placebo group (P < 0.0001). Cumulative incidence of severe adverse events was 3.1 per 1000 patient-weeks. Peripheral neuropathy and amenorrhea were the most frequent adverse events.

CONCLUSIONS: In this pilot randomized controlled trial on cases of UC refractory to immunosuppressive therapy, thalidomide compared with placebo resulted in improved clinical remission at 8 weeks of treatment and in longer term maintenance of remission. These findings require replication in larger clinical studies evaluating both thalidomide efficacy and safety.

VL - 21 IS - 8 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26185909?dopt=Abstract ER - TY - JOUR T1 - Failure of interferon-γ pre-treated mesenchymal stem cell treatment in a patient with Crohn's disease. JF - World J Gastroenterol Y1 - 2015 A1 - Taddio, Andrea A1 - Tommasini, Alberto A1 - Valencic, Erica A1 - Biagi, Ettore A1 - Decorti, Giuliana A1 - De Iudicibus, Sara A1 - Cuzzoni, Eva A1 - Gaipa, Giuseppe A1 - Badolato, Raffaela A1 - Prandini, Alberto A1 - Biondi, Andrea A1 - Ventura, Alessandro AB -

Mesenchymal stem cells (MSC) are cells of stromal origin which exhibit unlimited self-renewal capacity and pluripotency in vitro. It has recently been observed that MSC may also exert a profound immunosuppressive and anti-inflammatory effect both in vitro and in vivo with consequent potential use in autoimmune disorders. We present the case of a patient suffering from childhood-onset, multidrug resistant and steroid-dependent Crohn's disease who underwent systemic infusions of MSC, which led to a temporary reduction in CCR4, CCR7 and CXCR4 expression by T-cells, and a temporary decrease in switched memory B-cells, In addition, following MSC infusion, lower doses of steroids were needed to inhibit proliferation of the patient's peripheral blood mononuclear cells. Despite these changes, no significant clinical benefit was observed, and the patient required rescue therapy with infliximab and subsequent autologous hematopoietic stem cell transplantation. The results of biological and in vitro observations after MSC use and the clinical effects of infusion are discussed, and a brief description is provided of previous data on MSC-based therapy in autoimmune disorders.

VL - 21 IS - 14 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25892890?dopt=Abstract ER - TY - JOUR T1 - Genetic determinants for methotrexate response in juvenile idiopathic arthritis. JF - Front Pharmacol Y1 - 2015 A1 - Pastore, Serena A1 - Stocco, Gabriele A1 - Favretto, Diego A1 - De Iudicibus, Sara A1 - Taddio, Andrea A1 - d'Adamo, Pio A1 - Malusà, Noelia A1 - Addobbati, Riccardo A1 - Decorti, Giuliana A1 - Lepore, Loredana A1 - Ventura, Alessandro AB -

Juvenile idiopathic arthritis (JIAs) is the most common chronic rheumatic disease of childhood and is an important cause of disability. The folic acid analog methotrexate is the first choice disease-modifying anti-rheumatic drug in this disease, however, 35-45% of patients fail to respond. Molecular elements, such as variants in genes of pharmacological relevance, influencing response to methotrexate in JIA, would be important to individualize treatment strategies. Several studies have evaluated the effects of candidate genetic variants in the complex pathway of genes involved in methotrexate pharmacodynamics and pharmacokinetics, however, results are still contrasting and no definitive genetic marker of methotrexate response useful for the clinician to tailor therapy of children with JIA has been identified. Recently, genome-wide approaches have been applied, identifying new potential biological processes involved in methotrexate response in JIA such as TGF-beta signaling and calcium channels. If these genomic results are properly validated and integrated with innovative analyses comprising deep sequencing, epigenetics, and pharmacokinetics, they will greatly contribute to personalize therapy with methotrexate in children with JIA.

VL - 6 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25852556?dopt=Abstract ER - TY - JOUR T1 - Glucocorticoid pharmacogenetics in pediatric idiopathic nephrotic syndrome. JF - Pharmacogenomics Y1 - 2015 A1 - Cuzzoni, Eva A1 - De Iudicibus, Sara A1 - Franca, Raffaella A1 - Stocco, Gabriele A1 - Lucafò, Marianna A1 - Pelin, Marco A1 - Favretto, Diego A1 - Pasini, Andrea A1 - Montini, Giovanni A1 - Decorti, Giuliana AB -

Idiopathic nephrotic syndrome represents the most common type of primary glomerular disease in children: glucocorticoids (GCs) are the first-line therapy, even if considerable interindividual differences in thepir efficacy and side effects have been reported. Immunosuppressive and anti-inflammatory effects of these drugs are mainly due to the GC-mediated transcription regulation of pro- and anti-inflammatory genes. This mechanism of action is the result of a complex multistep pathway that involves the glucocorticoid receptor and several other proteins, encoded by polymorphic genes. Aim of this review is to highlight the current knowledge on genetic variants that could affect GC response, particularly focusing on children with idiopathic nephrotic syndrome.

VL - 16 IS - 14 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26419298?dopt=Abstract ER - TY - JOUR T1 - Patients' Induced Pluripotent Stem Cells to Model Drug Induced Adverse Events: A Role in Predicting Thiopurine Induced Pancreatitis? JF - Curr Drug Metab Y1 - 2015 A1 - Stocco, Gabriele A1 - Lanzi, Gaetana A1 - Yue, Fengming A1 - Giliani, Silvia A1 - Sasaki, Katsunori A1 - Tommasini, Alberto A1 - Pelin, Marco A1 - Martelossi, Stefano A1 - Ventura, Alessandro A1 - Decorti, Giuliana AB -

Induced pluripotent stem cells (iPSC) can be produced from adult cells by transfecting them with a definite set of pluripotency-associated genes. Under adequate growth conditions and stimulation iPSC can differentiate to almost every somatic lineage in the body. Patients' derived iPSC are an innovative model to study mechanisms of adverse drug reactions in individual patients and in cell types that cannot be easily obtained from human subjects. Proof-of concept studies with known toxicants have been performed for liver, cardiovascular and central nervous system cells: neurons obtained from iPSC have been used to elucidate the mechanism of chemotherapy-induced peripheral neuropathy by evaluating the effects of neurotoxic drugs such as vincristine. However, no study has been performed yet on pancreatic tissue and drug induced pancreatitis. Thiopurines (azathioprine and mercaptopurine) are immunosuppressive antimetabolite drugs, commonly used to treat Crohn's disease. About 5% of Crohn's disease patients treated with thiopurines develop pancreatitis, a severe idiosyncratic adverse event; these patients have to stop thiopurine administration and may require medical treatment, with significant personal and social costs. Molecular mechanism of thiopurine induced pancreatitis (TIP) is currently unknown and no fully validated biomarker is available to assist clinicians in preventing this adverse event. Hence, in this review we have reflected upon the probable research applications of exocrine pancreatic cells generated from patient specific iPS cells. Such pancreatic cells can provide excellent insights into the molecular mechanism of TIP. In particular three hypotheses on the mechanism of TIP could be explored: drug biotransformation, innate immunity and adaptative immunity.

VL - 17 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26526832?dopt=Abstract ER - TY - JOUR T1 - Role of oxidative stress mediated by glutathione-s-transferase in thiopurines' toxic effects. JF - Chem Res Toxicol Y1 - 2015 A1 - Pelin, Marco A1 - De Iudicibus, Sara A1 - Fusco, Laura A1 - Taboga, Eleonora A1 - Pellizzari, Giulia A1 - Lagatolla, Cristina A1 - Martelossi, Stefano A1 - Ventura, Alessandro A1 - Decorti, Giuliana A1 - Stocco, Gabriele AB -

Azathioprine (AZA), 6-mercaptopurine (6-MP), and 6-thioguanine (6-TG) are antimetabolite drugs, widely used as immunosuppressants and anticancer agents. Despite their proven efficacy, a high incidence of toxic effects in patients during standard-dose therapy is recorded. The aim of this study is to explain, from a mechanistic point of view, the clinical evidence showing a significant role of glutathione-S-transferase (GST)-M1 genotype on AZA toxicity in inflammatory bowel disease patients. To this aim, the human nontumor IHH and HCEC cell lines were chosen as predictive models of the hepatic and intestinal tissues, respectively. AZA, but not 6-MP and 6-TG, induced a concentration-dependent superoxide anion production that seemed dependent on GSH depletion. N-Acetylcysteine reduced the AZA antiproliferative effect in both cell lines, and GST-M1 overexpression increased both superoxide anion production and cytotoxicity, especially in transfected HCEC cells. In this study, an in vitro model to study thiopurines' metabolism has been set up and helped us to demonstrate, for the first time, a clear role of GST-M1 in modulating AZA cytotoxicity, with a close dependency on superoxide anion production. These results provide the molecular basis to shed light on the clinical evidence suggesting a role of GST-M1 genotype in influencing the toxic effects of AZA treatment.

VL - 28 IS - 6 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25928802?dopt=Abstract ER - TY - JOUR T1 - Role of Pharmacogenetics in Hematopoietic Stem Cell Transplantation Outcome in Children. JF - Int J Mol Sci Y1 - 2015 A1 - Franca, Raffaella A1 - Stocco, Gabriele A1 - Favretto, Diego A1 - Giurici, Nagua A1 - Decorti, Giuliana A1 - Rabusin, Marco AB -

Hematopoietic stem cell transplantation (HSCT) is an established therapeutic procedure for several congenital and acquired disorders, both malignant and nonmalignant. Despite the great improvements in HSCT clinical practices over the last few decades, complications, such as graft vs. host disease (GVHD) and sinusoidal obstructive syndrome (SOS), are still largely unpredictable and remain the major causes of morbidity and mortality. Both donor and patient genetic background might influence the success of bone marrow transplantation and could at least partially explain the inter-individual variability in HSCT outcome. This review summarizes some of the recent studies on candidate gene polymorphisms in HSCT, with particular reference to pediatric cohorts. The interest is especially focused on pharmacogenetic variants affecting myeloablative and immunosuppressive drugs, although genetic traits involved in SOS susceptibility and transplant-related mortality are also reviewed.

VL - 16 IS - 8 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26266406?dopt=Abstract ER - TY - JOUR T1 - Fate of lymphocytes after withdrawal of tofacitinib treatment. JF - PLoS One Y1 - 2014 A1 - Piscianz, Elisa A1 - Valencic, Erica A1 - Cuzzoni, Eva A1 - De Iudicibus, Sara A1 - De Lorenzo, Elisa A1 - Decorti, Giuliana A1 - Tommasini, Alberto KW - Antigens, CD KW - B-Lymphocytes KW - CD4-Positive T-Lymphocytes KW - CD8-Positive T-Lymphocytes KW - Cell Proliferation KW - Drug Administration Schedule KW - Humans KW - Janus Kinase 3 KW - Killer Cells, Natural KW - Lymphocyte Activation KW - Lymphocyte Count KW - Phytohemagglutinins KW - Piperidines KW - Primary Cell Culture KW - Protein Kinase Inhibitors KW - Pyrimidines KW - Pyrroles AB -

Tofacitinib (Tofa) is an inhibitor of Janus Kinase 3, developed for the treatment of autoimmune diseases and for the prevention of transplant rejection. Due to its selective action on proliferating cells, Tofa can offer a way to block T cell activation, without toxic effects on resting cells. However, few studies have investigated the effects of Tofa on lymphocyte activation in vitro. Our aim was to study the action of Tofa on different lymphocyte subsets after in vitro stimulation and to track the behaviour of treated cells after interruption of the treatment. Peripheral blood lymphocytes were stimulated in vitro with mitogen and treated with two concentrations of Tofa. After a first period in culture, cells were washed and further incubated for an additional time. Lymphocyte subsets, activation phenotype and proliferation were assessed at the different time frames. As expected, Tofa was able to reduce the activation and proliferation of lymphocytes in the first four days of treatment. In addition the drug led to a relative decrease of Natural Killer, B cells and CD8 T cells compared to CD4 T cells. However, treated cells were still viable after the first period in culture and begun to proliferate, strikingly, in a dose dependent manner when the drug was removed from the environment by replacing the culture medium. This novel data does not necessarily predict a similar behaviour in vivo, but can warn about the clinical use of this drug when a discontinuation of treatment with Tofa is considered for any reason.

VL - 9 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/24416411?dopt=Abstract ER - TY - JOUR T1 - Pharmacogenetics of azathioprine in inflammatory bowel disease: a role for glutathione-S-transferase? JF - World J Gastroenterol Y1 - 2014 A1 - Stocco, Gabriele A1 - Pelin, Marco A1 - Franca, Raffaella A1 - De Iudicibus, Sara A1 - Cuzzoni, Eva A1 - Favretto, Diego A1 - Martelossi, Stefano A1 - Ventura, Alessandro A1 - Decorti, Giuliana KW - 6-Mercaptopurine KW - Animals KW - Apoptosis KW - Azathioprine KW - Glutathione KW - Glutathione Transferase KW - Humans KW - Immunosuppressive Agents KW - Inflammatory Bowel Diseases KW - Oxidative Stress KW - Pharmacogenetics KW - Polymorphism, Genetic AB -

Azathioprine is a purine antimetabolite drug commonly used to treat inflammatory bowel disease (IBD). In vivo it is active after reaction with reduced glutathione (GSH) and conversion to mercaptopurine. Although this reaction may occur spontaneously, the presence of isoforms M and A of the enzyme glutathione-S-transferase (GST) may increase its speed. Indeed, in pediatric patients with IBD, deletion of GST-M1, which determines reduced enzymatic activity, was recently associated with reduced sensitivity to azathioprine and reduced production of azathioprine active metabolites. In addition to increase the activation of azathioprine to mercaptopurine, GSTs may contribute to azathioprine effects even by modulating GSH consumption, oxidative stress and apoptosis. Therefore, genetic polymorphisms in genes for GSTs may be useful to predict response to azathioprine even if more in vitro and clinical validation studies are needed.

VL - 20 IS - 13 U1 - http://www.ncbi.nlm.nih.gov/pubmed/24707136?dopt=Abstract ER - TY - JOUR T1 - TNF-α SNP rs1800629 and risk of relapse in childhood acute lymphoblastic leukemia: relation to immunophenotype. JF - Pharmacogenomics Y1 - 2014 A1 - Franca, Raffaella A1 - Rebora, Paola A1 - Athanasakis, Emmanouil A1 - Favretto, Diego A1 - Verzegnassi, Federico A1 - Basso, Giuseppe A1 - Tommasini, Alberto A1 - Valsecchi, Maria Grazia A1 - Decorti, Giuliana A1 - Rabusin, Marco KW - Adolescent KW - Antineoplastic Agents KW - Child KW - Child, Preschool KW - Drug Resistance, Neoplasm KW - Female KW - Genotype KW - Humans KW - Infant KW - Leukemia, Lymphocytic, Chronic, B-Cell KW - Male KW - Phenotype KW - Precursor Cell Lymphoblastic Leukemia-Lymphoma KW - Precursor T-Cell Lymphoblastic Leukemia-Lymphoma KW - Recurrence KW - Risk Assessment KW - Steroids KW - Tumor Necrosis Factor-alpha AB -

AIM: In the AIEOP-BFM ALL (Associazione Italiana Ematologia Oncologia Pediatrica-Berlin Frankfurt Münster acute lymphoblastic leukemia) 2000 protocol, 70% of relapsed patients had favorable prognostic features and fell within less intensive polychemotherapeutic regimens, suggesting the need for better assessing lower risk stratification.

MATERIALS & METHODS: A novel two-phase study design selected 614 children to be genotyped for TNF-α SNP rs1800629 (-308G>A). A weighted Cox model was applied to evaluate the SNP effect on hazard of relapse, adjusting for immunophenotype, risk group, age and gender and including interaction terms.

RESULTS: Significant interaction was found with immunophenotypes (p = 0.0007, with minor allele genotypes being adverse genetic markers in B-cell acute lymphoblastic leukemia and protective ones in T-cell acute lymphoblastic leukemia), and also with risk protocols (p = 0.0041, with minor allele genotypes as prognostic factor of relapse for standard risk patients [only one T-cell acute lymphoblastic leukemia in the subgroup analyzed]).

CONCLUSION: The presence of at least one A allele in TNF-α SNP rs1800629 should suggest a closer monitoring in B-cell acute lymphoblastic leukemia standard risk patients.

VL - 15 IS - 5 U1 - http://www.ncbi.nlm.nih.gov/pubmed/24798719?dopt=Abstract ER - TY - JOUR T1 - Association between BclI polymorphism in the NR3C1 gene and in vitro individual variations in lymphocyte responses to methylprednisolone. JF - Br J Clin Pharmacol Y1 - 2012 A1 - Cuzzoni, Eva A1 - De Iudicibus, Sara A1 - Bartoli, Fiora A1 - Ventura, Alessandro A1 - Decorti, Giuliana KW - Adaptor Proteins, Signal Transducing KW - Adolescent KW - Adult KW - Apoptosis Regulatory Proteins KW - Cyclin D1 KW - Dose-Response Relationship, Drug KW - Female KW - Genotype KW - Glucocorticoids KW - Humans KW - Lymphocytes KW - Male KW - Methylprednisolone KW - Middle Aged KW - Polymorphism, Single Nucleotide KW - Receptors, Glucocorticoid KW - Young Adult AB -

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT: In vitro lymphocyte steroid sensitivity has been suggested as a useful tool to predict in vivo response to glucocorticoid treatment in different inflammatory chronic diseases. A correlation between genetic polymorphisms and clinical response to glucocorticoids has been demonstrated in these patients.

WHAT THIS STUDY ADDS: The BclI polymorphism in the glucocorticoid receptor (NR3C1) gene is associated with higher methylprednisolone potency in vitro. The combined evaluation of the in vitro sensitivity to methylprednisolone and BclI polymorphism could represent an aid for physicians to adjust therapy a priori. AIM To evaluate the association between the in vitro sensitivity of peripheral blood mononuclear cells (PBMCs) to methylprednisolone (MP) and the presence of genetic polymorphisms involved in glucocorticoid (GC) response.

METHODS: In vitro MP inhibition of the proliferation of lymphocytes stimulated with concanavalin A was determined. Non linear regression of dose-response data was performed computing the MP concentration required to reduce proliferation to 50% (IC(50) ). The maximum inhibition achievable at the highest MP concentration (I(max) ) was also calculated. Moreover, the Taqman technique was used to analyze the BclI polymorphism in the NR3C1 gene and the Leu155His polymorphism in the NALP1 gene.

RESULTS: A significant association between the BclI mutated genotype and an increased in vitro sensitivity to GCs was observed.

CONCLUSIONS: The a priori evaluation of the BclI polymorphism, associated with a lymphocyte proliferation assay, could represent a useful diagnostic tool for the optimization of steroid treatment.

VL - 73 IS - 4 U1 - http://www.ncbi.nlm.nih.gov/pubmed/22008062?dopt=Abstract ER - TY - JOUR T1 - The effect of clodronate on a mevalonate kinase deficiency cellular model. JF - Inflamm Res Y1 - 2012 A1 - Zanin, Valentina A1 - Marcuzzi, Annalisa A1 - Piscianz, Elisa A1 - Vuch, Josef A1 - Bianco, Anna Monica A1 - Monasta, Lorenzo A1 - Decorti, Giuliana A1 - Crovella, Sergio KW - Adolescent KW - Adult KW - Alendronate KW - Animals KW - Anti-Inflammatory Agents KW - Apoptosis KW - Cell Line KW - Cells, Cultured KW - Child KW - Clodronic Acid KW - Female KW - Humans KW - Inflammation KW - Lipopolysaccharides KW - Male KW - Mevalonate Kinase Deficiency KW - Mice KW - Models, Biological KW - Monocytes KW - Nitric Oxide KW - Young Adult AB -

BACKGROUND: A potential anti-inflammatory effect of clodronate--an aminobisphosphonate--was described to antagonize the pro-inflammatory effects of the block in the mevalonate pathway, the main feature of a rare auto-inflammatory disease called mevalonate kinase deficiency (MKD).

OBJECTIVE: In this study we evaluated the potential anti-inflammatory effect of clodronate in MKD--a still orphan drug pediatric disease.

METHODS: We studied some biological parameters, nitric oxide production using Griess reagents and programmed cell death by flow cytometry, as common inflammatory parameters in MKD, in the presence of different doses of clodronate (1, 10 and 100 μM).

RESULTS: In our cellular model and in monocytes from patients with MKD, clodronate induced an increase in programed cell death and nitric oxide production in comparison with non-treated cells.

CONCLUSION: Our findings suggest that clodronate does not have an anti-inflammatory effect as previously reported but that it increases the epiphenomena of this pediatric disease.

VL - 61 IS - 12 U1 - http://www.ncbi.nlm.nih.gov/pubmed/22851203?dopt=Abstract ER - TY - JOUR T1 - Personalized therapies in pediatric inflammatory and autoimmune diseases. JF - Curr Pharm Des Y1 - 2012 A1 - Stocco, Gabriele A1 - De Iudicibus, Sara A1 - Franca, Raffaella A1 - Addobbati, Riccardo A1 - Decorti, Giuliana KW - Arthritis, Rheumatoid KW - Autoimmune Diseases KW - Child KW - Genetic Predisposition to Disease KW - Humans KW - Immunogenetic Phenomena KW - Individualized Medicine KW - Inflammation KW - Inflammatory Bowel Diseases KW - Pharmacogenetics AB -

Pediatric inflammatory and autoimmune diseases are a wide array of systemic or organ-specific conditions, characterized by an exaggerated immune reactivity, which generally occurs in immunogenetically predisposed children. Among the most important ones, in terms of their diffusion and morbidity in the population worldwide, pediatric inflammatory bowel disease (IBD) and juvenile rheumatoid arthritis (JRA) have to be considered. The aim of personalized therapy is to give to each patient the most appropriate drug and dose regimen, in order to maximize treatment response and reduce the risk of adverse events. In general, several therapeutic options exist for pediatric inflammatory and autoimmune conditions, therefore the perspective of pharmacological tools that allow identification of patients with increased risk of treatment issues related to a particular medication, in terms of lack of efficacy or increased probability of adverse events, is particularly desirable and promising. The present review will be focused on the personalized therapy approaches already available or in development for pediatric patients with IBD or JRA, comprising pharmacokinetic, pharmacodynamic and pharmacogenetic assays.

VL - 18 IS - 35 U1 - http://www.ncbi.nlm.nih.gov/pubmed/22726111?dopt=Abstract ER - TY - JOUR T1 - Differential action of 3-hydroxyanthranilic acid on viability and activation of stimulated lymphocytes. JF - Int Immunopharmacol Y1 - 2011 A1 - Piscianz, Elisa A1 - Cuzzoni, Eva A1 - De Iudicibus, Sara A1 - Valencic, Erica A1 - Decorti, Giuliana A1 - Tommasini, Alberto KW - 3-Hydroxyanthranilic Acid KW - Boronic Acids KW - Cell Survival KW - Cells, Cultured KW - Humans KW - Lymphocyte Activation KW - Lymphocytes KW - Manganese KW - Pyrazines AB -

Lymphocytes proliferation after antigen-driven activation leads to an increase in cell count, which could last some week, until apoptosis mechanisms allow the homeostatic control of the system. During the first days of this stimulation, activated lymphocytes display high resistance to apoptosis and to most immunosuppressive drugs. According to the literature, few compounds have been described to kill recently activated cells, by inhibiting metabolic processes fundamental to proliferation. The aim of our work was to evaluate comparatively these different compounds, in order to identify the best strategy to kill cells that have undergone proliferation, while sparing the repertoire of resting cells. After preliminary experiments, 3-HAA and bortezomib were selected as the most suitable compounds for our purposes. The possible synergic effect of 3-HAA with bortezomib or with manganese ions was also assessed. 3-HAA was confirmed to be the most reliable pharmacologic approach to inhibit proliferation with acceptable toxicity on resting cells. While in the case of PHA stimulation 3-HAA led to death of most lymphocytes, only a minor percentage of cells were killed after allo-stimulation, suggesting that the effect is proportional to the percentage of stimulated lymphocytes. Manganese ions further enhanced this effect, while results with bortezomib seemed to be less consistent. These results deserve further investigations to develop new procedures for targeting activated cells with pharmacological approaches.

VL - 11 IS - 12 U1 - http://www.ncbi.nlm.nih.gov/pubmed/21979495?dopt=Abstract ER - TY - JOUR T1 - The farnesyltransferase inhibitors tipifarnib and lonafarnib inhibit cytokines secretion in a cellular model of mevalonate kinase deficiency. JF - Pediatr Res Y1 - 2011 A1 - Marcuzzi, Annalisa A1 - De Leo, Luigina A1 - Decorti, Giuliana A1 - Crovella, Sergio A1 - Tommasini, Alberto A1 - Pontillo, Alessandra KW - Alendronate KW - Animals KW - Anti-Inflammatory Agents KW - Cell Line KW - Child KW - Child, Preschool KW - Cholesterol KW - Cytokines KW - Dose-Response Relationship, Drug KW - Enzyme Inhibitors KW - Farnesyltranstransferase KW - Humans KW - Inflammation Mediators KW - Lovastatin KW - Male KW - Mevalonate Kinase Deficiency KW - Mice KW - Monocytes KW - Phosphotransferases (Alcohol Group Acceptor) KW - Piperidines KW - Polyenes KW - Polyisoprenyl Phosphates KW - Polyunsaturated Alkamides KW - Pyridines KW - Quinolones KW - Terpenes AB -

The shortage of geranylgeranyl-pyrophosphate (GGPP) was associated to an increased IL-1β release in the autoinflammatory syndrome mevalonate kinase deficiency (MKD), a rare inherited disease that has no specific therapy. Farnesyltransferase inhibitors (FTIs) act at the end of mevalonate pathway. Two FTIs, tipifarnib (Tip) and lonafarnib (Lon), were therefore evaluated as possible therapeutical choices for the treatment of MKD. FTIs could lead to a redirection of the limited available number of mevalonate intermediates preferentially to GGPP synthesis, eventually preventing the uncontrolled inflammatory response. The effect of Tip and Lon on intracellular cholesterol level (ICL) and on proinflammatory cytokines secretion was evaluated in a cellular model of MKD, chemically obtained treating RAW 264.7 cells with lovastatin (Lova) and alendronate (Ald). The combination of FTIs with the isoprenoid geraniol (GOH) was also tested both in this model and in monocytes isolated from MKD patients. Tip and Lon proved to revert the ICL lowering and to significantly reduce the lipopolysaccharide-induced cytokines secretion in Ald-Lova -RAW 264.7 cells. This anti-inflammatory effect was amplified combining the use of GOH with FTIs. The effect of GOH and Tip was successfully replicated in MKD patients' monocytes. Tip and Lon showed a dramatic anti-inflammatory effect in monocytes where mevalonate pathway was chemically or genetically impaired.

VL - 70 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/21430599?dopt=Abstract ER - TY - JOUR T1 - Genetic predictors of glucocorticoid response in pediatric patients with inflammatory bowel diseases. JF - J Clin Gastroenterol Y1 - 2011 A1 - De Iudicibus, Sara A1 - Stocco, Gabriele A1 - Martelossi, Stefano A1 - Londero, Margherita A1 - Ebner, Egle A1 - Pontillo, Alessandra A1 - Lionetti, Paolo A1 - Barabino, Arrigo A1 - Bartoli, Fiora A1 - Ventura, Alessandro A1 - Decorti, Giuliana KW - Adolescent KW - Child KW - Drug Resistance KW - Female KW - Follow-Up Studies KW - Genotype KW - Glucocorticoids KW - Humans KW - Inflammatory Bowel Diseases KW - Male KW - Multivariate Analysis KW - Polymorphism, Genetic KW - Receptors, Glucocorticoid KW - Regression Analysis KW - Retrospective Studies KW - Sex Factors KW - Treatment Outcome AB -

BACKGROUND: Glucocorticoids (GCs) are used in moderate-to-severe inflammatory bowel diseases (IBD) but their effect is often unpredictable.

AIM: To determine the influence of 4 polymorphisms in the GC receptor [nuclear receptor subfamily 3, group C, member 1 (NR3C1)], interleukin-1β (IL-1β), and NACHT leucine-rich-repeat protein 1 (NALP1) genes, on the clinical response to steroids in pediatric patients with IBD.

METHODS: One hundred fifty-four young IBD patients treated with GCs for at least 30 days and with a minimum follow-up of 1 year were genotyped. The polymorphisms considered are the BclI in the NR3C1 gene, C-511T in IL-1β gene, and Leu155His and rs2670660/C in NALP1 gene. Patients were grouped as responder, dependant, and resistant to GCs. The relation between GC response and the genetic polymorphisms considered was examined using univariate, multivariate, and Classification and Regression Tree (CART) analysis.

RESULTS: Univariate analysis showed that BclI polymorphism was more frequent in responders compared with dependant patients (P=0.03) and with the combined dependant and resistant groups (P=0.02). Moreover, the NALP1 Leu155His polymorphism was less frequent in the GC responsive group compared with resistant (P=0.0059) and nonresponder (P=0.02) groups. Multivariate analysis comparing responders and nonresponders confirmed an association between BclI mutated genotype and steroid response (P=0.030), and between NALP1 Leu155His mutant variant and nonresponders (P=0.033). An association between steroid response and male sex was also observed (P=0.034). In addition, Leu155His mutated genotype was associated with steroid resistance (P=0.034). Two CART analyses supported these findings by showing that BclI and Leu155His polymorphisms had the greatest effect on steroid response (permutation P value=0.046). The second CART analysis also identified age of disease onset and male sex as important variables affecting response.

CONCLUSIONS: These results confirm that genetic and demographic factors may affect the response to GCs in young patients with IBD and strengthen the importance of studying high-order interactions for predicting response.

VL - 45 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/20697295?dopt=Abstract ER - TY - JOUR T1 - Molecular mechanism of glucocorticoid resistance in inflammatory bowel disease. JF - World J Gastroenterol Y1 - 2011 A1 - De Iudicibus, Sara A1 - Franca, Raffaella A1 - Martelossi, Stefano A1 - Ventura, Alessandro A1 - Decorti, Giuliana KW - Drug Resistance KW - Glucocorticoids KW - Humans KW - Inflammatory Bowel Diseases KW - P-Glycoproteins KW - Polymorphism, Genetic KW - Receptors, Glucocorticoid KW - Signal Transduction KW - Transcription, Genetic AB -

Natural and synthetic glucocorticoids (GCs) are widely employed in a number of inflammatory, autoimmune and neoplastic diseases, and, despite the introduction of novel therapies, remain the first-line treatment for inducing remission in moderate to severe active Crohn's disease and ulcerative colitis. Despite their extensive therapeutic use and the proven effectiveness, considerable clinical evidence of wide inter-individual differences in GC efficacy among patients has been reported, in particular when these agents are used in inflammatory diseases. In recent years, a detailed knowledge of the GC mechanism of action and of the genetic variants affecting GC activity at the molecular level has arisen from several studies. GCs interact with their cytoplasmic receptor, and are able to repress inflammatory gene expression through several distinct mechanisms. The glucocorticoid receptor (GR) is therefore crucial for the effects of these agents: mutations in the GR gene (NR3C1, nuclear receptor subfamily 3, group C, member 1) are the primary cause of a rare, inherited form of GC resistance; in addition, several polymorphisms of this gene have been described and associated with GC response and toxicity. However, the GR is not self-standing in the cell and the receptor-mediated functions are the result of a complex interplay of GR and many other cellular partners. The latter comprise several chaperonins of the large cooperative hetero-oligomeric complex that binds the hormone-free GR in the cytosol, and several factors involved in the transcriptional machinery and chromatin remodeling, that are critical for the hormonal control of target genes transcription in the nucleus. Furthermore, variants in the principal effectors of GCs (e.g. cytokines and their regulators) have also to be taken into account for a comprehensive evaluation of the variability in GC response. Polymorphisms in genes involved in the transport and/or metabolism of these hormones have also been suggested as other possible candidates of interest that could play a role in the observed inter-individual differences in efficacy and toxicity. The best-characterized example is the drug efflux pump P-glycoprotein, a membrane transporter that extrudes GCs from cells, thereby lowering their intracellular concentration. This protein is encoded by the ABCB1/MDR1 gene; this gene presents different known polymorphic sites that can influence its expression and function. This editorial reviews the current knowledge on this topic and underlines the role of genetics in predicting GC clinical response. The ambitious goal of pharmacogenomic studies is to adapt therapies to a patient's specific genetic background, thus improving on efficacy and safety rates.

VL - 17 IS - 9 U1 - http://www.ncbi.nlm.nih.gov/pubmed/21448414?dopt=Abstract ER - TY - JOUR T1 - Decreased cholesterol levels reflect a consumption of anti-inflammatory isoprenoids associated with an impaired control of inflammation in a mouse model of mevalonate kinase deficiency. JF - Inflamm Res Y1 - 2010 A1 - Marcuzzi, Annalisa A1 - Decorti, Giuliana A1 - Pontillo, Alessandra A1 - Ventura, Alessandro A1 - Tommasini, Alberto KW - Animals KW - Anti-Inflammatory Agents KW - Cholesterol KW - Disease Models, Animal KW - Inflammation KW - Interleukin-1beta KW - Male KW - Mevalonate Kinase Deficiency KW - Mice KW - Mice, Inbred BALB C KW - Random Allocation KW - Terpenes AB -

OBJECTIVE: The aim of this study was to evaluate, in a mouse model of mevalonate kinase deficiency (MKD), the possible link between inflammatory symptoms and serum cholesterol levels.

MATERIALS AND METHODS: Balb/c mice were treated with alendronate and bacterial muramyl dipeptide. Body temperature, interleukin-1 beta (IL-1 beta) secretion and serum cholesterol levels were measured.

RESULTS: An increased production of the pro-inflammatory cytokine IL-1 beta (p < 0.05) and a rise in body temperature (p < 0.05) was observed, while, in parallel, serum cholesterol concentration significantly decreased (p < 0.05). These effects were completely reversed when animals were treated with exogenous isoprenoids.

CONCLUSIONS: In the mouse model of MKD, the inflammatory response is associated with a reduction in cholesterol levels, and hence this parameter could be used as an indicator of isoprenoid consumption. In addition, plant derived isoprenoids could represent candidate treatments for this disease.

VL - 59 IS - 5 U1 - http://www.ncbi.nlm.nih.gov/pubmed/20174853?dopt=Abstract ER - TY - JOUR T1 - Fasting increases tobramycin oral absorption in mice. JF - Antimicrob Agents Chemother Y1 - 2010 A1 - De Leo, Luigina A1 - Di Toro, Nicola A1 - Decorti, Giuliana A1 - Malusà, Noelia A1 - Ventura, Alessandro A1 - Not, Tarcisio KW - Administration, Oral KW - Animals KW - Anti-Bacterial Agents KW - Biological Availability KW - Fasting KW - Injections, Intramuscular KW - Injections, Intravenous KW - Intestinal Absorption KW - Lactulose KW - Male KW - Mice KW - Mice, Inbred BALB C KW - Rhamnose KW - Tobramycin AB -

The pharmacokinetics of the aminoglycoside tobramycin was evaluated after oral administration to fed or fasting (15 h) mice. As expected, under normal feeding conditions, oral absorption was negligible; however, fasting induced a dramatic increase in tobramycin bioavailability. The dual-sugar test with lactulose and l-rhamnose confirmed increased small bowel permeability via the paracellular route in fasting animals. When experiments aimed at increasing the oral bioavailability of hydrophilic compounds are performed, timing of fasting should be extremely accurate.

VL - 54 IS - 4 U1 - http://www.ncbi.nlm.nih.gov/pubmed/20086144?dopt=Abstract ER - TY - JOUR T1 - Oxidative stress-based cytotoxicity of delphinidin and cyanidin in colon cancer cells. JF - Arch Biochem Biophys Y1 - 2010 A1 - Cvorovic, Jovana A1 - Tramer, Federica A1 - Granzotto, Marilena A1 - Candussio, Luigi A1 - Decorti, Giuliana A1 - Passamonti, Sabina KW - Anthocyanins KW - Antioxidants KW - Apoptosis KW - Caco-2 Cells KW - Camptothecin KW - Cell Line, Tumor KW - Colonic Neoplasms KW - Drug Resistance, Neoplasm KW - Glutathione KW - Glutathione Reductase KW - Humans KW - Oxidants KW - Oxidative Stress AB -

Colorectal cancer is the second most frequent cause of cancer death in the western world. Although the prognosis has improved after the introduction of newer anticancer drugs, the treatment of metastatic colorectal cancer still remains a challenge due to a high percentage of drug-resistant tumor forms. We aimed at testing whether anthocyanidins exerted cytotoxicity in primary (Caco-2) and metastatic (LoVo and LoVo/ADR) colorectal cancer cell lines. Both cyanidin and delphinidin, though neither pelargonidin nor malvidin, were cytotoxic in metastatic cells only. The cell line most sensitive to anthocyanidins was the drug-resistant LoVo/ADR. There, cellular ROS accumulation, inhibition of glutathione reductase, and depletion of glutathione could be observed. This suggests that anthocyanidins may be used as sensitizing agents in metastatic colorectal cancer therapy.

VL - 501 IS - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/20494645?dopt=Abstract ER - TY - JOUR T1 - Targeting farnesyl-transferase as a novel therapeutic strategy for mevalonate kinase deficiency: in vitro and in vivo approaches. JF - Pharmacol Res Y1 - 2010 A1 - De Leo, Luigina A1 - Marcuzzi, Annalisa A1 - Decorti, Giuliana A1 - Tommasini, Alberto A1 - Crovella, Sergio A1 - Pontillo, Alessandra KW - Adult KW - Animals KW - Anti-Inflammatory Agents KW - Cells, Cultured KW - Child, Preschool KW - Enzyme Inhibitors KW - Farnesyltranstransferase KW - Female KW - Humans KW - Male KW - Mevalonate Kinase Deficiency KW - Mevalonic Acid KW - Mice KW - Mice, Inbred BALB C KW - Monocytes KW - Polyenes KW - Polyisoprenyl Phosphates KW - Polyunsaturated Alkamides KW - Young Adult AB -

Mevalonate kinase deficiency (MKD) is a rare inborn auto-inflammatory disease due to the impairment of the pathway for the biosynthesis of cholesterol and non-sterol isoprenoids. The shortage of isoprenoids compounds and in particular of geranylgeranylpyrophosphate (GGPP) was recently associated to the MKD characteristic inflammatory attacks. The aim of this study is to demonstrate that the normalization of the mevalonate pathway intermediates levels and in particular of GGPP, through the specific inhibition of farnesyl-transferase (FT) with Manumycin A could ameliorate the inflammatory phenotype of MKD patients. The effect of Manumycin A was first evaluated in MKD mouse and cellular models, chemically obtained using the aminobisphosphonate alendronate (ALD), and then in monocytes isolated from 2 MKD patients. Our findings were compared to those obtained by using natural exogenous isoprenoids (NEIs). Manumycin A was able to significantly reduce the inflammatory marker serum amyloid A in ALD-treated Balb/c mice, as well as IL-1 beta secretion in ALD-monocytes and in MKD patients. These results clearly showed that, through the inhibition of FT, an increased number of mevalonate pathway intermediates could be redirected towards the synthesis of GGPP diminishing the inflammatory response. The importance in limiting the shortage of GGPP was emphasized by the anti-inflammatory effect of NEIs that, due to their biochemical structure, can enter the MKD pathway. In conclusion, manumycin A, as well as NEIs, showed anti-inflammatory effect in MKD models and especially in MKD-monocytes, suggesting novel approaches in the treatment of MKD, an orphan disease without any efficacious treatment currently available.

VL - 61 IS - 6 U1 - http://www.ncbi.nlm.nih.gov/pubmed/20206266?dopt=Abstract ER - TY - JOUR T1 - Usefulness of the measurement of azathioprine metabolites in the assessment of non-adherence. JF - J Crohns Colitis Y1 - 2010 A1 - Stocco, Gabriele A1 - Londero, Margherita A1 - Campanozzi, Angelo A1 - Martelossi, Stefano A1 - Marino, Sara A1 - Malusà, Noelia A1 - Bartoli, Fiora A1 - Decorti, Giuliana A1 - Ventura, Alessandro KW - 6-Mercaptopurine KW - Adolescent KW - Azathioprine KW - Child KW - Child, Preschool KW - Female KW - Guanine Nucleotides KW - Hepatitis, Autoimmune KW - Humans KW - Immunosuppressive Agents KW - Inflammatory Bowel Diseases KW - Male KW - Medication Adherence KW - Thionucleotides KW - Young Adult AB -

Azathioprine is a thiopurine immunosuppressive antimetabolite used to chronically treat inflammatory bowel disease and autoimmune hepatitis. Azathioprine treatment is a long-term therapy and therefore it is at risk for non-adherence, which is considered an important determinant of treatment inefficacy. Measurement of 6-thioguanine and 6-methylmercaptopurine nucleotides has been recently suggested as a screener for non-adherence detection. We describe four young patients in which non-adherence to azathioprine therapy was detected only through the measurement of drug metabolite concentrations, and the criterion for non-adherence was undetectable metabolite levels. After the identification of non-adherence, patients and their families were approached and the importance of a correct drug administration was thoroughly enlightened and discussed; this allowed obtaining a full remission in all subjects. Our observations support the use of undetectable metabolite levels as indicators of non-adherence to therapy in azathioprine treated patients. The additional level of medical supervision given by this assay allows getting a better adherence to medical treatment, which results in an improvement in the response to therapy; these benefits may justify the costs associated with the assay.

VL - 4 IS - 5 U1 - http://www.ncbi.nlm.nih.gov/pubmed/21122567?dopt=Abstract ER -