Therapeutic drug monitoring to improve outcome of anti-TNF drugs in pediatric inflammatory bowel disease

Introduction: Medical treatment of pediatric inflammatory bowel diseases (IBD) has been greatly changed by the introduction of a number of biologic agents that are able to target various players of the immune response. In particular, monoclonal antibodies against the pro-inflammatory cytokine TNF-alpha (TNF) such as infliximab, adalimumab, and golimumab are now in the clinics both in induction and maintenance therapy, and several efforts are currently ongoing to optimize the use of these drugs in children. Areas covered: This review focuses on therapeutic drug monitoring (TDM) of anti-TNF levels and antidrug antibodies (ADAs), in IBD children. A revision of the analytical assays used for assessing anti-TNF plasma levels is also provided. Expert opinion: Although there is a consensus across studies that higher anti-TNF trough levels are associated with a better clinical outcome, and that early anti-TNF serum measurements could be predictive of long-term response, it is still not clear what the best predictive time of sampling is and what the ideal target drug plasma concentration to achieve. Indeed, there are a number of published studies, particularly in pediatric cohorts, limited by the population size analyzed and more prospective large studies are needed to examine the value of these predictive markers

Emerging Insights on the Interaction Between Anticancer and Immunosuppressant Drugs and Intestinal Microbiota in Pediatric Patients

Diseases affecting the immune system, such as inflammatory bowel disease (IBD), juvenile idiopathic arthritis (JIA), and acute lymphoblastic leukemia (ALL), are pathological conditions affecting the pediatric population and are often associated with alterations in the intestinal microbiota, such as a decrease in bacterial diversity. Growing evidence suggests that gut microbiota can interfere with chemotherapeutic and immunosuppressant drugs, used in the treatment of these diseases, reducing or facilitating drug efficacy. In particular, the effect of intestinal microflora through translocation, immunomodulation, metabolism, enzymatic degradation, and reduction of bacterial diversity seems to be one of the reasons of interindividual variability in the therapeutic response. Although the extent of the role of intestinal microflora in chemotherapy and immunosuppression remains still unresolved, current evidence on bacterial compositional shifts will be taken in consideration together with clinical response to drugs for a better and personalized therapy. This review is focused on the effect of the intestinal microbiota on the efficacy of pharmacological therapy of agents used to treat IBD, JIA, and ALL

SERS of cells: What can we learn from cell lysates?

Surface-enhanced Raman spectroscopy (SERS) is a promising and emerging technique to analyze the cellular environment. We developed an alternative, rapid and label-free SERS-based method to get information about the cellular environment by analyzing cells lysates, thus avoiding the need to incorporate nanoparticles into cells. Upon sonicating and filtrating cells, we obtained lysates which, mixed with Au or Ag nanoparticles, yield stable and repeatable SERS spectra, whose overall profile depends on the metal used as substrate, but not on the buffer used for the lysis process. Bands appearing in these spectra were shown to arise mostly from the cytosol and were assigned to adenine, guanine, adenosine and reduced glutathione (GSH). Spectral differences among various cell types also demonstrated that this approach is suitable for cell type identification

Induced pluripotent stem cells for therapy personalization in pediatric patients: Focus on drug-induced adverse events

Adverse drug reactions (ADRs) are major clinical problems, particularly in special populations such as pediatric patients. Indeed, ADRs may be caused by a plethora of different drugs leading, in some cases, to hospitalization, disability or even death. In addition, pediatric patients may respond differently to drugs with respect to adults and may be prone to developing different kinds of ADRs, leading, in some cases, to more severe consequences. To improve the comprehension, and thus the prevention, of ADRs, the set-up of sensitive and personalized assays is urgently needed. Important progress is represented by the possibility of setting up groundbreaking patient-specific assays. This goal has been powerfully achieved using induced pluripotent stem cells (iPSCs). Due to their genetic and physiological species-specific differences and their ability to be differentiated ideally into all tissues of the human body, this model may be accurate in predicting drug toxicity, especially when this toxicity is related to individual genetic differences. This review is an up-to-date summary of the employment of iPSCs as a model to study ADRs, with particular attention to drugs used in the pediatric field. We especially focused on the intestinal, hepatic, pancreatic, renal, cardiac, and neuronal levels, also discussing progress in organoids creation. The latter are three-dimensional in vitro culture systems derived from pluripotent or adult stem cells simulating the architecture and functionality of native organs such as the intestine, liver, pancreas, kidney, heart, and brain. Based on the existing knowledge, these models are powerful and promising tools in multiple clinical applications including toxicity screening, disease modeling, personalized and regenerative medicine

Thiopurine metabolites variations during co-treatment with aminosalicylates for inflammatory bowel disease: effect of N-acetyl transferase polymorphisms

AIM: To evaluate variation of the concentration of thiopurine metabolites after 5-aminosalicylate (5-ASA) interruption and the role of genetic polymorphisms of N-acetyl transferase (NAT) 1 and 2. METHODS: Concentrations of thioguanine nucleotides (TGN) and methymercaptopurine nucleotides (MMPN), metabolites of thiopurines, were measured by high performance liquid chromatography in 12 young patients (3 females and 9 males, median age 16 years) with inflammatory bowel disease (6 Crohn's disease and 6 ulcerative colitis) treated with thiopurines (7 mercaptopurine and 5 azathioprine) and 5-ASA. Blood samples were collected one month before and one month after the interruption of 5-ASA. DNA was extracted and genotyping of NAT1, NAT2, inosine triphosphate pyrophosphatase (ITPA) and thiopurine methyl transferase (TPMT) genes was performed using PCR assays. RESULTS: Median TGN concentration before 5-ASA interruption was 270 pmol/8 x 108 erythrocytes (range: 145-750); after the interruption of the aminosalicylate, a 35% reduction in TGN mean concentrations (absolute mean reduction 109 pmol/8 7 108 erythrocytes) was observed (median 221 pmol/8 7 108 erythrocytes, range: 96-427, P value linear mixed effects model 0.0011). Demographic and clinical covariates were not related to thiopurine metabolites concentrations. All patients were wild-type for the most relevant ITPA and TPMT variants. For NAT1 genotyping, 7 subjects presented an allele combination corresponding to fast enzymatic activity and 5 to slow activity. NAT1 genotypes corresponding to fast enzymatic activity were associated with reduced TGN concentration (P value linear mixed effects model 0.033), putatively because of increased 5-ASA inactivation and consequent reduced inhibition of thiopurine metabolism. The effect of NAT1 status on TGN seems to be persistent even after one month since the interruption of the aminosalicylate. No effect of NAT1 genotypes was shown on MMPN concentrations. NAT2 genotyping revealed that 6 patients presented a genotype corresponding to fast enzymatic activity and 6 to slow activity; NAT2 genotypes were not related to thiopurine metabolites concentration in this study. CONCLUSION: NAT1 genotype affects TGN levels in patients treated with thiopurines and aminosalicylates and could therefore influence the toxicity and efficacy of these drugs; however the number of patients evaluated is limited and this has to be considered a pilot study

Expression of bilitranslocase in the vascular endothelium and its function as a flavonoid transporter

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Azathioprine Biotransformation in Young Patients with Inflammatory Bowel Disease: Contribution of Glutathione-S Transferase M1 and A1 Variants

The contribution of candidate genetic variants involved in azathioprine biotransformation on azathioprine efficacy and pharmacokinetics in 111 young patients with inflammatory bowel disease was evaluated. Azathioprine doses, metabolites thioguanine-nucleotides (TGN) and methylmercaptopurine-nucleotides (MMPN) and clinical effects were assessed after at least 3 months of therapy. Clinical efficacy was defined as disease activity score below 10. Candidate genetic variants (TPMT rs1142345, rs1800460, rs1800462, GSTA1 rs3957357, GSTM1, and GSTT1 deletion) were determined by polymerase chain reaction (PCR) assays and pyrosequencing. Statistical analysis was performed using linear mixed effects models for the association between the candidate variants and the pharmacological variables (azathioprine doses and metabolites). Azathioprine metabolites were measured in 257 samples (median 2 per patient, inter-quartile range IQR 1-3). Clinical efficacy at the first evaluation available resulted better in ulcerative colitis than in Crohn's disease patients (88.0% versus 52.5% responders, p = 0.0003, linear mixed effect model, LME). TGN concentration and the ratio TGN/dose at the first evaluation were significantly higher in responder. TPMT rs1142345 variant (4.8% of patients) was associated with increased TGN (LME p = 0.0042), TGN/dose ratio (LME p < 0.0001), decreased azathioprine dose (LME p = 0.0087), and MMPN (LME p = 0.0011). GSTM1 deletion (58.1% of patients) was associated with a 18.5% decrease in TGN/dose ratio and 30% decrease in clinical efficacy. GSTA1 variant (12.8% of patients) showed a trend (p = 0.049, LME) for an association with decreased clinical efficacy; however, no significant effect on azathioprine pharmacokinetics could be detected. In conclusion, GSTs variants are associated with azathioprine efficacy and pharmacokinetics

Long non-coding RNA gas5 and intestinal mmp2 and mmp9 expression: A translational study in pediatric patients with IBD

Background: The long non-coding RNA (lncRNA) growth arrest–specific transcript 5 (GAS5) seems to be involved in the regulation of mediators of tissue injury, in particular matrix metalloproteinases (MMPs), implicated in the pathogenesis of inflammatory bowel disease (IBD). We investigated the role of GAS5 in regulating MMP2 and MMP9 expression in pediatric patients with IBD and in vitro. Methods: In total, 25 IBD patients were enrolled: For each patient paired inflamed and non-inflamed biopsies were collected. RNA was extracted and GAS5, MMP2, and MMP9 were quantified by TaqMan assay. The expression of GAS5 and MMPs was also determined in the human monocytic THP1 cells differentiated into macrophages and stimulated with lipopolysaccharide (LPS). The function of GAS5 was assessed by overexpressing the lncRNA and evaluating the MMPs levels. Results: Real-time PCR results demonstrated a downregulation of GAS5 and an upregulation of both MMPs in inflamed tissues. In vitro data confirmed the trend observed in patients for the three genes: The stimulation with LPS promoted a downregulation of GAS5 while an increase of MMPs was observed. Overexpression experiments showed that higher levels of GAS5 lead to a decrease of both enzymes. Conclusion: These results provide new information about the role of GAS5 in IBD: The lncRNA could mediate tissue damage by modulating the expression of MMPs