Influence of CYP3A polymorphisms on tacrolimus pharmacokinetics in kidney transplant recipients.

Tacrolimus is characterized by a highly variable pharmacokinetics (PK) and a small therapeutic window. It is metabolized specifically by the CYP3A isoenzymes. This study aimed to determine, in kidney transplant patients, the influence of different genotypic clusters involving these SNPs CYP3A4*1B, CYP3A4*22, and CYP3A5*3 on Tacrolimus bioavailability during the first (PTP1) and the second (PTP2) posttransplant phase (PT). We included kidney transplant patients who received Tacrolimus and underwent drug monitoring by C0 monitoring. CYP3A4 and CYP3A5 genotyping were performed using PCR-RFLP. We classified the patients into four groups: Slow, Intermediate, rapid, and ultra-rapid metabolizers. We included 80 patients. The Tacrolimus dose-normalized C0 (C0/D ratio) was significantly decreased in intermediate, rapid, and ultra-rapid comparing with slow metabolisers. During PTP1 only CYP3A5*3 and CYP3A4*22 polymorphisms correlate significantly with C0/D ratio. Regardless of the PT phase and during the late one, only the CYP3A4 polymorphisms correlate significantly with the C0/D ratio. We identified that these SNPs are all associated independently with Tacrolimus exposure in different PT phases. Moreover, we are the first to define a genotypic cluster including the three CYP3A SNPs.

Development of a population pharmacokinetic model and Bayesian estimators for isoniazid in Tunisian tuberculosis patients.

This study aimed to develop a population pharmacokinetic model using full pharmacokinetic (PK) profiles of isoniazid (INH) taking into account demographic and genetic covariates and to develop Bayesian estimators for predicting INH area under the curve (AUC) in Tunisian tuberculosis patients. The INH concentrations in the building data set were fitted using a one- to three-compartment model. The impact of the different covariates was assessed on the PK parameters of the best model. The best limited sampling strategy (LSS) for estimating the INH AUC was selected by comparing the predicted values to an independent data set. INH PK was best described using a three-compartment model with lag-time absorption. The different studied covariates did not have any impact on the PK parameters of the building model. The Bayesian estimation using one-point concentrations gave the lowest values of prediction errors for the C3 LSS model. This model could be sufficient in routine activity for INH monitoring in this population.

Distribution of Genetic Polymorphisms of Genes Implicated in Thiopurine Drugs Metabolism.

Thiopurine-S-methyltransferase (TPMT) and inosine triphosphate pyrophosphatase (ITPA) are crucial enzymes involved in the metabolism of thiopurine drugs. Significant interethnic variation in the expression of TPMT and ITPA is caused by single nucleotide polymorphisms of genes encoding these proteins. The aim of this study was to describe the distribution of TPMT and ITPA polymorphisms in healthy Tunisian subjects and to establish the metabolizer status of thiopurine drugs in this population. A total of 309 healthy Tunisian subjects were recruited among blood donors of Fattouma Bourguiba Hospital of Monastir. A written informed consent was obtained from all subjects. Whole blood samples were collected from every subject in ethylenediaminetetraacetic acid tubes. TPMT (c.238 G > C, c.460 G > A and c.719A > G) and ITPA (c.94C > A and IVS2+21A > C) mutations were genotyped using polymerase chain reaction-restriction fragment length polymorphism. The observed frequencies of TPMT*3A and TPMT*3C alleles were both 0.8%. The phenotype distribution of TPMT was bimodal: 96.8% of subjects were extensive metabolizers and 3.2% were intermediate metabolizers. Genotyping of ITPA revealed frequencies of 9% and 3% for IVS2+21A > C and c.94C > A mutations, respectively. Accordingly, a trimodal phenotype distribution was found: 75.4% of the subjects were extensive metabolizers, 23.4% were intermediate metabolizers, and 1.2% wereslow metabolizers. Combination of TPMT and ITPA genotyping has revealed that a quarter of the Tunisian Population carries polymorphisms that reduce the metabolic activities of these enzymes.

Poikiloderma with neutropenia in a Tunisian patient with a novel C16orf57 gene mutation.

Poikiloderma with neutropenia (PN) is a genodermatosis characterized by poikiloderma, permanent neutropenia, recurrent infections, nail abnormalities, and palmoplantar hyperkeratosis. We report the case of a Tunisian patient with PN. Skin lesions started from the face and spread to the extremities and trunk. Neutropenia was initially periodic and concomitant with infections periods. DNA analysis identified a novel homozygous deletion of a 1-bp (c.161delC, p.P54RfsX60) in the C16orf57gene, presumed to be causative. This report presents the variability of the clinical manifestations and evolution of PN and emphasizes the importance of studying other patients with PN to better delineate mutations profile among populations.

Clinical profile of comorbidity of rare diseases in a Tunisian patient: a case report associating incontinentia pigmenti and Noonan syndrome.

BACKGROUND: Noonan syndrome (NS) is an autosomal dominant multisystem disorder caused by the dysregulation of several genes belonging to the RAS Mitogen Activated Protein Kinase (MAPK) signaling pathway. Incontinentia Pigmenti (IP) is an X-linked, dominantly inherited multisystem disorder. CASE PRESENTATION: This study is the first report of the coexistence of Noonan (NS) and Incontinentia Pigmenti (IP) syndromes in the same patient. We report on the clinical phenotype and molecular characterization of this patient. The patient was examined by a pluridisciplinary staff of clinicians and geneticist. The clinical diagnosis of NS and IP was confirmed by molecular investigations. The newborn girl came to our clinics due to flagrant dysmorphia and dermatological manifestations. The clinical observations led to characterize the Incontinentia Pigmenti traits and a suspicion of a Noonan syndrome association. Molecular diagnosis was performed by Haloplex resequencing of 29 genes associated with RASopathies and confirmed the NS diagnosis. The common recurrent intragenic deletion mutation in IKBKG gene causing the IP was detected with an improved PCR protocol. CONCLUSION: This is the first report in the literature of comorbidity of NS and IP, two rare multisystem syndromes.

ABCB1 and SLCO1B3 Gene Polymorphisms and Their Impact on Digoxin Pharmacokinetics in Atrial Fibrillation Patients among the Tunisian Population.

BACKGROUND: Digoxin is a substrate of P-glycoprotein (P-gp) and organic anion transporting polypeptide transporters that are encoded by ABCB1 and SLCO1B3 genes. Genetic polymorphisms in both genes may explain inter-individual variability of serum digoxin concentration (SDC). This study evaluates the possible effect of the most common ABCB1 and SLCO1B3 polymorphisms on SDC after a single oral dose of digoxin in Tunisian atrial fibrillation (AF) patients. METHODS: ABCB1 and SLCO1B3 genotypes were analyzed in 102 patients with AF who received digoxin (0.5 mg) without (group I, n = 58) or with the co-administration of P-gp inhibitors (group II, n = 44). SDCs were determined at 6 h following the oral dose. RESULTS: SDCs levels were significantly higher in patients who were co-administered P-gp inhibitors. No influence was noted in ABCB1 and SLCO1B3 polymorphisms on SDC in group I patients. However, SDCs values were significantly different among ABCB1 single nucleotide polymorphisms (SNPs) genotypes of 2677G>T/A (TT, GG>GT, p < 0.05) and 3435C>T (TT, CC>CT, p < 0.05) only in group II with no effect of 1236C>T and SLCO1B3 SNPs. CONCLUSION: Results suggest that P-gp inhibitors and ABCB1 gene polymorphisms may affect digoxin pharmacokinetics.

[Marshall syndrome: Clinical, radiological and genetical features of a Tunisian family]. / Syndrome de Marshall: Aspects cliniques, radiologiques et génétiques d'une famille tunisienne.

BACKGROUND: Marshall syndrome is a rare autosomal dominant skeletal dysplasia. It associates a particular facial dysmorphism with midface hypoplasia, ocular abnormalities and sensorineural hearing loss. It is caused by heterozygous mutations in COL11A1 gene coding the 1 chain of collagen XI. Stickler syndrome is the principal differential diagnosis of Marshall syndrome. AIM: Clinical and radiological study of Marshall syndrome in a Tunisian family with a linkage study of the COL11A1 gene to this disease. METHODS: We report the clinical and the radiological findings of a Tunisian family including 8 members affected by Marshall syndrome. The linkage of the COL11A1 gene to this disease was tested using the polymorphic microsatellite markers of DNA. RESULTS: A variability of the clinical expression of Marshall syndrome was reported. Specific Marshall phenotype and an overlapping phenotype between the Marshall and Stickler syndromes were observed among the affected members of this family. The ocular manifestations were also heterogeneous. Marshall syndrome's specific radiological signs were found. The linkage study supports the linkage of the abnormal phenotype to the COL11A1 gene. CONCLUSION: There is a variability of the clinical expression among the affected members of the study's family. We will continue searching the causative mutation to establish a clear genotype- phenotype correlation.

A novel mutation in FGD4/FRABIN causes Charcot Marie Tooth disease type 4H in patients from a consanguineous Tunisian family.

Charcot-Marie-Tooth (CMT) disease constitutes a clinically and genetically heterogeneous group of hereditary neuropathies characterized by progressive muscular and sensory loss in the distal extremities with chronic distal weakness, deformation of the feet, and loss of deep tendon reflexes. CMT4H is an autosomal recessive demyelinating subtype of CMT, due to mutations in FGD4/FRABIN, for which nine mutations are described to date. In this study, we describe three patients from a consanguineous Tunisian family, presenting with severe, early onset, slowly progressive, autosomal recessive demyelinating CMT, complicated by mild to severe kyphoscoliosis, consistent with CMT4H. In these patients, we report the identification of a novel homozygous frameshift mutation in FGD4: c.514_515insG; p.Ala172Glyfs*27. Our study reports the first mutation identified in FGD4 in Tunisian patients affected with CMT. It further confirms the important clinical heterogeneity observed in patients with mutations in FGD4 and the lack of phenotype/genotype correlations in CMT4H. Our results suggest that FGD4 should be screened in other early-onset CMT subtypes, regardless of the severity of the phenotype, and particularly in patients of consanguineous descent. In Tunisians, as in other populations with high consanguinity rates, screening of genes responsible for rare autosomal recessive CMT subtypes should be prioritized.

A novel CASR mutation in a Tunisian FHH/NSHPT family associated with a mental retardation.

The calcium-sensing receptor (CASR), a plasma membrane G-protein coupled receptor, is expressed in parathyroid gland and kidney, and controls systemic calcium homeostasis. Inactivating CASR mutations have previously been identified in patients with familial hypocalciuric hypercalcemia (FHH) and neonatal severe hyperparathyroidism (NSHPT). The aim of the present study is to determine the underlying molecular defect of FHH/NSHPT disease in a consanguineous Tunisian family. Mutation screening was carried out using RFLP-PCR and direct sequencing. We found that the proband is homozygous for a novel 15 bp deletion in the exon 7 (c.1952_1966del) confirming the diagnosis of NSHPT. All the FHH members were found to be heterozygous for the novel detected mutation. The mutation, p.S651_L655del, leads to the deletion of 5 codons in the second trans-membrane domain of the CASR which is thought to be involved in the processes of ligand-induced signaling. This alteration was associated with the evidence of mental retardation in the FHH carriers and appears to be a novel inactivating mutation in the CASR gene. Our findings provide additional support for the implication of CASR gene in the FHH/NSHPT pathogenesis.

Genotoxic damage in hospital workers exposed to ionizing radiation and metabolic gene polymorphisms.

Of all workers exposed globally to synthetic sources of radiation, medical personnel represent the largest group, but receive relatively low doses. Accidental or therapeutic acute radiation exposure of humans was observed to induce various forms of cytogenetic damage, including the possibility of increasing the incidence of micronuclei (MN) and chromosomal aberrations (CA). The aim of this study was to assess occupationally induced chromosomal damage in a large population of hospital workers exposed to low doses of ionizing radiation (IR). The cytokinesis-block MN and comet assays were used to examine peripheral blood lymphocytes (PBL) of 31 exposed workers to IR and 33 control subjects corresponding in gender, age, and smoking. Glutathione S-transferases (GSTM1, GSTT1, and GSTP1) are postulated to be involved in the detoxification of endogenous and exogenous genotoxicants. The association between these biomarkers and polymorphic genes of xenobiotic metabolizing enzymes was thus also assessed. MN frequency was significantly higher in the exposed subjects compared controls. Comet assay results showed a significant increase of tail length in workers exposed to IR. Data obtained suggest that GSTM1, GSTT1, and GSTP1 polymorphism do not modify significantly the genotoxic potential of IR. Therefore, the exposed medical personnel need to carefully apply radiation protection procedures and minimize, as low as possible, IR exposure to avoid possible genotoxic effects.

Codeine-induced acute generalized exanthematous pustulosis without IL36RN mutations.

Recent studies have suggested an association between mutations in the IL-36RN gene and the onset of pustular generalized. In the literature, only one case of acute generalized exanthematous pustulosis (AGEP) induced by codeine in a patient with IL36RN mutation has been reported. Herein, we reported an unusual case of AGEP caused by codeine in a patient with a history of psoriasis and confirmed by an oral provocation test. In this case, we have shown that the IL36RN gene mutation is not a constant condition in drug-induced AGEP. Clinicians should be aware of this side effect of codeine especially, in patients with a history of psoriasis. More studies are needed to clarify the association between drug-induced AGEP and IL36RN gene mutations.

Interplay of socio-economic factors, consanguinity, fertility, and offspring mortality in Monastir, Tunisia.

AIM: To assess the association among social status, prevalence of consanguineous marriages, and the effects of consanguinity on reproductive behavior and mortality in Tunisia. METHODS: The study included data on a total of 1741 live-births born from November 1989 to October 1990 in the maternity ward of the University-Hospital Fattouma Bourguiba of Monastir, Tunisia. After delivery, women filled out a questionnaire on the age of the parents at marriage, the number of pregnancies and abortions, the number of neonatal and post-neonatal deaths, and deaths of children under 5 years. Three categories of marriages were distinguished as follows: marriages between first cousins, marriages between cousins of other degree, and non consanguineous marriages. RESULTS: Consanguineous marriages represented 432 (24.81%) of the unions. Most consanguineous marriages were contracted between first cousins (n=303; 70.13%). Consanguineous couples had a lower age at marriage and a higher fertility index than non-consanguineous couples. The rates of spontaneous abortions and stillbirths were not correlated with consanguinity. However, higher rates of neonatal and post-neonatal deaths, and deaths of children younger than 5 years were observed in consanguineous couples. CONCLUSION: Fertility index and mortality, especially in the first year of life, were significantly higher in consanguineous marriages. This important socio-economical factor needs to be considered in assessing equity on health in specific social and cultural contexts.

Association among education level, occupation status, and consanguinity in Tunisia and Croatia.

AIM: To investigate the association between education level, occupation status (a proxy for socio-economic status), and consanguinity in 2 large data sets from Tunisia and Croatia countries with different attitudes toward consanguinity. METHODS: The sample of 1016 students, attending 5 university institutions in Monastir, Tunisia, were interviewed about the educational level and occupation status of their parents and the degree of parental relatedness. In Croatia, a sample of 1001 examinees from 9 isolated island populations was interviewed about their own educational level, occupation status, and consanguinity. RESULTS: Prevalence of consanguinity (offspring of second cousins or closer) among 1016 Tunisian students was 20.1%, and 9.3% among 1001 Croatian isolates. In Tunisia, the association between consanguinity and both parental degree of education and parental occupation status was highly significant in women (P<0.001), but not significant in men. In Croatia, no statistically significant associations were noted, although there was a consistent trend of increased prevalence of consanguinity with lower education level or occupation status in both genders, but more pronounced in women. CONCLUSION: Association between education level, socio-economic status, and consanguinity needs to be taken into account in inbreeding studies in human populations. The relationship may be specific for each studied population and highly dependent on the cultural context. It is generally more pronounced among women in most settings.

The effect of titanium dioxide nanoparticles on neuroinflammation response in rat brain.

Titanium dioxide nanoparticles (TiO2 NPs) are widely used for their whiteness and opacity in several applications such as food colorants, drug additives, biomedical ceramic, and implanted biomaterials. Research on the neurobiological response to orally administered TiO2 NPs is still limited. In our study, we investigate the effects of anatase TiO2 NPs on the brain of Wistar rats after oral intake. After daily intragastric administration of anatase TiO2 NPs (5-10 nm) at 0, 50, 100, and 200 mg/kg body weight (BW) for 60 days, the coefficient of the brain, acethylcholinesterase (AChE) activities, the level of interleukin 6 (IL-6), and the expression of glial fibrillary acidic protein (GFAP) were assessed to quantify the brain damage. The results showed that high-dose anatase TiO2 NPs could induce a downregulated level of AChE activities and showed an increase in plasmatic IL-6 level as compared to the control group accompanied by a dose-dependent decrease inter-doses, associated to an increase in the cerebral IL-6 level as a response to a local inflammation in brain. Furthermore, we observed elevated levels of immunoreactivity to GFAP in rat cerebral cortex. We concluded that oral intake of anatase TiO2 NPs can induce neuroinflammation and could be neurotoxic and hazardous to health.

Hematological, biochemical, and toxicopathic effects of subchronic acetamiprid toxicity in Wistar rats.

Acetamiprid is one of the most widely used neonicotinoids. This study investigates toxic effects of repeated oral administration of three doses of acetamiprid (1/20, 1/10, and 1/5 of LD50) during 60 days. For this, male Wistar rats were divided into four different groups. Hematological, biochemical, and toxicopathic effects of acetamiprid were evaluated. According to the results, a significant decrease in the body weight gain at the highest dose 1/5 of LD50 of acetamiprid was noticed. An increase in the relative liver weight was also observed at this dose level. The hematological constituents were affected. A significant decrease in RBC, HGB, and HCT in rats treated with higher doses of acetamiprid (1/10 and 1/5 of LD50) was noted. However, a significant increase in WBC and PLT were observed at the same doses. Furthermore, acetamiprid induced liver toxicity measured by the increased activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphates (ALPs), and lactate dehydrogenase (LDH) which may be due to the loss of hepatic membrane architecture and hepatocellular damage. In addition, exposure to acetamiprid resulted in a significant decrease in the levels of superoxide dismutase and catalase activities (p ≤ 0.01) with concomitant increase in lipid peroxidation in rat liver. These findings highlight the subchronic hepatotoxicity of acetamiprid.

Toxicopathic changes and genotoxic effects in liver of rat following exposure to diazinon.

In general, people may come in contact with mixtures of insecticides through domestic use, consumption of contaminated food or drinks, and/or living close to treated areas. We analyzed the toxic effects of diazinon on histological structure of liver and hematological parameters in male rats. DNA-damaging potential of diazinon was also investigated using the comet assay in blood cells and the micronucleus test in bone marrow. Two groups of six male rats orally received different amounts of diazinon: 1/50 and 1/25 LD 50 for 4 weeks (5 day/week). The present study showed that diazinon caused hypertrophy of sinusoids, central vein, and portal triad, in addition to the formation of oedema, vacuoles, hemorrhage, necrosis, and lymphoid infiltration in rats' liver. A significant decrease in red blood cells, hemoglobin, hematocrite levels, and platelet counts was observed in the treated groups. However, the white blood cell count increased. Micronucleus test results revealed aneugenic effects of diazinon. Furthermore, we noticed an increase in comet tail length in treated groups. So, the comet assay confirmed the genotoxic potential of diazinon in vivo. On the assumption that all alterations observed in rats could be observed in human, it is necessary to raise the awareness about the health risk posed by this insecticide.

Histopathological and genotoxic effects of chlorpyrifos in rats.

This study aims to investigate the effects of chlorpyrifos's sub-acute exposure on male rats. Two groups with six animals each were orally treated, respectively, with 3.1 mg/kg b w and 6.2 mg/kg b w of chlorpyrifos during 4 weeks. The genotoxic effect of chlopyrifos was investigated using the comet assay and the micronucleus test. Some hematological and liver's histopathological changes were also evaluated. Results revealed that chlorpyrifos induced histopathological alterations in liver parenchyma. The lymphoid infiltration observed in liver sections and the increase in white blood cells parameter are signs of inflammation. A significant increase in the platelet' count and in polychromatic erythrocytes/normochromatic erythrocytes (PCE/NCE) ratio was observed in chlorpyrifos-treated groups which could be due to the stimulatory effect of chlorpyrifos on cell formation in the bone marrow at lower doses. In addition, the increase of bone marrow micronucleus percentage and the comet tail length revealed a genotoxic potential of chlorpyrifos in vivo.

Study of the T16189C variant and mitochondrial lineages in Tunisian and overall Mediterranean region.

The mitochondrial DNA (mtDNA) variant T16189C has been investigated in several metabolic diseases. In this study, we aimed to estimate the frequency of the T16189C variant in Tunisian and other Mediterranean populations and to evaluate the impact of this variant on the phylogeny of Mediterranean populations. Blood sample of 240 unrelated Tunisian subjects were recruited from several Tunisian localities. The hypervariable region 1 of the mtDNA were amplified and sequenced. Additional sequences (N = 4921) from Mediterranean populations were compiled from previous studies. The average frequency of T16189C variant in Tunisia (29%) is similar to that observed in North African and Near Eastern populations. Our findings showed positive correlation of the T16189C variant with Sub-Saharan and North African lineages, while a negative correlation was found with the Eurasian haplogroups, reaching its maximum with the Eurasian haplogroup H. The principal component analyses showed a high internal heterogeneity between Tunisian localities. At the Mediterranean scale, Tunisians are closer to North African (Algerian and Moroccan) and Near Eastern populations (Syrians and Palestinians) than to Europeans.

Influence of combined CYP3A4 and CYP3A5 single-nucleotide polymorphisms on tacrolimus exposure in kidney transplant recipients: a study according to the post-transplant phase.

AIM: The present study investigated in Tunisian renal transplant patients, genetic polymorphisms of CYP3A4 -392A>G and CYP3A5 6986A>G and their influence on tacrolimus (Tac) pharmacokinetics during early and late post-transplant (PT) phases and established customized ranges of Tac doses matching the C0 target levels according to CYP3A4 and CYP3A5 genotype combination and the PT phase. PATIENTS & METHODS: We included adult Tunisian patients having received Tac for de novo kidney grafts and undergone a therapeutic drug monitoring of Tac by morning C0 monitoring during early (1 to 90 days) and late (over 90 days) PT phases. The genomic DNA was extracted from peripheral blood mononuclear cells using a salting-out procedure. CYP3A4 promoter (rs2740574; -392A>G) and CYP3A5 (rs776746; 6986A>G) SNP genotyping was analyzed using PCR-RFLP. RESULTS: Fifty-two patients were enrolled in the study. During the early PT phase, the CYP3A5 polymorphism but not that of CYP3A4, correlates significantly with Tac dose-normalized C0 (C0/D ratio). During the late PT phase, the effect of CYP3A4 polymorphism becomes significant and that of CYP3A5 becomes nonsignificant on Tac C0/D Tac. The mean daily doses (mg/kg) matching therapeutic C0, regardless of the CYP3A genotypes, were 0.16 ± 0.05 and 0.10 ± 0.05 during early and late PT phase, respectively. Carriers of the CYP3A4*1B allele require higher doses to maintain the C0 in the therapeutic range during the two PT phases. However, patients carrying the CYP3A5*1 require significant higher Tac doses, only during the early phase. CONCLUSION: Our data support a critical role of the CYP3A5 6986A>G and CYP3A4 -392A>G polymorphisms on the variation of Tac exposure during the early and the late PT phase, respectively. The establishment of customized Tac doses, according to CYP3A4/CYP3A5 genotype combination and the PT time, may allow preventing graft rejection and improving the safety profile of this drug. Further studies are needed to investigate this issue.