Céline Narjoz

Reverse transcriptase-PCR quantification of mRNA levels from cytochrome (CYP)1, CYP2 and CYP3 families in 22 different human tissues.

Objective The aim of this work was to study simultaneously the expression profile of the 23 CYP mRNAs of CYP1, CTP2 and CYP3 families in 22 different human tissues namely adrenal gland, bladder, bone marrow, colon, fetal liver, heart, kidney, liver, lung, mammary gland, ovary, placenta, prostate, salivary gland, skeletal muscle, small intestine, spleen, testis, thymus, thyroid, trachea and uterus. Methods Analysis of the mRNA levels of each of these CYP isoforms was performed on total RNA from pooled specimens of human organs using reverse transcriptase-PCR-based CYP mRNA assays previously validated for their sensitivity and their specificity. Results Our results confirmed previously reported data in the literature concerning isoforms expression in the most currently studied tissues. Moreover, they provided a great deal of new information, mainly about the expression of mRNA of little-known CYP isoforms. Among the 23 CYP isoforms studied, 12 were mainly hepatic (CYP1A2, 2A6, 2A7, 2A13, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, 3A4 and 3A43). Two CYP mRNAs were predominantly expressed in several extrahepatic tissues: CYP1B1 mRNA was the predominant CYP in seven extrahepatic tissues (bone marrow, kidney, mammary gland, prostate, spleen, thyroid and uterus) and CYP2J2 in four extrahepatic tissues (heart, placenta, salivary gland and skeletal muscle). Finally, some CYPs were nearly exclusively expressed in only one extrahepatic tissue. CYP2R1 was found in testis, CYP2U1 in the thymus and CYP2F1 in the respiratory tract (lung and trachea). Conclusion This description will broaden the understanding of the physiological functions of these CYPs.

Early Sorafenib-Induced Toxicity Is Associated with Drug Exposure and UGTIA9 Genetic Polymorphism in Patients with Solid Tumors: A Preliminary Study

Background Identifying predictive biomarkers of drug response is of key importance to improve therapy management and drug selection in cancer therapy. To date, the influence of drug exposure and pharmacogenetic variants on sorafenib-induced toxicity remains poorly documented. The aim of this pharmacokinetic/pharmacodynamic (PK/PD) study was to investigate the relationship between early toxicity and drug exposure or pharmacogenetic variants in unselected adult outpatients treated with single-agent sorafenib for advanced solid tumors. Methods Toxicity was recorded in 54 patients on days 15 and 30 after treatment initiation and sorafenib exposure was assessed in 51 patients. The influence of polymorphisms in CYP3A5, UGT1A9, ABCB1 and ABCG2 was examined in relation to sorafenib exposure and toxicity. Clinical characteristics, drug exposure and pharmacogenetic variants were tested univariately for association with toxicities. Candidate variables with p<0.1 were analyzed in a multivariate analysis. Results Gender was the sole parameter independently associated with sorafenib exposure (p = 0.0008). Multivariate analysis showed that increased cumulated sorafenib (AUCcum) was independently associated with any grade ≥3 toxicity (p = 0.037); UGT1A9 polymorphism (rs17868320) with grade ≥2 diarrhea (p = 0.015) and female gender with grade ≥2 hand-foot skin reaction (p = 0.018). Using ROC curve, the threshold AUCcum value of 3,161 mg/L.h was associated with the highest risk to develop any grade ≥3 toxicity (p = 0.018). Conclusion In this preliminary study, increased cumulated drug exposure and UGT1A9 polymorphism (rs17868320) identified patients at high risk for early sorafenib-induced severe toxicity. Further PK/PD studies on larger population are warranted to confirm these preliminary results.

UGT1A1 genotype and irinotecan therapy: general review and implementation in routine practice

Irinotecan is a major drug in the treatment of advanced colorectal cancer. Its active form is the SN38 metabolite, which is cleared by the biliary route after glucuronidation by uridine diphosphate–glucuronosyltransferase 1A1 (UGT1A1). UGT1A1 activity exhibits a wide intersubject variability, in part related to UGT1A1 gene polymorphisms. The present review on the impact of the deficient UGT1A1*28 variant on irinotecan efficacy and toxicity was produced by a French joint workgroup comprising the Group of Clinical Onco‐pharmacology (GPCO‐Unicancer) and the National Pharmacogenetics Network (RNPGx). It clearly emerges that for irinotecan doses at least equal to 180 mg/m2, patients homozygous for the UGT1A1*28 allele are at increased risk of developing hematological and/or digestive toxicities. Irinotecan dose reduction is thus recommended in homozygous *28/*28 patients. In addition, this personalized medicine strategy aims to secure high‐dose irinotecan administration (≥240 mg/m2) that have proven to be safe in homozygous *1/*1 patients only. The clinical relevance of this test is discussed in terms of treatment efficacy improvement, as increasing the irinotecan dose appears to be safe in patients not bearing a deficient allele. Best execution practices, cost‐effectiveness, and result interpretation are discussed with the aim of facilitating the implementation of this analysis in clinical practice. The existence of networks of laboratories performing this test in routine hospital treatment, as in France, offers the prospect of widespread screening, thus guaranteeing equal access to safe treatment and optimized therapy for patients receiving irinotecan‐based therapy in advanced colorectal cancer.

APOL1 Polymorphisms and Development of CKD in an Identical Twin Donor and Recipient Pair

We report an occurrence of progressive loss of transplant function and ultimately transplant failure after living related kidney transplantation involving monozygotic twin brothers of Afro-Caribbean origin who were both heterozygous for the G1 and G2 kidney disease risk alleles in the APOL1 gene, which encodes apolipoprotein L-I. A 21-year-old man with end-stage kidney disease of unknown cause received a kidney from his brother, who was confirmed as a monozygotic twin by microsatellite analysis. Thirty months after transplantation, the patient presented with proteinuria and decreased estimated glomerular filtration rate; a biopsy of the transplant showed typical focal segmental glomerulosclerosis lesions. He received steroid therapy, but progressed to kidney failure 5 years later. The twin brother had normal kidney function without proteinuria at the time of transplantation; however, 7 years later, he was found to have decreased estimated glomerular filtration rate (40mL/min/1.73m(2)) and proteinuria (protein excretion of 2.5g/d). APOL1 genotyping revealed that both donor and recipient were heterozygous for the G1 and G2 alleles. This case is in stark contrast to the expected course of kidney transplantation in identical twins and suggests a role for APOL1 polymorphisms in both the donor and recipient.

Interindividual variability in TPMT enzyme activity: 10 years of experience with thiopurine pharmacogenetics and therapeutic drug monitoring

BACKGROUND & AIMS TPMT activity and metabolite determination (6-thioguanine nucleotides [6-TGN] and 6-methylmercaptopurine nucleotides [6-MMPN]) remain controversial during thiopurine management. This study assessed associations between patient characteristics and TPMT activity, and their impact on metabolite levels. PATIENTS & METHODS A retrospective review of the laboratory database from a French university hospital identified 7360 patients referred for TPMT phenotype/genotype determination, and/or for 6-TGN/6-MMPN monitoring. RESULTS Four TPMT phenotypes were identified according to TPMT activity distribution: low, intermediate, normal/high and very high. Based on 6775 assays, 6-TGN concentrations were 1.6-fold higher in TPMT-deficient patients compared with TPMT-normal patients. Azathioprine dose and TPMT genotype were significant predictors of metabolite levels. Furthermore, 6-MMPN and 6-MMPN: 6-TGN ratios were, respectively, 1.6- and 2.2-fold higher in females than in males, despite similar TPMT, 6-TGN and azathioprine doses. An unfavorable ratio (≥20) was associated with a slightly higher TPMT activity. CONCLUSION These results illustrate the usefulness of pharmacogenomics and metabolite measurement to improve the identification of noncompliance and patients at high risk for toxicity or therapeutic resistance. Original submitted 13 November 2013; Revision submitted 30 January 2014.

Important role of CYP2J2 in protein kinase inhibitor degradation: a possible role in intratumor drug disposition and resistance.

We have investigated in vitro the metabolic capability of 3 extrahepatic cytochromes P-450, CYP1A1, 1B1 and 2J2, known to be over-expressed in various tumors, to biotransform 5 tyrosine kinase inhibitors (TKI): dasatinib, imatinib, nilotinib, sorafenib and sunitinib. Moreover, mRNA expression of CYP1A1, 1B1, 2J2 and 3A4 in 6 hepatocellular and 14 renal cell carcinoma tumor tissues and their surrounding healthy tissues, was determined. Our results show that CYP1A1, 1B1 and especially 2J2 can rapidly biotransform the studied TKIs with a metabolic efficiency similar to that of CYP3A4. The mRNA expression of CYP1A1, 1B1, 2J2 and 3A4 in tumor biopsies has shown i) the strong variability of CYP expression and ii) distinct outliers showing high expression levels (esp. CYP2J2) that are compatible with high intratumoral CYP activity and tumor-specific TKI degradation. CYP2J2 inhibition could be a novel clinical strategy to specifically increase the intratumoral rather than plasma TKI levels, improving TKI efficacy and extending the duration before relapse. Such an approach would be akin to beta-lactamase inhibition, a classical strategy to avoid antibiotic degradation and resistance.

Genomic consequences of cytochrome P450 2C9 overexpression in human hepatoma cells.

Cytochrome P450 2C9 (P450 2C9) is one of the most important P450 isoforms in the human liver, as it metabolizes numerous exogenous and endogenous substrates. Moreover, it is inducible by several compounds, such as rifampicin, phenobarbital, and NSAIDs (nonsteroidal anti-inflammatories). The aim of this study was to investigate the global cellular consequences of P450 2C9 overexpression at the transcriptional level using an untargeted approach: pangenomic microarrays. Recombinant adenovirus was used to express P450 2C9 instead of an inducer to prevent a per se effect of inducer or its metabolites. P450 2C9 overexpression induced endoplasmic reticulum (ER) stress and regulated genes implicated in the unfolded protein response (UPR) as heat shock protein (HSP) (we studied particurlarly HSPA5 and HSPB1) and in the endoplasmic reticulum associated degradation (ERAD) system as Sec61 and ubiquitin and proteasome pathways. UPR and ERAD are two mechanisms of adaptative response to ER stress. Moreover, activation of Akt was observed in HepG2 cells that overexpress P450 2C9 and might participate in the cellular adaptive response to stress, thus leading to the activation of cell survival pathways. UPR and ERAD should be caused by accumulation of native and misfolded P450 2C9 protein. Our results indicated that P450 2C9 overexpression did not lead to toxicity but induced an ER stress due to protein overexpression rather than mono-oxygenase activity. The ER stress triggered activation of the adaptative response and of pathways leading to cell survival.

Collapsing glomerulopathy associated lupus in a black female with homozygous APOL1 mutation

Collapsing glomerulopathy(CG), characterized by collapse of the glomerular capillary loops onto the mesangial stalks is rarely associated to systemic lupus erythematosus (SLE). Recently a genetic predisposition to HIV associated nephropathy (HIVAN) has been shown in Afro-Americans: MYH9 polymorhism in 2008 and then APOL1 variants (G1 and G2 alleles) in 2010 were shown to be strongly associated with HIVAN. We describe here for the first time the association of CG in a young Afro-American female with SLE having a homozygous mutation of APOL1. The clinical history, laboratory findings and immunofluorescence all confirmed a diagnosis of SLE. However, studies for factors associated with collapsing glomerulopathy in other situations were consistently negative. As this Afro-American patient developed a CG, we performed genotyping of APOL1. It was found that she is homozygotic for the G2 allele of APOL1. Despite

Microsatellite instability analysis in uterine cavity washings as a screening tool for endometrial cancer in Lynch syndrome

Although patients with Lynch syndrome (LS) are at high risk of endometrial cancer, gynecologic screening has been poorly investigated and diagnostic value of current screening tests remains unclear. Microsatellite instability (MSI) phenotype is found in more than 90% of endometrial cancers developed in LS patients. Here we report the first two cases of unstable endometrial tumors with detection of MSI in uterine cavity washings cells. This new technique may be a promising screening tool in LS.

TPMT status determination: the simplest is the most effective?

Dear Sir, We carefully read the article by Hindorf and Appell stating that genotyping should be considered the primary choice for pre-treatment evaluation of thiopurine methyltransferase (TPMT) function.1 As evidence since many years, TPMT status should be determined before a thiopurine treatment to identify patients at risk for severe adverse events, as bone marrow suppression, and to propose an individualized dosage.2 Hindorf and Appell mentioned “it is not reasonable to check both genotype and phenotype in all patients”. We agree …