Jacques Riby

Genome-wide association study of follicular lymphoma identifies a risk locus at 6p21.32

To identify susceptibility loci for non-Hodgkin lymphoma subtypes, we conducted a three-stage genome-wide association study. We identified two variants associated with follicular lymphoma at 6p21.32 (rs10484561, combined P = 1.12 × 10−29 and rs7755224, combined P = 2.00 × 10−19; r2 = 1.0), supporting the idea that major histocompatibility complex genetic variation influences follicular lymphoma susceptibility. We also found confirmatory evidence of a previously reported association between chronic lymphocytic leukemia/small lymphocytic lymphoma and rs735665 (combined P = 4.24 × 10−9).

Genetic variants at 6p21.33 are associated with susceptibility to follicular lymphoma.

We conducted genome-wide association studies of non-Hodgkin lymphoma using Illumina HumanHap550 BeadChips to identify subtype-specific associations in follicular, diffuse large B-cell and chronic lymphocytic leukemia/small lymphocytic lymphomas. We found that rs6457327 on 6p21.33 was associated with susceptibility to follicular lymphoma (FL; N = 189 cases, 592 controls) with validation in another 456 FL cases and 2,785 controls (combined allelic P = 4.7 × 10−11). The region of strongest association overlapped C6orf15 (STG), located near psoriasis susceptibility region 1 (PSORS1).

Regulation of CYP1A1 by indolo [3, 2-b] carbazole in murine hepatoma cells

To determine the basis for unexpected differences in CYP1A1 inducing potencies and efficacies for the diet-derived indole derivative, indolo[3,2-b]carbazole (ICZ) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), we conducted a systematic analysis of events involved in the induced expression of CYP1A1 in murine hepatoma-derived cell lines (Hepa-1). In contrast to the effects of TCDD, induction kinetics and CYP1A1 mRNA half-life were dependent on ICZ concentration, and the response from low doses of inducer was transient due to rapid clearance of ICZ. TCDD and ICZ produced the same maximum response (i.e. equal efficacies) from a TCDD-responsive CAT reporter construct in Hepa-1 cells. When measured by the immediate responses associated with CYP1A1 expression, including cellular uptake of inducer, receptor transformation and binding to DRE (gel mobility shift assay), initiation of transcription (nuclear run-on assay), and short-term accumulation of mRNA (Northern blot assay), ICZ also exhibited an efficacy equal to that of TCDD and a potency that corresponds to its receptor affinity. ICZ is a potent and selective noncompetitive inhibitor of ethoxyresorufin O-deethylase activity (K = 1.5 nM). Taken together these results indicate that ICZ is a bifunctional modulator of CYP1A1 expression with intrinsic efficacy equal to that of TCDD.

GWAS of follicular lymphoma reveals allelic heterogeneity at 6p21.32 and suggests shared genetic susceptibility with diffuse large B-cell lymphoma.

Non-Hodgkin lymphoma (NHL) represents a diverse group of hematological malignancies, of which follicular lymphoma (FL) is a prevalent subtype. A previous genome-wide association study has established a marker, rs10484561 in the human leukocyte antigen (HLA) class II region on 6p21.32 associated with increased FL risk. Here, in a three-stage genome-wide association study, starting with a genome-wide scan of 379 FL cases and 791 controls followed by validation in 1,049 cases and 5,790 controls, we identified a second independent FL–associated locus on 6p21.32, rs2647012 (ORcombined = 0.64, Pcombined = 2×10−21) located 962 bp away from rs10484561 (r2<0.1 in controls). After mutual adjustment, the associations at the two SNPs remained genome-wide significant (rs2647012:ORadjusted = 0.70, Padjusted = 4×10−12; rs10484561:ORadjusted = 1.64, Padjusted = 5×10−15). Haplotype and coalescence analyses indicated that rs2647012 arose on an evolutionarily distinct haplotype from that of rs10484561 and tags a novel allele with an opposite (protective) effect on FL risk. Moreover, in a follow-up analysis of the top 6 FL–associated SNPs in 4,449 cases of other NHL subtypes, rs10484561 was associated with risk of diffuse large B-cell lymphoma (ORcombined = 1.36, Pcombined = 1.4×10−7). Our results reveal the presence of allelic heterogeneity within the HLA class II region influencing FL susceptibility and indicate a possible shared genetic etiology with diffuse large B-cell lymphoma. These findings suggest that the HLA class II region plays a complex yet important role in NHL.

Intestinal absorption of fructose in the rat

The objective of this study was to investigate the mechanisms involved in intestinal absorption of fructose. The results indicate that adult rats readily absorbed 0.4 g of fructose, an amount equivalent to 1.4-1.6 g fructose/kg body wt. Acute malabsorption of fructose occurred with doses greater than 0.6 g (2.1-2.4 g/kg body wt). Continued exposure to dietary fructose resulted in a decrease in the evidence of colonic fermentation. Glucose or galactose administered with fructose enhanced the absorption of fructose. The greatest absorption was observed when equal amounts of fructose and glucose were given simultaneously. If glucose was ingested as a polymer (starch or dextrin), the stimulatory effect was dependent on the digestibility of the polymer. Sucrose given with the fructose and glucose diminished the absorption of fructose. Acarbazone, a specific inhibitor of alpha-glucosidases, including sucrase, also inhibited the facilitating effect of glucose and galactose in absorption of fructose. These results give evidence for joint absorption of the two monosaccharides, fructose and glucose.

Common variation at 6p21.31 (BAK1) influences the risk of chronic lymphocytic leukemia

We performed a meta-analysis of 3 genome-wide association studies to identify additional common variants influencing chronic lymphocytic leukemia (CLL) risk. The discovery phase was composed of genome-wide association study data from 1121 cases and 3745 controls. Replication analysis was performed in 861 cases and 2033 controls. We identified a novel CLL risk locus at 6p21.33 (rs210142; intronic to the BAK1 gene, BCL2 antagonist killer 1; P = 9.47 × 10(-16)). A strong relationship between risk genotype and reduced BAK1 expression was shown in lymphoblastoid cell lines. This finding provides additional support for polygenic inheritance to CLL and provides further insight into the biologic basis of disease development.

ENRICHMENT OF CYSTEINYL ADDUCTS OF HUMAN SERUM ALBUMIN

We report a method to enrich cysteinyl adducts of human serum albumin (HSA), representing biomarkers of exposure to systemic electrophiles. Because the major site of HSA adduction is the single free sulfhydryl group at Cys34, we used thiol-affinity resins to remove mercaptalbumin (i.e., unadducted HSA) from the cysteinyl adducts. Electrospray ionization mass spectrometry was used to detect mercaptalbumin and HSA-Cys34 modifications before and after enrichment of HSA. Differences in adduct content were detected across samples of freshly isolated, archived, and commercial HSA. Cysteinylated and glycosylated adducts were present in all samples, with abundances decreasing in the following order: commercial HSA>archived HSA>fresh HSA. After enrichment of HSA, mercaptalbumin was no longer observed in mass spectra. The ratios of HSA adducts post-/preenrichment, quantified via the Bradford assay and gel electrophoresis, were 0.029 mg adducts/mg HSA in fresh HSA and 0.323 mg adducts/mg HSA in archived HSA. The apparent elevation of adduct levels in archived samples could be due to differences in specimen preparation and storage rather than to differences in circulating HSA adducts. We conclude that thiol-affinity resins can efficiently remove mercaptalbumin from HSA samples prior to characterization and quantitation of protein adducts of reactive systemic electrophiles.

A sandwich enzyme-linked immunosorbent assay for adducts of polycyclic aromatic hydrocarbons with human serum albumin

Adducts of benzo[a]pyrene-diolepoxide (BPDE) with blood nucleophiles have been used as biomarkers of exposure to polycyclic aromatic hydrocarbons (PAHs). The most popular such assay is a competitive enzyme-linked immunosorbent assay (ELISA) that employs monoclonal antibody 8E11 to detect benzo[a]pyrene tetrols following hydrolysis of BPDE adducts from lymphocyte DNA or human serum albumin (HSA). Here we used 8E11 as the capture antibody in a sandwich ELISA to detect BPDE-HSA adducts directly in 1-mg samples of HSA or 20 microl of serum/plasma. The assay employs an anti-HSA antibody for detection, and this is amplified by an avidin/biotinylated horseradish peroxidase complex. The sandwich ELISA has advantages of specificity and simplicity and is approximately 10 times more sensitive than the competitive ELISA. To validate the assay, HSA samples were assayed from three populations with known high PAH exposures (coke oven workers), medium PAH exposures (steel factory control workers), and low PAH exposures (volunteer subjects) (n=30). The respective geometric mean levels of BPDE-HSA adducts--67.8, 14.7, and 1.93 ng/mg HSA (1010, 220, and 28.9 fmol BPDE equiv/mg HSA)--were significantly different (P<0.05). The sandwich ELISA will be useful for screening PAH exposures in large epidemiologic studies and can be extended to other adducts for which capture antibodies are available.

Development of ornithine metabolism in the mouse intestine.

ABSTRACT: Circulating arginine available for synthesis of protein is produced in the kidney of the adult mammal by the action of the last two enzymes of the urea cycle, argininosuccinate synthase and argininosuccinate lyase. In a previous publication, we reported the presence of a complete biosynthetic pathway for arginine in the intestine of the neonatal mouse at a time when no other endogenous sources of arginine were available. Our present study was aimed at the determination of the source of ornithine used by the intestine of the neonatal mouse for the synthesis of arginine. We established the developmental profile of the two intestinal mitochondrial enzymes, pyrroline 5-carbox-ylate synthase and ornithine aminotransferase, responsible for the conversion of glutamate to ornithine. Both enzymatic activities were found to be significantly elevated throughout the suckling period with a peak of activity during the 2nd wk of life. Glutamate dehydrogenase activity in the intestine did not appear to be developmentally regulated during the suckling and weaning periods; therefore, this enzyme was used as a convenient marker to quantify mitochondrial preparations. Ornithine decarboxylase activity was undetectable in the intestine of the mouse during the suckling period and was detected briefly at weaning, indicating that ornithine synthesized in the intestinal mitochondria is probably not diverted actively into the polyamine pathway and is available for synthesis of arginine by the enzymes of the urea cycle.

PRRC2A and BCL2L11 gene variants influence risk of non-Hodgkin lymphoma: results from the InterLymph consortium

Many common genetic variants have been associated with non-Hodgkin lymphoma (NHL), but individual study results are often conflicting. To confirm the role of putative risk alleles in B-cell NHL etiology, we performed a validation genotyping study of 67 candidate single nucleotide polymorphisms within InterLymph, a large international consortium of NHL case-control studies. A meta-analysis was performed on data from 5633 B-cell NHL cases and 7034 controls from 8 InterLymph studies. rs3789068 in the proapoptotic BCL2L11 gene was associated with an increased risk for B-cell NHL (odds ratio = 1.21, P random = 2.21 × 10(-11)), with similar risk estimates for common B-cell subtypes. PRRC2A rs3132453 in the HLA complex class III region conferred a reduced risk of B-cell NHL (odds ratio = 0.68, P random = 1.07 × 10(-9)) and was likewise evident for common B-cell subtypes. These results are consistent with the known biology of NHL and provide insights into shared pathogenic components, including apoptosis and immune regulation, for the major B-cell lymphoma subtypes.