title: Combined polymerase chain reaction methods to detect c-myc/IgH rearrangement in childhood Burkitt's lymphoma for minimal residual disease analysis.
abstract: OBJECTIVE
  The translocation t(8;14)(q24;q32), involving the c-myc gene (8q24) and the immunoglobulin heavy chain (IgH) locus (14q32), represents about 75% of all chromosomal translocations in Burkitt's lymphoma (BL). Due to the large variability of the breakpoint region, only the recently improved long-distance LD-polymerase chain reaction (PCR) allows the specific c-myc/IgH fusion to be identified at the genomic level. The sensitivity of the LD-PCR is only 10(-2) to 10(-3) due to the relatively large size of the amplification products (1 to 10 kbp). We, therefore, established a more sensitive nested PCR with a specific primer combination for each patient based on sequence analysis of the variant breakpoint regions.
  
  
  METHODS
  Using the combined PCR methods, we analyzed bone marrow and peripheral blood without visible blasts at diagnosis from 18 patients with t(8;14)-positive BL.
  
  
  RESULTS
  In tests employing dilutions of genomic DNA from the BL cell line CA-46 in the T-cell lymphoma cell line KARPAS-299, which lacks the t(8;14), the sensitivity increased 100-fold, to 10(-5). However, the investigation of 18 c-myc/IgH-positive BL patients with each breakpoint-specific nested PCR showed an inter-patient variability of sensitivity between 10(-3) and 10(-5). Using this assay, the rearrangement was detected in 4/16 bone marrow samples and in 6/15 peripheral blood samples without visible blasts at diagnosis.
  
  
  CONCLUSIONS
  Using the combined PCR methods the detection of c-myc/IgH reaches a level of sensitivity required for the evaluation of minimal residual disease (MRD) in BL patients. Furthermore, the results highlight the importance of verifying the patient-specific sensitivity level for individual MRD monitoring.
authors.affiliation: Dept. of Pediatric Hematology and Oncology, Children's University Hospital, Giessen, Germany. kerstin.busch@paediat.med.uni-giessen.de
authors.forename: Kerstin
authors.initials: K
authors.lastname: Busch
authors.forename: Arndt
authors.initials: A
authors.lastname: Borkhardt
authors.forename: Wilhelm
authors.initials: W
authors.lastname: Wössmann
authors.forename: Alfred
authors.initials: A
authors.lastname: Reiter
authors.forename: Jochen
authors.initials: J
authors.lastname: Harbott
mesh_terms: D000293:Adolescent; D002051:Burkitt Lymphoma / Q000150:complications / Q000235:genetics*; D045744:Cell Line, Tumor; D002648:Child; D002675:Child, Preschool; D002883:Chromosomes, Human, Pair 14 / Q000235:genetics*; D002898:Chromosomes, Human, Pair 8 / Q000235:genetics*; D017931:DNA Primers; D005260:Female; D005803:Genes, Immunoglobulin / Q000235:genetics; D016259:Genes, myc / Q000235:genetics*; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D008297:Male; D018365:Neoplasm, Residual / Q000150:complications / Q000175:diagnosis*; D016133:Polymerase Chain Reaction / Q000379:methods*; D014178:Translocation, Genetic
chemical_list: D017931:DNA Primers; D007143:Immunoglobulin Heavy Chains