title: Improved clonality assessment in germinal centre/post-germinal centre non-Hodgkin's lymphomas with high rates of somatic hypermutation.
abstract: BACKGROUND
  PCR detects clonal rearrangements of the Ig gene in lymphoproliferative disorders. False negativity occurs in germinal centre/post-germinal centre lymphomas (GC/PGCLs) as they display a high rate of somatic hypermutation (SHM), which causes primer mismatching when detecting Ig rearrangements by PCR.
  
  
  OBJECTIVE
  To investigate the degree of SHM in a group of GC/PGCLs and assess the rate of false negativity when using BIOMED-2 PCR when compared with previously published strategies.
  
  
  METHODS
  DNA was isolated from snap-frozen tissue from 49 patients with GC/PGCL (23 diffuse large B cell lymphomas (DLBCLs), 26 follicular lymphomas (FLs)) and PCR-amplified for complete (VDJH), incomplete (DJH) and Ig kappa/lambda rearrangements using the BIOMED-2 protocols, and compared with previously published methods using consensus primers. Germinal centre phenotype was defined by immunohistochemistry based on CD10, Bcl-6 and MUM-1.
  
  
  RESULTS
  Clonality detection by amplifying Ig rearrangements using BIOMED-2 family-specific primers was considerably higher than that found using consensus primers (74% DLBCL and 96% FL vs 69% DLBCL and 73% FL). Addition of BIOMED-2 DJH rearrangements increased detection of clonality by 22% in DLBCL. SHM was present in VDJH rearrangements from all patients with DLBCL (median (range) 5.7% (2.5-13.5)) and FL (median (range) 5.3% (2.3-11.9)) with a clonal rearrangement.
  
  
  CONCLUSIONS
  Use of BIOMED-2 primers has significantly reduced the false negative rate associated with GC/PGCL when compared with consensus primers, and the inclusion of DJH rearrangements represents a potential complementary target for clonality assessment, as SHM is thought not to occur in these types of rearrangements.
authors.affiliation: Haemato-Oncology Laboratory, Department of Haematology, Belfast City Hospital, Belfast, Northern Ireland. mark.catherwood@bll.n-i.nhs.uk
authors.forename: Mark A
authors.initials: MA
authors.lastname: Catherwood
authors.forename: David
authors.initials: D
authors.lastname: Gonzalez
authors.forename: Caroline
authors.initials: C
authors.lastname: Patton
authors.forename: Edwina
authors.initials: E
authors.lastname: Dobbin
authors.forename: Lakshmi
authors.initials: L
authors.lastname: Venkatraman
authors.forename: H Denis
authors.initials: HD
authors.lastname: Alexander
mesh_terms: D004273:DNA, Neoplasm / Q000235:genetics; D054278:Embryonal Carcinoma Stem Cells; D015322:Gene Rearrangement, B-Lymphocyte; D018858:Germinal Center / Q000473:pathology*; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D016393:Lymphoma, B-Cell / Q000235:genetics / Q000276:immunology / Q000473:pathology; D008224:Lymphoma, Follicular / Q000235:genetics / Q000276:immunology / Q000473:pathology; D016403:Lymphoma, Large B-Cell, Diffuse / Q000235:genetics / Q000276:immunology / Q000473:pathology; D008228:Lymphoma, Non-Hodgkin / Q000235:genetics* / Q000276:immunology / Q000473:pathology; D014411:Neoplastic Stem Cells / Q000473:pathology*; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D027041:Somatic Hypermutation, Immunoglobulin
chemical_list: D004273:DNA, Neoplasm; D007143:Immunoglobulin Heavy Chains