Lack of IKBA coding region mutations in primary mediastinal large B-cell lymphoma and the host response subtype of diffuse large B-cell lymphoma

The role of inhibitor of kappa Bα (IKBA) mutations in lymphoid malignancies with constitutive NF-κB signaling remains to be defined. We recently characterized the molecular signatures of primary mediastinal large B-cell lymphoma (MLBCL) and 3 subtypes of diffuse large B-cell lymphoma (DLBCL).^1,2  The primary MLBCL signature had striking similarities to that of classic Hodgkin lymphoma (cHL), a clinically related disorder.^1,3  Like cHL, primary MLBCL exhibited nuclear localization of the c-REL subunit of NF-κB and increased expression of multiple NF-κB target genes.^1,4  In addition, MLBCL cells transduced with an IκBα superrepressor exhibited markedly decreased NF-κB activity and significantly increased apoptosis, confirming the role of IκBα and the NF-κB pathway in MLBCL cell survival.⁴  Of interest, the newly identified host response (HR) subtype of DLBCL also had significantly increased coordinate expression of multiple NF-κB target genes, implicating the NF-κB survival pathway in this additional subtype of LBCL.^2,4 

Previous studies suggest that DLBCLs that share features with normal in vitro activated B cells (“ABC-like” tumors) also exhibit NF-κB activation and express a more restricted set of NF-κB target genes.^4,5  In addition, “ABC-like” DLBCL cells transduced with an IκBα superrepressor had markedly decreased tumor cell survival.⁵ 

In earlier analyses of potential genetic bases for constitutive NF-κB activation in lymphoid malignancies, somatic mutations of IKBA were described in a subset of cHL.^6-8  In contrast, IKBA mutations were rare in a recently described small series of “ABC-like” DLBCLs.⁹ IKBA mutations were not found in 9 of 10 of such differentiation-associated DLBCLs; a single tumor had both a somatically mutated and wild-type copy of IKBA.⁹ 

To determine whether IKBA mutations were present in primary MLBCLs or HR DLBCLs, we subjected 26 MLBCL and 16 HR DLBCL RNAs to reverse-transcriptase-polymerase chain reaction (RT-PCR) of the entire coding region of IKBA cDNA and sequenced the resulting IKBA PCR products. Only 2 single nucleotide changes in the IKBA coding region (bp 95-1048) were identified (C to T, position 175; and C to T, position 400); neither nucleotide change altered the IKBA coding sequence.¹⁰  Both of the identified single nucleotide changes are recognized IKBA single nucleotide polymorphisms (SNPs; SNP IDs: rs1957106 and rs10782383) (http://www.ncbi.nlm.nih.gov/SNP). The polymorphism at position 175 was detected in 7 of 16 HR tumors, 2 of which were homozygous (T/T), and 12 of 26 MLBCLs, all of which were heterozygous (T/C). The polymorphism at position 400 was detected in 16 of 16 HR tumors, 7 of which were homozygous (C/C), and 25 of 26 MLBCLs, 16 of which were C/C. To exclude the possibility that infiltrating normal cells in primary MLBCLs and HR DLBCLs reduced the sensitivity of the assay, we identified the previously described IKBA mutation in L428 lymphoma cells⁸  using L428 RNA admixed with 4-fold higher concentrations of RNA from a cell line with wild-type IKBA (DHL6).

Taken together, our data and that of Thomas et al⁹  indicate that mutations of IKBA are largely absent in LBCL subtypes, including MLBCL, HR DLBCL, and the differentiation-associated “ABC-like” DLBCLs. These results suggest that alternative mechanisms are likely responsible for the constitutive activation of NF-κB in these tumors.