Clonotypic B cells in classic Hodgkin lymphoma

To the editor:

Jones and colleagues described 2 Hodgkin lymphoma (HL) cell lines that contain small subpopulations of aldehyde dehydrogenase (ALDH)⁺CD20⁺CD30⁻Igλ⁺ B cells, which have clonogenic potential and give rise to the typical CD30⁺ Hodgkin and Reed-Sternberg (HRS) cells.¹  Importantly, the authors also reported that in the peripheral blood (PB) of HL patients, ALDH^high cells are detectable, that are often clonally related to the HRS cells. These cells may represent HL-initiating cells, perhaps cancer stem cells.

First indications for the existence of HRS cell clone members among small CD30⁻, morphologically normal cells were reported 10 years ago. In several HL the same numerical chromosomal abnormalities were present in HRS cells and in some small (CD30⁻) cells.^2,3  However, such abnormalities are not the best clonal markers in HL.⁴  In another study of HL with Epstein-Barr virus (EBV)⁺ HRS cells, CD30⁻ small EBV⁺ lymphocytes in the HL lymph nodes were not (with perhaps rare exceptions) members of the HRS clone.⁵  As HRS cells show a clonal EBV infection pattern, putative CD30⁻ clone members should also be EBV-infected. Thus, members of the HRS cell clone are usually absent among CD30⁻ B cells, at least in EBV⁺ cases. Vockerodt and colleagues analyzed multiple PB and bone marrow samples from 2 HL patients, using a highly sensitive HRS cell clone–specific PCR, but HRS cell–specific amplificates were undetectable.⁶  Hence, clonotypic cells were very infrequent or absent in the PB of these patients.

Jones and coworkers present 3 pieces of evidence that clonotypic B cells are present in the PB of HL patients among ALDH^highCD27⁺CD19⁺ B cells.¹  However, none of these is convincing. First, in the heavy chain fragment length analysis, the isolated HRS cell population surprisingly gave a polyclonal pattern with many fragments of different lengths, so that also seeing 2 bands with the same length as those obtained from the PB B cells does not prove clonal identity. Second, the fragment length analysis of Vκ light chain rearrangements is unsuitable to demonstrate clonal identity, because the V-J joints of light chain rearrangements show little length variation; 66% of polyclonal VκJκ joints have an identical CDRIII length of 27 bp.⁷  Third, sequences of V(D)J joints are principally an ideal clonal marker for B cells. However, the VκJκ sequences obtained from HRS cells and PB ALDH^highCD27⁺CD19⁺ B cells surprisingly stop 5′ of the V-J joint, so that their potential clonal relationship is not demonstrated. The pattern of somatic mutations in the sequences actually argues against a clonal relationship: as V genes in HRS cells show nearly never intraclonal diversity,⁸  the finding of different mutations only in the Vκ sequences from the HRS and PB B cells suggests that the latter do not carry the clonotypic V gene rearrangement of the HRS cells.

It will also be important in future studies to clarify whether indeed only the CD30⁻ cells in HL cell lines have clonogenic potential, because small mononuclear CD30⁺ Hodgkin cells were not analyzed separately from large multinuclear Reed-Sternberg cells, and mononuclear Hodgkin cells have a high proliferative potential.⁹  Moreover, it will be critical to determine whether B cells in the PB with clonotypic rearrangements—if their existence can be confirmed—are members of the malignant HRS cell clone or represent premalignant B cells that persisted in the patients.