Clonotypic B Cells Circulate in Hodgkin's Lymphoma (HL).

Abstract

Reed-Sternberg (RS) cells, the hallmark of HL, are clonal B lineage cells generally characterized by their co-expression of CD15 and CD30. RS cells also occasionally express the plasma cell (PC) cell surface antigen CD138 (syndecan-1), and we thus hypothesized that RS may represent aberrant PC differentiation. We found that RS cells from the L428 HL cell line exhibited an expression profile similar to normal PC and those from multiple myeloma (MM): high expression of the PC transcription factor Blimp-1 and low expression of the germinal center B cell markers Pax-5 and Bcl-6. We recently demonstrated that CD138+ MM PC, like their normal counterparts, have limited replicative potential; rather, self-renewing CD138neg clonotypic B cells appear to be responsible for the initiation and maintenance of MM, giving rise to the differentiated CD138+ MM PC (Matsui et al Blood 103: 2332–2336, 2004). To determine if RS cells similarly were differentiated progeny of rare HL cancer stem cells, we studied HL cell lines (L428, KM-H2) and found that each line contained a small (<5%) subpopulation of cells that did not express the RS markers CD15 and CD30 present on most of the cells. This small subpopulation instead resembled memory B cells (CD19+CD20+CD27+) and in addition possessed most of the clonogenic capacity of the HL cell lines; this suggested that RS cells, similar to the neoplastic PC in MM, are not the cells responsible for the growth and maintenance of HL. Using Aldefluor, a fluorescent aldehyde that is a substrate for aldehyde dehydrogenase (ALDH), we previously showed that ALDH is highly expressed in both normal and MM stem cells. The clonogenic subpopulation within the HL cell lines also expressed high ALDH activity, while the predominant RS cells exhibited low ALDH activity. To determine if clonotypic memory B cells were similarly present in HL patients, CD19+ cells were isolated from the marrow or blood of 5 HL patients. The bulk CD19+ cells, as well as those that were ALDHlow, isolated from the HL patients were a mixture of non-clonal naive and memory B cells. However, the ALDHhigh CD19+ cells were a highly enriched population of immunoglobulin (Ig) light chain-restricted CD27+ memory B cells that represented 0.7 to 3% of the circulating CD19+ cells, in 4/5 patients (none with evidence of marrow involvement) studied. These ALDHhigh CD19+ cells also displayed clonal Ig gene rearrangement by polymerase chain reaction (PCR) amplification. In two of these patients, CD15+CD30+ RS cells were isolated from a fresh diagnostic lymph node and contained the same clonal Ig gene rearrangement as the circulating ALDHhigh CD19+ B cells. Thus, clonotypic memory B cells can be found in both HL cell lines and patients. These data suggest that these clonotypic memory B cells circulate in relatively high numbers even in early stage patients, and may represent the HL stem cells. Our results also suggest that therapies, such as rituximab, that target these putative HL stem cells may have clinical activity in HL patients.