Abstract
Abstract 2425
Mechanisms promoting chronic lymphocytic leukemia (CLL) transformation to Richter syndrome (RS) are poorly understood and might involve antigen stimulation. We explored intraclonal diversification (ID) of immunoglobulin (IGHV) genes in RS in order to: i) follow the evolutionary history of the RS tumor clone over time; ii) understand whether antigen stimulation sustains the growth of DLBCL cells once RS is established. The study was based on 11 clonally related CLL/RS pairs, 7 clonally unrelated RS, and, for comparison, 20 de novo DLBCL. Fifty IGHV subclones were analysed per sample. Mutations observed in only one subclone were defined unconfirmed (UCM). Partially shared mutations were defined confirmed (CM). Cases were scored positive for ID only in the presence of confirmed mutations. For each sample, the normalized mutation frequency (NMF) of ongoing somatic hypermutation was calculated. Phylogenetic analyses was performed with MEGA4. Most (10/11, 90.9%) clonally related RS originated from an ancestor clone that was already present at the time of CLL diagnosis (Fig 1A, 1B, 1C). One single RS case had a transformation pattern compatible with sequential evolution from a secondary CLL subclone (Fig. 1D). All secondary CLL subclones that were documented in the CLL phase disappeared once transformation had occurred, and were substituted by the dominant DLBCL clone with its own descendants. Paired analsysis of clonally related CLL/RS samples documented that NMF significantly decreased during evolution from the CLL phase (median: 0.76 × 10−3) to the RS phase (median: 0.13 × 10−3)(p=.013). Accordingly, at transformation, the ID of IGHV genes switched off in 6/11 (54.5%) clonally related CLL/RS pairs. Clonally related RS that upon transformation maintained ID of IGHV genes were characterized by a higher prevalence of aberrant somatic hypermutation of proto-oncogenes compared to cases lacking ID (0/6 vs 3/4, 75.0%, respectively; p=.033). Also, RS cases that maintained ID of IGHV genes acquired more frequently c-MYC translocation at the time of transformation (0/6 vs 2/4, respectively; p=.133). Clonally unrelated RS are characterized by the development of a de novo DLBCL in the context of CLL. Despite morphologic and phenotypic similarities, clonally unrelated RS differed from de novo DLBCL in terms of NMF (median: 1.18 × 10−3 range: vs median: 0.08 × 10−3 range, respectively; p=.016) and prevalence of cases harboring ID (6/7, 85.7% vs 6/20, 30.0%, respectively; p=.024). The NMF was also significantly higher in clonally unrelated RS compared to clonally related cases (p=.001). These data indicate that: i) RS transformation is due to the expansion of a common ancestor clone that gains selective advantage over other subclones of the CLL phase; ii) antigen stimulation exerts different roles in clonally related RS, clonally unrelated RS and de novo DLBCL; iii) more than 50% clonally related RS switched off ID at the time of transformation, suggesting that the DLBCL clone has become independent of antigen stimulation for its sustainment; iv) clonally related RS that maintained ID of IGHV genes at transformation may take advantage of ID as a mechanism for accumulating genetic instability.
No relevant conflicts of interest to declare.
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