Stereotyped patterns of somatic hypermutation in subsets of patients with chronic lymphocytic leukemia: implications for the role of antigen selection in leukemogenesis

Abstract

Somatic hypermutation (SHM) features in a series of 1967 immunoglobulin heavy chain gene (IGH) rearrangements obtained from patients with chronic lymphocytic leukemia (CLL) were examined and compared with IGH sequences from non-CLL B cells available in public databases. SHM analysis was performed for all 1290 CLL sequences in this cohort with less than 100% identity to germ line. At the cohort level, SHM patterns were typical of a canonical SHM process. However, important differences emerged from the analysis of certain subgroups of CLL sequences defined by: (1) IGHV gene usage, (2) presence of stereotyped heavy chain complementarity-determining region 3 (HCDR3) sequences, and (3) mutational load. Recurrent, "stereotyped" amino acid changes occurred across the entire IGHV region in CLL subsets carrying stereotyped HCDR3 sequences, especially those expressing the IGHV3-21 and IGHV4-34 genes. These mutations are underrepresented among non-CLL sequences and thus can be considered as CLL-biased. Furthermore, it was shown that even a low level of mutations may be functionally relevant, given that stereotyped amino acid changes can be found in subsets of minimally mutated cases. IntroductionDeveloping B cells generate a vast repertoire of antibody specificities through somatic recombination of distinct variable (V), diversity (D) (heavy chain only), and joining (J) genes to form the variable domain exons of immunoglobulins (IG). 1 Unlike heavy chain complementarity determining regions (HCDR) 1 and 2, which are entirely encoded by the IGHV gene, HCDR3 is created de novo by the VDJ recombination process. 1 The skewing of diversity to the HCDR3 implies that HCDR3 sequences are the principal determinants of specificity, at least in the primary repertoire. 2,3 However, HCDR3 diversity is not enough to realize the full potential of antibody diversity. 4 Furthermore, unconventional antigens, such as B-cell superantigens, may be recognized not via the CDRs but rather via the framework regions (FRs). 5 Somatic hypermutation (SHM) of IG variable genes forms a second round of diversification after somatic recombination which increases antibody diversity. 6 SHM has long been thought to occur mainly in the germinal centers (GCs) after antigen stimulation and in a manner dependent on T-cell help. 7 Recent reports, however, suggest that SHM can be T-cell independent and may also occur outside classic GCs. [8][9][10][11][12][13] In recent years, the mutational status of IGHV genes has been established as one of the most important molecular genetic markers in defining prognostic subgroups of chronic lymphocytic leukemia (CLL). CLL patients who carry IGHV genes with 98% identity or more to the closest germ line gene ("unmutated") follow a more aggressive clinical course and have strikingly shorter survival than patients carrying IGHV genes with less than 98% identity to germ line ("mutated"). 14,15 The 98% cutoff was chosen as a shortcut to exclude potential polymorphic variants [16][17][18][19] and has been used by the majority of st...

Over 30% of patients with splenic marginal zone lymphoma express the same immunoglobulin heavy variable gene: ontogenetic implications

“…37,38 However, no such bias is evident in other B-cell lymphomas, for example, CLL or MCL, where among the rearrangements using the IGHV1-2 gene, alleles *02 and *04 are detected at roughly similar frequencies. 8,24 This restricted usage of allele *04 in SMZL is noteworthy, given that it differs from allele *02 by a single amino acid, and can be considered as a molecular argument for selection.…”

“…Furthermore, interpretation of results for all cases was performed de novo in the context of this study, following the approaches reported in our recent publications. 8,24,28 Molecular modeling In order to explore the possible modifications in the antibody structure caused by a single amino-acid replacement---namely, the arginine (R)-totryptophan (W) substitution at position VH FR3-75 to obtain an IGHV1-2*04-rearranging antibody---we performed a molecular dynamics study of the human anti-polyhydroxybutyrate antibody F v : IGHV1-2*02 IGKV1-39*01/1D-39*01 (Protein Data Bank code 2D7T). Replacement of the VH FR3-75 R (germline for IGHV1-2*02) by the W residue (germline for IGHV1-2*04) was executed by the COOT algorithm.…”

“…Additionally, many lymphoma subtypes are characterized by somatic hypermutation (SHM) in IGHV genes, suggestive of antigen-driven affinity maturation. 7,8 Previous studies by us and others have shown that SMZL shows a biased IG gene repertoire and is characterized by heterogeneity with regard to the mutational status of the IG receptors. 4,9 --23 Individual studies have highlighted different IG sequence features, including repertoire differences between SMZL, SDRL and hairy cell leukemia-variant; 4,15 intraclonal diversification of IG genes through ongoing SHM; 12,17,18,23 potential analogies between SMZL and hairy cell leukemia-variant; 15 and, very recently, the existence of cases sharing quasi-identical antigen-binding sites (stereotyped B-cell receptors).…”

Immunoglobulin sequence analysis and prognostication in CLL: guidelines from the ERIC review board for reliable interpretation of problematic cases

“…From a biological standpoint, there are data to suggest that low mutational load in CLL productive rearrangements (leading to a germline identity of X98% but o100%) may have been functionally selected, perhaps providing the clone with certain antigenic reactivities. [15][16][17] The unproductive rearrangement would have been similarly targeted by the SHM process but remained free of any functional constraints and thus left to acquire significantly more mutations of a random nature. Whatever the underlying explanation for this phenomenon, at present, a prognostically relevant interpretation is lacking.…”

Next generation RNA‐sequencing in prognostic subsets of chronic lymphocytic leukemia

“…All patients were diagnosed according to recently revised criteria and displayed the typical CLL immunophenotype (CD5 1 /CD19 1 /CD23 1 ) [21]. Subsets were defined according to previously described criteria [3,22] (Supporting Information Table 8S). All samples were collected at follow-up (median 82 and range 6-167 months after diagnosis) and contained 90% (average 93%) tumor cells.…”