Conserved Amino Acid Sequences Occur in a Limited Group of CLL Patients with Predominantly Mutated IGH Rearrangements.
Authors: Duke Veronique M. , Yogashangari Birithini , Gandini Domenica , Sherrington Paul D. , Mehta Atul , Hoffbrand A. Victor , Foroni Letizia 
Published: October 2019

Assessment of IGH mutation rate combined with other clinical and immunological parameters has become an accepted prognostic indicator in the management of CLL patients, mutated-CLL patients showing a better overall outcome than patients with VH germline genes. Through the availability of a large cohort of sequenced CLL cases two features of interest have emerged. Firstly, we and others observed a biased usage of individual VH genes in both the mutated and germline groups (Duke et al, 2003; Hematologica; 88: 1259). Secondly, some investigators have suggested the existence of a “common” antigen stimulation as an underlying cause of the observed B cell expansion in a proportion of cases (Messmer et al, 2004; Blood; 103:3490).

Aims. The aims of this study were twofold: 1) to investigate the level of aminoacid homology of the IgH CDR3 region in 179 CLL patients; 2) to investigate a possible correlation with stage and other clinical parameters known to be significant in assessing CLL patients.

Methods. FR1-JH amplified sequences were analysed using the MEGALIGN program which allow protein translation and DNA and aminoacid (AA) sequence alignments for all framework(FR) and complementarity determining regions (CDR). Only VH genes with four or more patients were included in the following analysis.

Results. Of 179 patients, 147 (82%) fulfilled the criteria set for analysis, i.e. VH genes used in ≥4 patients. Informative results are summarised in Table 1. Overall, ten patients of 147 suitable for analysis (6.8%) were found to have homologous CDR3 regions.

Results of CDR3 AA homology analysis

VH gene Patients (no) Homologouse seq (no) % VH 1–69 25 2 sets (5 patients) 20 VH 3–21 10 3 30 VH 3–23 16 2 12.5 VH 4–34 11 50% homology in 2 patients 18 TOTAL 62 10 16

None of the remaining 85 cases showed AA homology, particularly in the CDR3 region among rearrangements involving VH1-2 (14 cases), VH1-3 (5 cases) VH2-5 (8 cases), VH3-7 (9 cases), VH3-15 (6 cases), VH3-30 (14 cases) VH3-48 (12 cases) VH4-39, VH4-59, VH4-61 (4 cases each) and VH5 (5 cases).

Of the 10 patients that had homologues sequences eight (80%) had been classified as CLL-mutated VH gene, while only two (20%) were in the germline group. There was a predominance of disease Stage A (6 stage A, 1 stage B; for cases with available information) and most were male (n=7) females(n=2). The median age was 67 years (56–73years) the median WBC was 43.05 (16.7–69.4).

In conclusion, between 12.5 and 30% of patients with individual VH gene rearrangements of frequently used IGH genes (VH1-69, VH3-21, VH3-23 and VH4-34) show homology within their CDR3 region. Although in some instances increased mutation rate may be associated with poorer outcome (as for the BCL6 gene) in CLL patients AA homology appears to be associated with better outcome even in patients with rearrangement of the VH1-69 gene, which when in its germline is associated with poor outcome. Whether this is an antigen or T cell mediated event, remains to be investigated as more information will become available in the future.