The prognostic significance of various 13q14 deletions in chronic lymphocytic leukemia

Abstract

Purpose: To further our understanding of the biology and prognostic significance of various chromosomal 13q14 deletions in chronic lymphocytic leukemia (CLL).

Experimental design: We analyzed data from SNP 6.0 arrays to define the anatomy of various 13q14 deletions in a cohort of 255 CLL patients and have correlated two subsets of 13q14 deletions (type I exclusive of RB1 and type II inclusive of RB1) with patient survival. Furthermore, we measured the expression of the 13q14-resident microRNAs by quantitative PCR (Q-PCR) in 242 CLL patients and subsequently assessed their prognostic significance. We sequenced all coding exons of RB1 in patients with monoallelic RB1 deletion and have sequenced the 13q14-resident miR locus in all patients.

Results: Large 13q14 (type II) deletions were detected in approximately 20% of all CLL patients and were associated with shortened survival. A strong association between 13q14 type II deletions and elevated genomic complexity, as measured through CLL-FISH or SNP 6.0 array profiling, was identified, suggesting that these lesions may contribute to CLL disease evolution through genomic destabilization. Sequence and copy number analysis of the RB1 gene identified a small CLL subset that is RB1 null. Finally, neither the expression levels of the 13q14-resident microRNAs nor the degree of 13q14 deletion, as measured through SNP 6.0 array-based copy number analysis, had significant prognostic importance.

Conclusions: Our data suggest that the clinical course of CLL is accelerated in patients with large (type II) 13q14 deletions that span the RB1 gene, therefore justifying routine identification of 13q14 subtypes in CLL management.

Figure 1

Genomic copy number heatmap display of chromosome 13q of 255 CLL cases ranked by the position of centromeric 13q14 deletion break points: Copy number heatmap displays for paired DNA samples based on SNP 6.0 array profiling were generated using dChipSNP. Left panel: CD3+ or buccal DNA; Right panel: CLL CD19+ DNA. Blue indicates copy loss, red indicates copy gain. Each column represents one patient.

Figure 2A-D

Normalized expression of miR15a and miR16-1 versus SNP 6.0 array-based copy number (CN) estimates for typical 13q14 deletions: Panels A-D: Displayed are mean delta Ct values (miR15a or 16-1 minus housekeeping microRNAs) as single dots (y-axis) versus SNP 6.0 profiling-based CN estimates for 13q14 deletions (x-axis) grouped by 13q14 deletion status. Red numbers indicate mean delta Ct values for CLL groups with CN<1 or ≥1 and 13q14 deletions or non-del13q14 status. Black dots: CLL cases with unresolved miR gene status.

Figure 3A-H

Deletions 13q14 types I or II and OS in CLL (Kaplan-Meier plots): Panels A-D: Deletion 13q14 types I or II and OS in CLL. UT: untreated at enrollment; UT+T: untreated or relapsed at enrollment. A,B: OS from date of enrollment, C,D: OS from date of diagnosis; Panels E-H: pairwise groupings by 13q14 status (I or II) and any associated FISH lesions. The suffix “complex” indicates a 13q14 deletion with any (≥1) coexisting FISH-25 finding.

Figure 4A-H

Deletion 13q14 types I or II and associated SNP 6.0 array profiling-based aCNA and OS in CLL (Kaplan-Meier plots): Panels A-D: OS from date of enrollment. Panels E-H: OS from date of diagnosis. UT: untreated at enrollment; UT+T: untreated or relapsed at enrollment. The suffix “sc” indicates sub-chromosomal aCNA present at indicated thresholds.

Figure 5A-F

Identification of somatically acquired RB1 mutations and results of Q-PCR-based RB1 expression analysis: A: Delta Ct values (Ctm RB1 minus Ctm GAPDH) with each dot representing the mean of duplicate measurements in individual patients (N=160). Groupings are by 13q14 status. B: RB1 mutation results in CD19+-derived DNA versus CD3+- derived DNA.

Figure 6A-B

Identification of somatically acquired miR16.1 mutations: miR16.1 mutation results in CD19+-derived DNA versus CD3+- derived DNA. The nucleotide sequence of the miR16.1 gene located within 13q14 is indicated and miR16.1-3p and miR16.1-5p are highlighted in yellow.