SNP-Array Profiling Identifies Complex Aberrations and Candidate Genes in Myeloproliferative Neoplasms with Leukemic Transformation.

Myeloproliferative neoplasms (MPN) represent a heterogeneous group of acquired hematopoietic stem cell disorders. Clonality leads to exceeding production of myeloid cells resulting in an inherent tendency for thrombotic and hemorrhagic complications as well as transformation into acute myeloid leukemia (sAML). While vascular complications predominantly account for the morbidity in essential thrombocythemia (ET) and polycythemia vera (PV), the mortality of MPN is significantly related to leukemic transformation. Secondary AML occurs more frequently in primary and secondary myelofibrosis (PMF and SMF) than in ET and PV, and the risk for leukemic transformation increases with the duration of the disease. The molecular basis underlying the progression of MPN is poorly understood. Clonal evolution due to genomic instability is considered to play an important role.

To identify genomic lesions associated with leukemic transformation, we applied 250K single-nucleotide polymorphism (SNP) arrays that allow for genome-wide screening of both copy-number alterations (CNAs) and copy-neutral runs of homozygosity (ROH) at high resolution.

An unpaired SNP-array analysis of 23 sAML samples was performed [former diagnosis: ET, n=5; PV, n=7; PMF, n=9; SMF, n=2;]. An own set of 30 reference samples was used for normalization. CNAs and ROH were analyzed by CNAG 3.0 software. Aberrations were compared with the 250K SNP-array dataset of 151 MPN patients [ET, n=45; PV, n=45; PMF, n=47, SMF, n=14]. In one sAML patient corresponding SNP-array data from the time of ET diagnosis were available.

CNAs were present in 15 of 23 (65%) sAML patients. Thirty-five percent of cases (n=8) exhibited complex genomic aberrations with up to 20 CNAs in one patient (range 5–20). The most frequent larger (>5 Mb) CNAs were trisomy 8 (n=7), gain of 1q, loss of 5q, and deletion in 6p25-pter (16.2–26.7 Mb) and 20q11-q13 (13.6–16.9 Mb) in three cases each, followed by gain in 3q24-qter (51.7 and 54.1 Mb) in two patients. Of note, one case with deletion in 17p12-pter (64.4 Mb) encompassing TP53 and a second with gain in 21q22.12-qter (11.8 Mb) were identified; in the latter one the proximal breakpoint of the gain was located at RUNX1. Smaller CNAs (<5 Mb) were restricted to single cases with four cases exhibiting micro-deletions ranging from 0.7 to 2.7 Mb in size. Interestingly, three chromosomal regions harbour single genes: 11p11.2 (FOLH1), 18q21.2 (TCF4), and 21q22.12 (RUNX1). ROH comprising the terminal end of the chromosome were detectable in 13 of 23 (57%) sAML cases. The most frequent ROH included the JAK2 locus in 9p24 (n=6; 15.6–38.7 Mb), followed by ROH in 17p13-pter (16.3 and 17.7 Mb) covering TP53 and an overlapping segment in 1p32-pter (53 Mb) affecting MPL in two cases each. All cases with 9p ROH were JAK2 V617F mutated, whereas the MPL W515L mutation was found in one of the two 1p ROH cases. Moreover, sequencing analyses in both patients with ROH in 17p revealed TP53 missense mutations in exon 7 and exon 8, respectively. In addition, non-recurrent ROH covering the long arm of chromosome 7, 11, and 21 as well as ROH in the chromosomal segments 14q32-qter (12.6 Mb) and 17q31-qter (31 Mb) were identified. In one sAML patient SNP-array data performed at the time of ET were available for comparative analysis. While 20q deletion was present as sole aberration in ET, complex genomic aberrations (7 CNAs) were identified after development of sAML. Merging the results from our recent 250K SNP-array analysis (Stegelmann et al., Blood 2008; 112: Abstract #2794) on 61 PMF and SMF cases with data from this study, we were able to identify a second case with micro-deletion in 12q24. The commonly deleted region of both cases is 1.3 Mb in size and encompasses TCF1 as a novel recurrent aberration in MPN.

In summary, our data on a large series of well-defined sAML cases that evolved from MPN demonstrate that 250K SNP-array profiling is an excellent tool to identify genomic aberrations. In contrast to MPN, genomic alterations in sAML are characterized by a marked complexity reflecting both genomic instability and genetic heterogeneity of sAML. However, in our study several regions of interest including recurrently affected candidate genes such as TCF1, RUNX1, and TP53 were identified that need to be further investigated on a single gene level.

No relevant conflicts of interest to declare.