Analysis of Gene Expression Patterns and Gene Copy Number Changes in Human NK Cell Malignancies.

Natural killer (NK)-cell lymphomas/leukemias are a group of aggressive malignancies distinct from the indolent large granular lymphocyte (LGL) proliferations. In the World Health Organization (WHO) classification, NK-cell malignancies are divided into two groups (1) extranodal NK-cell lymphoma, nasal type (ENKL), and (2) aggressive NK-cell leukemia (ANKL). Only three studies of genome wide alteration have been published in these malignancies. We studied eight NK lymphoma cell lines (NK-92, KHYG1, YT, HANK-1, NK-YS, SNK-6, SNK-3 and KAI3) and seven NK lymphomas/leukemias specimens to:

1. determine DNA gains and losses in comparative genomic hybridizations (CGH) and compare these alterations with those reported literature to define recurring chromosomal abnormalities,

2. obtain transcription profiles of NK malignancies and compare them with normal NK cells(resting or IL-2 stimulated), and

3. correlate the genomic profiles with transcriptional profiles to identify possible candidate genes involved in lymphoma development.

Whole genome tiling arrays, containing 26,819 bacterial artificial chromosomes (BAC) were employed for CGH study.

Our CGH study identified frequent recurrent abnormalities that have been reported in all previous studies. These include gains in 17q11.2–q21 (frequency: 40%); 1q22–q23.1 (frequency: 50 %); and 7q21.13–q22.1(frequency: 50%) and all these regions showed at least a frequency of 17% in other reports. A number of regions were also described in at least one other published reports (e.g. 1q32.2–q41, 6p25.5 and 17q 21.2). Regions of loss with a frequency ≥ 20% including 11q23.1–23.2(20%); 7p22.1–p22.2 (40%); 6q21(50%); 6q25.1–25.3(50%); and 17p13–13.1(50%) showed consistent results with previous reports. 6q21 loss was detected in two ANKL cell lines (NK92 and KHYG1) but not in ENKL cell lines . Gains of 8p12–q21.12, 8q22.1–q24.13, and 8q24.21 were also unique to ANKL cell lines.

Transcriptional profiling using affymetrix gene chips on these cell lines and tumor specimen were compared with normal resting and IL-2 activated NK cells. We observed significant down-regulation (6–9 folds) of killer cell Ig-like receptor (KIR) (KIR2DL3, KIR2DS1, KIR3DL1, KIR3DL2) in NK-cell lymphoma as compared to normal NK cells. This may be related to the restricted clonal expression of the KIRs in neoplasms. High expression of pro-proliferative genes (e.g. Cyclin B1, A2, BUB1, CDC2, CDC20, CDCA1, CDCA3, and CDC7) was observed in NK lymphoma. On examining gene signatures/pathways (using BRB-array Tools), we observed significant increased expression of genes (p<0.001) involved in oxidative-phosphorylation /electron transport chain and MYC target signature in NK-cell lymphoma. A number of genes involved in TGF-beta signaling (p<0.0001) were down-regulated in lymphoma samples as compared to normal cells. This may correlate with the deletion of two crucial regulators (TGFBR3 and SMAD7) of this pathway, located at 1p32–33 and 18q21.1 regions respectively. In summary, combination of the genomic and transcriptional profiling by microarrays may lead to the identification of candidate genes in genomic alterations and provide clues to the pathogenesis of NK-cell malignancy.