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CCND1 (cyclin D1) at 11q13.3 encodes an important cell cycle regulator, cyclin D1 that modulates the cell cycle G1/S transition. Amplification and overexpression of this gene are frequently found in various human cancers. By gene-based array comparative genomic hybridization (array CGH) (Vysis) analysis, we have revealed that copy number gains of CCND1 in 1/2 NPC cell lines (HK1) and 2/3 xenografts (Xeno-2117 and Xeno-NPC8). The copy number gains of CCND1 in the 3 NPC samples were further been confirmed by FISH analysis. Amplifications were detected in the 2 xenografts, Xeno-2117 and Xeno-NPC8. By quantitative RT-PCR, consistent CCND1 overexpression was found in the 3 NPC samples with copy number gains. Xeno-2117 with the highest level of CCND1 amplification (3.43-fold increase) showed 4.47-fold increase in CCND1 mRNA expression while compared with the normal nasopharyngeal cells. Overexpression of cyclin D1 in these samples was also confirmed by Western blot analysis. The copy number aberration and protein expression of CCND1 were also examined on 21 primary NPCs by FISH and immunohistochemical analysis. Among the 21 primary tumors, gains of CCND1 were detected in 16 cases (76%), while only 2 cases (9%) showed high-level amplification. Overexpression of CCND1 was also found in 12 out of 21 (57%) primary tumors. A correlation of copy number changes with overexpression of CCND1 in the NPC samples (P<0.005, Fisher’s Exact Test) was shown. The findings imply that CCND1 is a target oncogene at 11q13 for NPC. To further elucidate the role of CCND1 activation in NPC cells, siRNA targeting CCND1 was then transfected into a NPC cell line HK1. Quantitative RT-PCR and Western blot analysis showed knockdown of more than 90% protein expression after transfection. Cell proliferation analysis showed more than 70% decrease in cell growth of siRNA-treated HK1 cell line. By flow cytometry analysis, S to G2 phase arrest in the cell cycle was found in the siRNA-treated cells. The findings suggested that activation of CCND1 play an important role in NPC tumorigenesis.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]