Quantitative real-time PCR does not show selective targeting of p14(ARF) but concomitant inactivation of both p16(INK4A) and p14(ARF) in 105 human primary gliomas

In many human cancers, the INK4A locus is frequently mutated by homozygous deletions, By alternative splicing this locus encodes two non-related tumor suppressor genes, p16(INK4A) and p14(ARF) (p19(ARF) in mice), which regula te cell cycle and cell survival in the retinoblastoma protein (pRb) and p53 pathways, respectively, In mice, the role of p16(INK4A) as th, critical tu mor suppressor gene at the INK4A locus was challenged when it was found tha t p19(ARF) only knock-out mice developed tumors, including gliomas, We have analysed the genetic status of the INK4A locus in 105 primary gliomas usin g both microsatellite mapping (MSM) and quantitative realtime PCR (QRT-PCR) . Comparison of the results of the two methods revealed agreement in 67% of the tumors examined, In discordant cases, fluorescence in situ hybridizati on (FISH) analysis was always found to support QRT-PCR classification, Dire ct assessment of p14(ARF) exon 1 beta, p16(INK4A) exon 1 alpha and exon 2 b y QRT-PCR revealed 43 (41%) homozygous and eight (7%) hemizygous deletions at the INK4A locus. In 49 (47%) gliomas, both alleles were retained, In add ition, QRT-PCR, but not MSM, detected hyperploidy in five (5%) tumors, Dele tion of p14(ARF) was always associated with co-deletion of p16(INK4A) and i ncreased in frequency upon progression from low to high grade gliomas, Shor ter survival was associated with homozygous deletions of INK4A in the subgr oup of glioblastoma patients older than 50 years of age (P = 0.025, Anova t est single factor, alpha =0.05).