Fine-mapping loss of gene architecture at the CDKN2B (p15INK4b), CDKN2A (p14ARF, p16INK4a), and MTAP genes in head and neck squamous cell carcinoma.
OBJECTIVE
To identify the extent and the smallest region of loss for CDKN2B(INK4b), CDKN2A(ARF,INK4a), and MTAP. Homozygous deletions of human chromosome 9p21 occur frequently in malignant cell lines and are common in squamous cell carcinoma of the head and neck (HNSCC). This complex region encodes the tumor suppressor genes cyclin-dependent kinase 2B (CDKN2B) (p15(INK4b)) and CDKN2A (p14(ARF), p16(INK4a)) and the housekeeping gene methylthioadenosine phosphorylase (MTAP).


METHODS
A targeted probe panel designed to finely map the region of 9p21 loss comprised 3 probes for CDKN2B(INK4b), 7 for CDKN2A(ARF, INK4a), and 3 for MTAP and was interrogated using the multiplex ligation-dependent probe amplification assay (MLPA). The MLPA genomic copy number alterations for CDKN2A were validated using real-time polymerase chain reaction.


METHODS
Six HNSCC primary (A) and recurrent or metastatic (B) cell lines were examined: UMSCC-11A/11B, UMSCC-17A/17B, and UMSCC-81A/81B.


RESULTS
Cell line UMSCC-11B retained all 9p loci tested in the region. Cell lines UMSCC-17A/B indicated homozygous deletion of CDKN2A(ARF, INK4a) starting at p16(INK4) exon 1alpha to include exons 2 and 3. Homozygous loss was indicated for CDKN2B(INK4b) and CDKN2A(ARF,INK4a) in UMSCC-11A, and UMSCC-81A. Cell line UMSCC-81B indicated retention of all 9p loci except for exon 1alpha (p16(INK4a)). Selective loss of the 3' end of MTAP was observed in UMSCC-11A. Genomic alterations by fine-mapping MLPA were validated at the DNA level for CDKN2A.


CONCLUSIONS
We identified exon 1alpha (p16(INK4a)) as the smallest region of loss in the CDKN2A(ARF, INK4a) gene. The frequency and precise loss of CDKN2B(INK4b), CDKN2A(ARF, INK4a), and MTAP in the prognosis of 9p21-deleted HNSCC may provide impetus for use of these targets as therapeutic biomarkers in head and neck cancer.
Worsham
Maria J
MJ
Department of Otolaryngology-Head and Neck Surgery and Research Division, Henry Ford Hospital, Detroit, MI 48202, USA. mworsha1@hfhs.org
Chen
Kang Mei
KM
Tiwari
Nivedita
N
Pals
Gerard
G
Schouten
Jan P
JP
Sethi
Seema
S
Benninger
Michael S
MS
D002294:Carcinoma, Squamous Cell / Q000235:genetics*; D045744:Cell Line, Tumor; D002874:Chromosome Mapping; D002899:Chromosomes, Human, Pair 9 / Q000235:genetics*; D050762:Cyclin-Dependent Kinase Inhibitor p15 / Q000235:genetics; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics; D004273:DNA, Neoplasm / Q000032:analysis; D017353:Gene Deletion; D020868:Gene Silencing; D016147:Genes, Tumor Suppressor; D019942:Genes, p16; D006258:Head and Neck Neoplasms / Q000235:genetics*; D006801:Humans; D011683:Purine-Nucleoside Phosphorylase / Q000235:genetics*; D025803:Tumor Suppressor Protein p14ARF / Q000235:genetics
C493966:CDKN2B protein, human; D050762:Cyclin-Dependent Kinase Inhibitor p15; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004273:DNA, Neoplasm; D025803:Tumor Suppressor Protein p14ARF; D011683:Purine-Nucleoside Phosphorylase; C021784:5'-methylthioadenosine phosphorylase