Alteration of the p53 and Rb tumor suppressor pathways by p16/INK4a and p14/ARF promoter methylation and loss of protein expression in recurrent and nonrecurrent meningiomas.

2064 Background: Promoter methylation inactivates tumor suppressor genes (TSG). The INK4a/ARF locus encodes p16INKa and p14ARF cell cycle regulatory proteins, which control Rb and p53 TSG pathways. Most meningiomas are slow growing tumors, but in spite of complete surgical removal, the recurrence rate at 5 years is 5%, rising to 19% in long-term follow-up. However, there are no markers predictive of this evolution. Epigenetic changes in low-grade meningiomas have not been previously addressed. To get insights into the possible role of p16INK4a and p14ARF TSG alterations in grade 1 meningiomas, we study the methylation status and protein expression of these genes in 140 specimens of meningiomas: 29 nonrecurrent and 57 recurrent in one, two or three times. Methods: Methylation specific PCR and bisulfate modification followed by bisulfate genomic sequencing of CpG islands and staining with p16INKa and p14ARF antibodies (Ab’s). Results: Our data show p16INK4a and p14ARF methylation in 43.4% and14.2% meningiomas respectively. Methylation of p16INK4a is found in a similar proportion in non-recurrent meningiomas (37.9%) and the first biopsy of recurrent cases (38.8%) and increases to a 52.3% in successive biopsies of recurrent cases. Methylation of p14ARF occurs in 13.8% of nonrecurrent vs. 9.6% recurrent meningiomas (first biopsy) and 19.6% of successive recurrent meningiomas. Loss of p16INK4a and p14ARF protein expression was shown in 52.7% and 18.6% of meningiomas respectively. p16INK4a and p14ARF methylation was associated with loss of protein expression in 54.7% and 18.8% of meningiomas. Loss of p16INK4a and p14ARF expression was associated with unmethylated promoters in 52.9% and 17.6% of cases respectively. Conclusions: Epigenetic changes of p16INK4a and p14ARF genes and loss of protein expression leading to Rb and p53 TSG pathways alterations, may have a pathogenic role in human meningiomas. Loss of p16INK4a and p14ARF protein expression associated with unmethylated promoters, could be due to loss of heterozigosity or gen mutation. Increase of p16INK4a and p14ARF methylation along the following biopsies of recurrent cases suggests a possible role of methylation in tumor progression.