Abnormal nuclear and nucleolar immunoreactivity of p16INK4a represents a frameshift alteration in CDKN2A: the E6H4 clone recognizes the p14ARF::p16INK4a chimeric protein

ABSTRACT

In p16INK4a immunostaining, block positivity (i.e., strong nuclear/nuclear-cytoplasmic staining) is a widely acknowledged surrogate marker of HPV-associated cancers. Thus, immunohistochemistry using the anti-p16INK4a clone E6H4 has been approved as an in vitro diagnostic tool for HPV-associated cancers. This study revealed a distinctive pattern of immunostaining with clone E6H4-strong nuclear with nucleolar staining-which can easily be confused with the HPV-associated pattern. This peculiar pattern can be caused by frameshift deletions or insertions within exon 2 of the INK4a/ARF locus. Since p16INK4a and p14ARF are coded in the same sequence on chromosome 9p21, these mutations may inactivate p16INK4a and convert the reading frame for the 3' region of the p14ARF sequence into p16INK4a, producing a chimeric protein. Recognizing this pattern is essential to avoid misdiagnosis of HPV association and inappropriate treatment.