ORIGINAL ARTICLE
Efficiency of immunohistochemical p16 expression and HPV typing in cervical squamous intraepithelial lesion grading and review of the p16 literature

https://doi.org/10.1016/j.prp.2007.03.010Get rights and content

Abstract

Diagnosing and grading cervical cancer precursors is challenging. This study investigates the presence of HPV infection, the expression of p16, and any correlation between these two findings.

H&E-stained slides of cervical loop excision materials diagnosed as LSIL and HSIL were reviewed. An immunohistochemical panel consisting of p16 as well as of all HPV types and HR-HPV types was applied. Staining of p16 was evaluated according to distribution extent and degree of intensity.

All HSIL cases and 80% of LSIL cases were positive for p16. In HSIL cases, the staining distribution was as follows: 50% full thickness, 45% basal, and 5% rare. The staining intensity for the same cases was strong in 70%, variable in 20%, and weak in 10% accordingly. In LSIL cases, staining distribution was basal in 58.3% and rare in 41.7%. None of the LSIL cases showed full thickness of p16 positivity. The staining intensity of the same cases was strong in 25%, variable in 16.7%, and weak in 58.3%. Of all cases, 48.6% were positive for screening kit (all HPV types), and 31.4% of all cases were positive for HR-HPV. The distribution of this positivity was 35% for HSIL and 26.6% for LSIL cases. The total HPV-type positivity rate was 48.6%, the distribution being 50% for HSIL and 46.6% for LSIL cases.

p16 is a highly sensitive marker for cervical epithelial dysplasia. Strong and full thickness staining of p16 in the cervix epithelium is highly supportive of HSIL, while weak and basal/rare staining favors LSIL. All HPV-positive cases were also p16-positive, but no statistically significant relationship between HPV infection positivity and the intensity and distribution of p16 was found. HPV is not helpful in the grading of SIL, as an unignorable rate of HR-HPV positivity (26.6%) was detected in LSIL group.

Introduction

Occasionally, it is challenging to render an accurate histopathologic diagnosis of cervical cancer precursors, which is an essential determiner in prognosis and survival. In particular, the differential diagnosis between reactive states and LSIL, and LSIL and HSIL may be difficult [6], [7], [9], [20]. At this point, the need for objective and reliable diagnostic tools cannot be ignored.

p16, a tumor suppressor protein and also a cyclin-dependent kinase inhibitor, whose overexpression has repeatedly been reported to be typical of dysplastic and neoplastic epithelium of cervix [29], [1], [15], slows down the cell cycle by inactivating the cyclin-dependent kinases that phosphorylate retinoblastoma protein (pRb) [26], [16]. The viral oncogenes E6 and E7 of HPV, whose expression is associated with malignant transformation of the cervical epithelial cells [27], [31], have the ability to bind and inactivate pRb. The pRb status has a remarkable influence on the expression of p16. p16 expression in cervical lesions is hypothesized to be caused by the functional inactivation of pRb by HPV-E6 and E7 proteins.

Many studies have proposed that p16 is a useful biomarker especially for HR (high risk)-HPV type-related cervical neoplasia [16, 18, 33, 35, 39, 45, 56, 58], and also for predicting SIL progression [13], [22], [10]. Moreover, a few other studies have recently concluded that there exists a significant association between cervical lesion grade and p16 staining distribution and intensity [17], [12], [11]. As for the localization of p16 staining, there are studies that report predominant cytoplasmic staining [15] as well as predominant nuclear staining [30], and a combination of both [23].

In this study, we used immunohistochemistry to investigate the staining pattern of p16 on the basis of distribution and intensity, and the presence of HPV in squamous intraepithelial lesions of the cervix. In this way, it was expected to define the role of p16 expression and HPV typing in the diagnosis and grading of the precursor lesions of the cervix.

Section snippets

Materials and methods

Hematoxylin-eosin stained slides from formalin-fixed, paraffin-embedded tissue sections from cervical loop excision materials, formerly diagnosed as LSIL and HSIL, were reviewed by the same pathologist. From the group of cases with confirmed diagnosis, we selected a LSIL group of 15 cases and a HSIL group of 20 cases. From each chosen paraffin-embedded tissue block, three serial sections were taken.

Results

All HSIL cases (20/20) and 80% of LSIL cases (12/15) were positive with p16. Staining distribution in the HSIL cases was as follows: 50% full thickness, 45% basal, and 5% rare (Fig. 1k, l). The staining intensity of the same cases was strong in 70%, variable in 20%, and weak in 10%, accordingly. Of p16-positive LSIL cases, staining distribution was basal in 58.3% and rare in 41.7%. None of the LSIL cases showed full thickness of p16 positivity (Fig. 1m, n). The staining intensity of the same

Discussion

Despite extensive studies of cervical cancer precursors, the interobserver variation in the histopathologic interpretation of cervical biopsy specimens still constitutes a dilemma [14], [28]. The search for a specific diagnostic biomarker to solve the problem of lack of interobserver reproducibility in the histologic diagnosis of cervical intraepithelial lesions revealed p16, a tumor supressor protein, whose overexpression has been frequently shown in cervical cancers and its precursors: Klaes

Acknowledgements

The authors would like to thank to Ünal Şeref for technical support and to Lokman Kale for archival assistance.

References (31)

  • F.X. Bosch et al.

    The casual relation between human papillomavirus and cervical cancer

    J. Clin. Pathol.

    (2002)
  • F.X. Bosch et al.

    Prevalence of human papillomavirus in cervical cancer: a worldwide perspective

    J. Natl. Cancer Inst.

    (1995)
  • G.M. Clifford et al.

    Human papillomavirus types in invasive cervical cancer worldwide: a meta-analysis

    Br. J. Cancer

    (2003)
  • R.T. Grenko et al.

    Variance in the interpretation of cervical biopsy specimens obtained for atypical squamous cells of undetermined significance

    Am. J. Clin. Pathol.

    (2000)
  • M.C. Guimaraes et al.

    Immunohistochemical expression of p16INK4a and bcl-2according to HPV type and to the progression of cervical squamous intraepithelial lesions

    J. Histochem. Cytochem.

    (2005)
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