Prospective evaluation of p16/Ki-67 dual-stained cytology for managing women with abnormal Papanicolaou cytology: PALMS study results

Cancer Cytopathol. 2015 Jun;123(6):373-81. doi: 10.1002/cncy.21542. Epub 2015 Apr 17.


Background: Testing for the presence of the human papillomavirus (HPV) is widely accepted for triaging Papanicolaou cytology results categorized as atypical squamous cells of undetermined significance (ASC-US). In contrast, HPV testing has limited use in triaging cytological low-grade squamous intraepithelial lesions (LSILs) due to prevalence rates of typically >80%. In the current study, the authors assessed the diagnostic performance of p16/Ki-67 dual-stained cytology in triaging ASC-US and LSIL cases within the prospective, multicentric Primary ASC-US LSIL Marker Study (PALMS).

Methods: A total of 575 ASC-US cases and 529 LSIL cases from a cohort of 27,349 women who were prospectively enrolled into the PALMS study in 5 European countries were tested with p16/Ki-67 dual-stained cytology and Hybrid Capture 2 (HC2) HPV testing. Colposcopy-guided biopsy results of cervical intraepithelial neoplasia of grade 2 or worse (CIN2+) were used as clinical endpoints.

Results: p16/Ki-67 dual-stained cytology demonstrated comparable (ASC-US: 94.4% for dual-stained cytology vs 100% for HC2 testing; P = .317) or lower (LSIL: 85.7% for dual-stained cytology vs 98.4% for HC2 testing; P = .005) sensitivity for CIN2+, but higher levels of specificity compared with HC2 HPV testing in both ASC-US (78.7% vs 60.4%; P<.001) and LSIL (53.3% vs 15.6%; P<.001) cases. Positive predictive values for CIN2+ were substantially higher for dual-stained cytology versus HC2 HPV testing, especially in LSIL, and in ASC-US cases for women aged <30 years.

Conclusions: The clinical usefulness and efficiency of triaging women with ASC-US or LSIL Papanicolaou cytology results by p16/Ki-67 dual-stained cytology testing has been confirmed in this prospective, pan-European study. The high positive predictive value of dual-stained cytology for the presence of high-grade CIN may help to reduce the number of unnecessary colposcopy referrals.

Keywords: CINtec PLUS; Primary ASC-US LSIL Marker Study (PALMS); atypical squamous cells of undetermined significance (ASC-US); cervical cytology; human papillomavirus (HPV); low-grade squamous intraepithelial lesion (LSIL); p16/Ki-67 dual-stained cytology; triage.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adolescent
  • Adult
  • Aged
  • Biomarkers, Tumor / analysis
  • Cervical Intraepithelial Neoplasia / diagnosis*
  • Cervical Intraepithelial Neoplasia / metabolism
  • Cyclin-Dependent Kinase Inhibitor p16 / metabolism*
  • Cytodiagnosis
  • Female
  • Follow-Up Studies
  • Humans
  • Ki-67 Antigen / metabolism*
  • Middle Aged
  • Neoplasm Grading
  • Prognosis
  • Prospective Studies
  • ROC Curve
  • Squamous Intraepithelial Lesions of the Cervix / diagnosis*
  • Squamous Intraepithelial Lesions of the Cervix / metabolism
  • Uterine Cervical Neoplasms / diagnosis*
  • Uterine Cervical Neoplasms / metabolism
  • Vaginal Smears*
  • Young Adult


  • Biomarkers, Tumor
  • Cyclin-Dependent Kinase Inhibitor p16
  • Ki-67 Antigen