Evaluation of cervical cone biopsies for coexpression of p16INK4a and Ki-67 in epithelial cells.

Diffuse overexpression of p16(INK4a) in basal and parabasal cells of cervical epithelium is a hallmark of human papillomavirus-mediated transformation. Focal p16(INK4a) expression is occasionally observed in nondysplastic epithelium. In normal cells, expression of p16(INK4a) triggers cell cycle arrest. However, cells undergoing transformation in intraepithelial lesions actively proliferate. To prove that the different expression patterns of p16(INK4a) , i.e., focal versus diffuse, reflect biologically different entities, we hypothesized that p16(INK4a) -positive cells in epithelia displaying focal p16(INK4a) expression pattern do not coexpress proliferation-associated Ki-67 protein, while p16(INK4a) -positive cells in lesions with diffuse p16(INK4a) expression may do. A total of 138 cervical cone biopsies were stained for the expression of p16(INK4a) and Ki-67 using a primary antibody cocktail. All metaplastic lesions (n = 21) displayed focal staining for p16(INK4a) , and in all of these lesions p16(INK4a) -positive cells were found to be negative for Ki-67 expression. Diffuse expression of p16(INK4a) was observed in 12/21 (57.1%) cervical intraepithelial neoplasia (CIN) 1 lesions, all of them simultaneously showed Ki-67 immunoreactivity in a large proportion of p16(INK4a) -positive cells. Seventeen of 23 (73.9%) CIN2 lesions and all 27 (100%) CIN3/carcinoma in situ (CIS) as well as all 46 (100%) carcinoma cases displayed diffuse and combined expression of p16(INK4a) and Ki-67. Coexpression of Ki-67 and p16(INK4a) in the same cell is entirely restricted to cervical lesions displaying diffuse p16(INK4a) expression, whereas in lesions with focal p16(INK4a) expression, p16(INK4a) -expressing cells are negative for Ki-67. Thus, diffuse expression of p16(INK4a) reflects lesions with proliferation-competent cells, while p16(INK4a) -expressing cells associated with focal expression patterns are cell cycle arrested.

Novel concepts in cervical cancer screening: a comparison of VIA, HPV DNA test and p16INK4a/Ki-67 dual stain cytology in Western Kenya.

BACKGROUND: Screening of unvaccinated women remains essential to mitigate the high morbidity/mortality of cervical cancer. Here, we compared visual inspection with acetic acid (VIA), recommended by WHO as the most cost-effective screening approach in LMICs, with HPV-based screening, and usage of p16INK4a/Ki-67 dual stain cytology. METHODS: We prospectively enrolled women participating in a VIA-based cervical cancer screening program in two peri-urban health centers of Kenya. Consenting women had a VIA examination preceded by collection of a liquid-based cytology sample from the cervix stored in PreservCyt medium (Hologic®). Analysis of all samples included a hrHPV DNA test and evaluation of a p16INK4a /Ki-67 (CINtecPLUS®) dual stained slide that was prepared using the ThinPrep® 2000 Processor and evaluated by a pathologist trained in the methodology. RESULTS: In 701 of a total of 800 women aged 18-64 years, all three investigations were performed and data could be analyzed. The HPV, VIA and dual stain cytology positivity were 33%, 7%, and 2% respectively. The HPV positivity rate of VIA positive cases was 32%. The five most common HPV types were HPV16, 52, 68, 58 and 35. The OR among HIV infected women of an HPV infection, VIA positivity and positive dual stain cytology were 2.6 (95%CI 1.5-4.3), 1.9 (95%CI 0.89-4.4) and 3.4 (95%CI 1.07-10.9) respectively. The sensitivity of VIA to detect a p16INK4a/Ki-67 positive transforming infection was 13% (95%CI 2-38). CONCLUSIONS: Primary HPV testing appears feasible and should be considered as a primary screening test also in LMICs. The poor sensitivity of VIA renders it unsuitable as a triage test for HPV positive women. The utility of p16INK4a/Ki-67 dual stain cytology as a triage test for HPV positive women in LMICs should be further studied.

Home-based HPV self-sampling assisted by a cloud-based electronic data system: Lessons learnt from a pilot community cervical cancer screening campaign in rural Ethiopia.

Primary HPV testing and triage of HPV-positive women is an effective cervical cancer screening strategy. Such a multi-visit screening algorithm is also promising for community-based screening in resource-poor communities, provided a robust tracking system is in place. A cervical cancer screening campaign was conducted in a rural community in Ethiopia. All women aged 25-65 years were offered genital self-sampling using the Evalyn Brush®. Samples were HPV-DNA-tested at a central laboratory. Key indicators were captured on tablet computers and linked by a cloud-based information system. HPV-positive women were examined at the local clinic using portable colposcopy, p16/Ki-67 dual stain cytology and biopsy examination. CIN2+ women were referred for LEEP to the referral hospital. Of 749 enumerated age-eligible women 634 (85%, (95% CI 82-88)) consented to screening, 429 samples were adequate for HPV testing, giving a total testing coverage of 57% (95% CI 53-62). The hrHPV prevalence was 14% (95% CI 5-22), 72% (95% CI 60-84) attended the clinic for a triage examination. Home-based HPV-DNA self-sampling and clinic-based triage assisted by cloud-based information technology is feasible in rural Ethiopia. Key components of such strategy are broad community awareness, high competency of community workers, and establishment of an adequate self-sampling and HPV-DNA testing platform.

Clonal history of papillomavirus-induced dysplasia in the female lower genital tract.

BACKGROUND: Dysplastic lesions of the vagina or the vulva often occur in women who have a previous history of cervical dysplasia. Most lesions in the female lower genital tract are induced by infections with high-risk oncogenic human papillomaviruses (HR-HPVs), including HPV16 and HPV18. HR-HPV genomes frequently integrate into host cell chromosomes at random sites. We analyzed viral integration sites in multiple metachronous lesions of the lower genital tract from women previously treated for HR-HPV-positive cervical dysplasia or cancer to determine whether the metachronous lesions emerged from a single common preexisting dysplastic cell clone or as consequence of independent HR-HPV infection events in the female lower genital tract. METHODS: From among 1500 patients with anogenital lesions, seven patients with high-grade vaginal or vulvar lesions and with a previous history of cervical disease (five with prior high-grade cervical dysplasia and two with a history of cervical cancer) were included in this study. Integration sites of HPV16 or HPV18 in vaginal or vulvar lesions were mapped by an adaptor ligation polymerase chain reaction (PCR) method. The sequence information was used to design an integrate-specific PCR assay that was applied to DNA extracted from archival paraffin-embedded material derived from biopsy samples of cervical lesions. RESULTS: Identical HPV DNA integration loci were found in vaginal or vulvar and cervical samples of all lesions available for four of the five patients with a prior history of high-grade cervical dysplasia and for both patients with a history of cervical cancer. CONCLUSIONS: These data indicate that high-grade dysplastic lesions in the female lower genital tract may emerge primarily as monoclonal lesions from a transformed cell population derived from the uterine cervix.