Factors associated with cervical intraepithelial neoplasia among women in Kisumu County and the diagnostic performance of p16/ki-67 Biomarkers

Abstract

Cervical intraepithelial neoplasia (CIN) is a premalignant lesion of the cervix that can progress to cervical cancer (CC) if not identified early and treated as appropriate. Globally, incidence of CC is estimated at 660,000 annually with the greatest burden found in sub-Saharan Africa (SSA) and about 40.1 per100,000 reported in Kenya of which 9 per 30,000 screened are admitted in Kisumu County with advanced stage cancer annually. Besides HPV infection, the definitive factors promoting CC burden in SSA are still not clear; although imbalance in cervicovaginal microbiome especially HSV-2 and CT have been suggested in studies conducted in Africa, differently from those conducted in other regions. Moreover, controversy surrounding hormonal contraceptives use and incidence of CIN in some regions remained unresolved. This study examined the effects of coinfections and contraceptives use on incidence of CIN in the region. Currently, HPV DNA test is the primary tool for CC screening with VIA serving as a triage test where Pap cytology is not available. However, Pap cytology is more subjective and inaccessible to majority patients while VIA is limited by low sensitivity. A more efficient point of care test (POCT) that guarantees a one-day patient visit complete with test results and treatment is still lacking. Integration of p16/ki-67 biomarkers in the current CC screening program, specifically to identify cases of cervical dysplasia with subsequent treatment in a single visit is currently explored in a number of countries. This study compared the diagnostic performance of p16/ki-67 biomarkers with VIA using histology as a standard. In a hospital based cross-sectional study, the risk factors associated with CIN and the diagnostic performance of p16/ki-67 cytology among women in Kisumu County were assessed. The specific objectives were to determine the associations between coinfections and hormonal contraceptive use with incidence of CIN; and compared the diagnostic performance of p16/ki-67 cytology with that of VIA. A total of 517 referrals with cervicovaginal abnormalities were sequentially enrolled at a referral center. Blood samples were collected for HIV1/2 and HSV-2 tests; cervical swabs collected for HPV and p16/ki-67 tests; urine collected for CT test; and biopsy collected for histology. HIV infection was tested via Determine HIV-1/2 and confirmed via first response HIV1/2 card tests; HSV-2 was tested via HerpeSelect-2 enzyme immunoassay; HPV was tested via GeneXpert HPV assay; p16/ki-67 biomarkers were tested via CINtec PLUS test; CT was tested via GeneXpert CT/NG assay and biopsies were tested via hematoxylin and eosin stains. Categorical variables were summarized using test of proportions and the differences examined using Pearson's Chi-square test. Logistic regression was used to test the associations between coinfections and contraceptives use with incidence of CIN, and McNemar χ2.test used to compare the performance of p16/ki-67 and VIA with a p-value < 0.05 considered as statistically significant. Among 189 colposcopic biopsy confirmed, CIN1, CIN2, ≥CIN3 and normal biopsy were 56(29.6%), 27(14.3%), 12(6.3%), and 94(49.7%) respectively. The overall prevalence of CIN was 18.4% (95/517) with high grade ≥CIN2 of 7.54% (39/517) equivalent to 32.5 per 100,000 women per year. HPV/HIV coinfection (infected vs. uninfected: aOR 4.45; 95% CI 2.53–7.92, p < 0.001); HPV/HSV-2 coinfection (infected vs. uninfected: aOR 5.67, 95% CI: 2.61–12.40, p <0.001); HPV/CT coinfection (infected vs. uninfected: aOR 6.03; 95% CI 3.00-12.2, p < 0.001) were significantly associated with CIN, but not contraceptives use [aOR 0.73, 95% CI 0.41–1.25, p = 0.079]. Regarding the diagnostic performance of p16 /ki-67 and VIA; the sensitivity of p16/Ki-67 test was 84.6% (95%CI: 69.5-94.1) greater than VIA 59.0% (95%CI: 42.1–74.4), p = 0.024; specificity was 44.0% (95%CI: 35.9-52.3) lower than VIA 62.0% (95%CI: 53.7- 69.8); p= 0.003; PPV was 28.2 (95%CI, 21.7–36.3) similar to VIA 28.8% (95%CI, 28.1 – 34.5); p = 0.251; NPV was 91.7% (95%CI, 87.8–94.7) greater than VIA 85.3% (95%CI, 80.0–89.7); p = 0.002 with a fair agreement of kappa 0.109; (95%CI, 0.02-0.196), p= 0.012. In conclusion, coinfections with HIV, HSV-2 and CT were associated with CIN but not contraceptives use. Immunostain p16/ki-67 is a fairly improved diagnostic test than VIA and can serve as an alternative test.