University of Groningen
Methylation analysis for the identification of cervical lesions to improve cervical cancer screening in a Chinese population
Li, Na
DOI:
10.33612/diss.134442856
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Publication date: 2020
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Li, N. (2020). Methylation analysis for the identification of cervical lesions to improve cervical cancer screening in a Chinese population. University of Groningen. https://doi.org/10.33612/diss.134442856
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Chapter 6
Chapter 6
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Summary
Cervical cancer is one of the most life-threatening diseases in women worldwide. The incidence of cervical cancer in western countries has dramatically decreased, among other factors due to the introduction of national population-based cervical screening programs 1. However, cervical cancer screening needs further improvement. The widely used cytology-based screening methodology has important limitations such as low sensitivity and subjective interpretation of morphological alterations 2, 3. More and more countries are therefore shifting to adopt high-risk human papillomavirus (hrHPV) testing as primary population-based screening programs due to its improved sensitivity for the detection of cervical intraepithelial neoplasia 2 lesions or worse (CIN2+). The lower specificity of the hrHPV test, however, especially in younger women, may lead to many unnecessary colposcopy referrals and over-treatment 4. To reduce these
unwarranted referrals, a triage test for hrHPV-positive women is urgently needed.
DNA promoter hypermethylation is now recognized as an early and frequent epigenetic alteration in cervical cancer 5, 6. At the University Medical Centre Groningen (UMCG), multiple
methylation markers with high sensitivity and high specificity to detect CIN2+ lesions were identified and their diagnostic performance was validated in a Dutch cytologically abnormal cohort, derived from the “old” population-based screening program, based on primary cytological examination and triage by hrHPV testing. To analyze whether these Dutch methylation markers are also predictive for CIN2+/CIN3+ (e.g. sensitivity/specificity) using cervical scrapings collected from women from different geographic areas and races, in chapter 2 women from a Chinese cohort were investigated. Methylation analysis for six markers (ANKRD18CP, C13ORF18, EPB41L3, JAM3, SOX1 and ZSCAN1) was performed using quantitative methylation-specific PCR (QMSP) in cervical scrapings. All 6 individual markers showed enhanced methylation levels and higher frequencies with increasing severity of the underlying lesion. In samples with an abnormal cytology, sensitivity to detect CIN2+ lesions was 79%, 76% and 72% for the 3 panels (C13ORF18/EBP41L3/JAM3, C13ORF18/ANKRD18CP/ JAM3 and ZSCAN1/SOX1, respectively) with a specificity of 57%, 65% and 68%. For the first 2 panels these diagnostic characteristics were similar to those observed in a Dutch cohort, while for ZSCAN1/SOX1 the sensitivity was higher in the Chinese cohort, but with a lower specificity. In hrHPV-positive samples, similar sensitivity and specificity for the detection of CIN2+ were
found as for the abnormal cytology cohort, which were also non-inferior to HPV16/18 genotyping. The latter technique has also been proposed as a possible triage tool. Our analysis revealed that the diagnostic performances were highly comparable for C13ORF18/EBP41L3/ JAM3 and C13ORF18/ANKRD18CP/JAM3 methylation marker panels in both cytological abnormal and hrHPV-positive women in both Chinese and Dutch cohorts. In conclusion, methylation panels as identified in a Dutch population might also be applicable for triage testing in cervical cancer screening in China.
Numerous other studies have been reported on the identification and/or clinical validation of various methylation markers to improve the early diagnosis of cervical cancer. In chapter 3, a systematic review was performed to collect and summarize studies that report on the analysis of methylation markers for the early detection of (pre-) malignant cervical cancer in hrHPV-positive cervical scrapings and/or self-sampled materials by QMSP, and to identify those methylation markers with the highest diagnostic potential. In this review, thirty-six unique genes described in 29 studies were used for analysis. Six genes (CADM1, EPB41L3, FAM19A4, JAM3, MAL and POU4F3) were observed with a sensitivity of 53%-87% and specificity of 60%-95% to detect CIN2+ in hrHPV-positive women. The highest combined sensitivity with specificity to detect CIN2+ lesions was reported for JAM3/ANKRD18CP (76% and 83%) and C13ORF18/JAM3/ANKRD18CP (77% and 81%), respectively.
Based on this systematic review, two methylation markers (PAX1 and ZNF582) were further investigated for their potential role for early cervical cancer detection in an Asian population. In
chapter 4 a meta-analysis on ZNF582 was performed, comprising 7 studies with in total 1749
patients from different Chinese cohorts and irrespective of hrHPV status. The pooled sensitivity of ZNF582 methylation for CIN3+ was estimated to be 71% with a specificity of 81% and an area under the curve (AUC) of 0.85. Our analysis suggested that a sequential combination of hrHPV and ZNF582 methylation test (AUC, sensitivity and specificity of 0.876, 75% and 87%, respectively) achieved higher diagnostic accuracy than a single hrHPV test (AUC, sensitivity and specificity of 0.669, 96% and 41%, respectively).
As the diagnostic performance of methylation analysis of PAX1 and ZNF582 so far was only estimated in women from Asian populations, in chapter 5 their potential diagnostic role was evaluated in parallel in a Dutch and Chinese cohort (as described in chapter 2). PAX1 and
Chapter 6
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ZNF582 methylation showed high specificity (91-97%) for CIN2+ in both abnormal cytology and hrHPV-positive samples with moderate sensitivity (43-61%) in both cohorts. Sensitivity for CIN3+ improved to 55-86%, but with a lower specificity (85-93%). In both the abnormal cytology and hrHPV-positive samples, sensitivity and specificity to detect CIN2+ lesions were similar between both cohorts. Sensitivity could be improved further with ~20% by combinations of PAX1 or ZNF582 with different previously identified CIN2+-specific methylation markers (ANKRD18P, C13ORF18, EPB41L3, JAM3, SOX1 and ZSCAN1). The diagnostic performance of PAX1 or ZNF582 methylation panels showed higher specificity to detect CIN2+/CIN3+ lesions as triage test either in abnormal cytology or hrHPV-positive scrapings. These markers as triage test might significantly decrease the number of unnecessary colposcopies. However, randomized controlled trials and further large prospective studies are needed to confirm our findings.
References
1. Peto, J., C. Gilham, O. Fletcher, et al., The cervical cancer epidemic that screening has prevented in the UK. Lancet, 2004. 364(9430):249-56.
2. Greene, J., The Pap Test: 20th Century Success Story, 21st Century Has-Been. Manag Care, 2018. 27(2):24-7. 3. Mustafa, R.A., N. Santesso, R. Khatib, et al., Systematic reviews and meta-analyses of the accuracy of HPV
tests, visual inspection with acetic acid, cytology, and colposcopy. Int J Gynaecol Obstet, 2016. 132(3):259-65.
4. Aitken, C.A., H.M.E. van Agt, A.G. Siebers, et al., Introduction of primary screening using high-risk HPV DNA
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5. Kelly, H., Y. Benavente, M.A. Pavon, et al., Performance of DNA methylation assays for detection of
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6. Kremer, W.W., R.D.M. Steenbergen, D.A.M. Heideman, et al., The use of host cell DNA methylation
analysis in the detection and management of women with advanced cervical intraepithelial neoplasia: a review. Bjog, 2020.
