ASCL1 and LHX8 methylation in HPV-positive self-collected samples 103 4 samples to paired clinician-collected cervical samples and cervical tissue specimens. Our findings on agreement are in line with previous data comparing methylation marker performance in paired samples, reporting moderate to good agreement 16, 18, 20, 22, 23, 41, 42. A limitation of our study is that our results could have been affected by verification bias. We did not have a histology endpoint for hrHPV-positive women with two consecutive normal cytology results, given that these women were referred to the next screening round in accordance with the current guidelines of the Dutch screening program. Nonetheless, this effect seems minimal hrHPV-positive women with two consecutive normal cytology results have a very low risk of CIN3+. In conclusion, the ASCL1/LHX8 methylation marker panel, alone or in combination with genotyping, constitutes a feasible direct triage method for detecting CIN3+ in hrHPVpositive women participating in routine screening by self-sampling. Our results support further clinical validation in prospective screening studies using HPV self-sampling.
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