Methylation of CpG sites in the upstream regulatory region, physical status and mRNA expression of HPV-6 in adult-onset laryngeal papilloma

// Zeyi Deng 1, 2 , Taro Ikegami 2 , Asanori Kiyuna 2 , Chunlin Zhang 1, 2, 3 , Tao Zhang 1 , Sen Matayoshi 2 , Takayuki Uehara 2 , Hiroyuki Maeda 2 , Mikio Suzuki 2 and Akira Ganaha 2 1 Department of Otorhinolaryngology, Head and Neck Surgery, Zhujiang Hospital, Southern Medical University, Guangzhou, China 2 Department of Otorhinolaryngology, Head and Neck Surgery, Graduate School of Medicine, University of the Ryukyus, Okinawa, Japan 3 Department of Otorhinolaryngology, Head and Neck Surgery, Affiliated Hospital of Zunyi Medical College, Zunyi, China Correspondence to: Zeyi Deng, email: jordandzy@hotmail.com Keywords: adult-onset laryngeal papilloma, human papillomavirus-6, methylation, physical status, mRNA expression Received: November 05, 2016      Accepted: July 13, 2017      Published: August 03, 2017 ABSTRACT The methylation status of HPV-6 upstream regulatory region (URR) in adult-onset laryngeal papillomatosis (AO-LP) remains unclear. The purpose of this study was to investigate the methylation status of URR and the physical status of HPV-6, as well as the dynamic variations of viral load and mRNA expression in AO-LP. We examined 18 specimens from 11 patients with AO-LP by real-time polymerase chain reaction (PCR), bisulfite-sequencing PCR, and amplification of papilloma oncogene transcripts. HPV-6 was identified in 9 of 11 patients (81.8%), and all the 15 specimens derived from 9 HPV-6-positive cases contained only episomal HPV-6 transcripts with intact E2 . Three HPV-6-positive patients developed recurrent lesions, and HPV-6 copy numbers and mRNA expression decreased after surgical treatment. Among the 96 CpG sites (16/case), 67 (69.8%) were unmethylated, while 23 (30.2%) were heterogeneous (≥ 1 methylated CpG clone). High viral loads and episomal status of HPV-6 were frequently observed in AO-LP; thus, persistent E6/E7 mRNA expression of LR-HPV-6 may be associated with AO-LP recurrences. Hypomethylation and scattered patterns of methylated CpGs at the URR of HPV-6 were identified in AO-LP.