Methylation at 3′LCR of HPV16 can be affected by patient age and disruption of E1 or E2 genes

Amaro-Filho, Sergio M.; Bertoni, Neilane; Brant, Ayslan Castro; Vidal, João Paulo Castello Branco; Felix, S.; Cavalcanti, Sílvia M. B.; Carestiato, Fernanda Nahoum; Martins, Luciana; Almeida, Liz Maria de; Moreira, Miguel Ângelo Martins

doi:10.1016/j.virusres.2017.01.022

Cited by 8 publications

(10 citation statements)

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“…In this paper, we compare and describe type-specific characteristics of HPV16, HPV18 and HPV45 in CC samples with respect to 3′LCR methylation and disruption of E1/E2 open reading frames (ORFs). Moreover, we intent to verify whether methylation was associated with E1/E2 disruption in HPV18 and HPV45, as previously reported in HPV16 [17] .…”

Section: Introductionmentioning

confidence: 72%

“…Samples were collected at diagnosis from patients treated at the Instituto Nacional de Câncer (INCA - Rio de Janeiro, Brazil) referred for treatment between June 2011 and March 2014. The set of HPV16 + and HPV18 + samples was selected as previously reported [17] in according to their histopathological presentation while all HPV45 + tumors were included in view of their lower number in the pool with respect to HPV16 + and HPV18 + samples ( Table 1 ). DNA isolation and HPV identification was as previously described [27] .…”

Section: Methodsmentioning

confidence: 99%

“…PCR products were submitted to pyrosequencing in a PyroMark Q24 platform (Qiagen, Hilden, Germany) following a standard protocol [17] . Briefly, streptavidin beads, PyroMark binding buffer (Qiagen, Hilden, Germany) and PCR products were mixed and incubated on a shaking platform.…”

Section: Methodsmentioning

confidence: 99%

“…The presence of intact E1 / E2 was identified by PCR amplification of overlapping fragments encompassing the E1 and E2 coding regions of HPV16, HPV18 and HPV45 using primers pairs listed in Table 3 , with some of them previously published [32] , [33] . PCR was carried out as described [17] with different annealing temperatures shown in Table 3 . Negative reactions were repeated to confirm lack of amplification of target regions.…”

Section: Methodsmentioning

confidence: 99%

“…Methylation at this segment has shown to be complex, with conflicting findings among CC samples; most available data being restricted to the HPV16 genotype [15] , [16] . Moreover, clinical factors have also been found to affect methylation levels [17] .…”

Section: Introductionmentioning

confidence: 99%

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HPV DNA methylation at the early promoter and E1/E2 integrity: A comparison between HPV16, HPV18 and HPV45 in cervical cancer

Amaro-Filho

Chaves

Felix

et al. 2018

Papillomavirus Research

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ObjectivesTo compare and describe type-specific characteristics of HPV16, HPV18 and HPV45 in cervical cancer with respect to 3′LCR methylation and disruption of E1/E2.MethodsThe methylation level of 137 cervical cancer samples (70 with HPV16, 37 with HPV18, and 30 with HPV45) of Brazilian patients was analyzed by pyrosequencing. PCR amplifications were performed to characterize E1 and E2 disruption as an episomal surrogate.ResultsThe 3′LCR of HPV16 showed a higher methylation at all CpG sites (7%, 9%, 11%, 10% and 10%) than homologous HPV18 regions (4%, 5%. 6%, 9% and 5%) and HPV45 regions (7%, 7% and 5%). Presence of intact E1/E2 was associated with higher HPV16 and HPV18 methylation levels at all CpG sites (p < 0.05). Disruption of E1/E2 was more frequently found in HPV45 (97%) and HPV18 (84%) than in HPV16 DNA (30%). HPV16 disruption was more frequently found in E1 (48%) unlike HPV18, where it was found in E2 (61%). Concomitant disruption of E1/E2 was most frequent in HPV45 (72%).ConclusionsThe findings showed a higher methylation associated with intact E1/E2 for HPV16 and HPV18. The closely phylogenetic related HPV18 and HPV45 share a similar methylation level and the frequency of viral genome disruption.

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Section: Introductionmentioning

confidence: 72%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%