Differences in serological IgA responses to recombinant baculovirus-derived human papillomavirus E2 protein in the natural history of cervical neoplasia

Abstract

Summary Infection with certain types of human papillomavirus (HPV) presents a high risk for the subsequent development of cervical intraepithelial neoplasia (CIN) and cervical carcinoma. Immunological mechanisms are likely to play a role in control of cervical HPV lesions.The HPV E2 protein has roles in virus replication and transcription, and loss of E2 functions may be associated with progression of cervical neoplasia. Accordingly, it is of interest to monitor immune responses to the E2 protein, and previous studies have reported associations between serological reactivity to E2 peptide antigens and cervical neoplasia. In order to investigate serological responses to native, full-length E2 protein, we expressed HPV-1 6 E2 proteins with and without an N-terminal polyhistidine tag using the baculovirus system. Purified HPV-1 6 E2 protein was used to develop enzyme-linked immunosorbent assays to detect serological IgG and IgA responses in cervical neoplasia patients and controls. We found that serum IgA levels against the E2 protein were elevated in CIN patients relative to normal control subjects but were not elevated in cervical cancer patients. Moreover, there appeared to be a gradient of response within cervical neoplasia such that the highest antibody levels were seen in lower grades of neoplasia up to CIN 2, whereas lower levels were observed in CIN 3 and still lower levels in cervical carcinoma. These findings suggest that the IgA antiboay response to E2 may associate with stage and progression in cervical neoplasia.Keywords: human papillomavirus; serology; E2 protein; cervical neoplasia; baculovirus Human papillomaviruses comprise a large group of DNA viruses that exclusively infect epithelium. Certain types of HPV are capable of infecting genital mucosal epithelium, which can result in cervical intraepithelial neoplasia (CIN). CIN is classified by histopathology into stages 1, 2 and 3, which are thought to represent progressively advanced precursor lesions of cervical carcinoma (CaCx). HPV infection is a prerequisite for the genesis of almost all CIN and cervical carcinomas Schiffman et al, 1993). Most CIN contains detectable HPV DNA. HPV-16, -18 and related types have been classed as 'high-risk' genital HPV types because of their associations with high-grade CIN and CaCx (reviewed in Walboomers et al, 1994).Genital-type HPV genomes are approximately 8 kbp in length and comprise six open reading frames (ORFs) encoding early functions (E1-E7) and two late ORFs encoding the capsid proteins. The E2 ORF encodes an approximately 45-kDa nuclear phosphoprotein that binds to specific sequence elements within the HPV long control region (LCR). Binding of the E2 protein to these elements functions in regulation of HPV transcription and replication. Binding of E2 to sites near the constitutive early promoter of HPV-16 or -18 can negatively regulate expression of the two major

Expression patterns of the human papillomavirus type 16 transcription factor E2 in low- and high-grade cervical intraepithelial neoplasia

“…Serological studies of anti-E2 IgA in serum from a separate group of patients 23 have indicated that IgA levels against E2, previously high in CIN I, are significantly lower and almost undetectable in higher-grade CIN patients. It should be possible to develop an ELISA or quantitative test to automate such detections, which will be less fraught with false-positive and sensitivity issues than PCR-based methods, not only on histological sections taken from cervical biopsies, but also in smear samples to complement Papanicolou stain, although the significance of this aspect would require further testing in a properly controlled epidemiological study of larger patient numbers.…”

The oncogenic potential of human papillomaviruses: a review on the role of host genetics and environmental cofactors

“…A relationship between the level of serum IgA anti‐E2 antibodies and cervical neoplasia progression as well as tumour stage has been demonstrated. As suspected, lower levels of IgA anti‐E2 antibodies correlated with tumour progression 68 . The significance of this finding is unclear, especially as the E2 gene is frequently deleted in CC cells 69 .…”

“…Multiple HPV types have been implicated in the promotion of cervical SCC development. The HPV types found in cervical neoplastic cells were classified by Muñoz et al into those associated with low (6, 11, 13, 40, 42-44, 54, 61, 62, 70, 72, 74, 81) and high (16,18,31,33,35,39,45,51,52,56,58,59,68,73,82) risk of progression to malignancy. In addition, the authors concluded that HPV types 26, 53 and 66 should be considered probably carcinogenic due to the extremely small number of affected patients.…”

Serology for human papillomavirus

“…4 A study which used E2 produced in insect cells indicates that there is a correlation between the detection of anti-E2 IgA and severity of lesions, although these antibodies disappear over the course of neoplasia evolution. 5 Unfortunately, the sensitivity and specificity of these assays have been evaluated in only a handful of studies. Given that there is a strong upregulation of E4 production throughout the viral cycle, anti-E4 antibody has been proposed as a marker for viral replication.…”