Phage display for site-specific immunization and characterization of high-risk human papillomavirus specific E7 monoclonal antibodies

The paper describes a method to use filamentous phage to display specific regions of proteins for immunization in order to direct the immune response towards a pre-defined region of the protein. The method called site-specific immunization (SSI) was evaluated using the E7 protein of oncogenic (high-risk) human papillomavirus (HPV) type 16 as a model system. This protein consists of sequence blocks also present in other viral and cellular proteins and in the corresponding protein of low-risk HPVs. A fragment of the HPV16 E7 oncoprotein specific for a group of high-risk viruses was identified by sequence comparison and displayed on filamentous phages in fusion with the major phage coat protein VIII. The recombinant phages triggered an immune response in mice against the full-length HPV16 E7 protein. Fusion of B-lymphocytes from the immunized animals with myeloma cells resulted in three hybridomas producing monoclonal antibodies (MAbs) with reactivity against the endogenous E7 protein. The specificity of the MAbs for the HPV16 E7 protein in cancer cell lines was confirmed by Western blot analyses and immunocytochemistry. The epitope of each MAb was roughly mapped by determining the reactivity against overlapping E7 fragments displayed on phage particles. The mimotopes of the MAbs were further determined by biopanning against a randomized peptide library displayed on phage and found to be unique for a sub-set of high-risk HPV E7 proteins. The combination of different phage display techniques for immunization and epitope mapping was efficient for generation and characterization of highly specific MAbs.     

人乳头瘤病毒检测对宫颈癌术后局部复发的预测性分析

目的 本研究主要就人乳头瘤病毒(HPV)检测在宫颈癌术后局部复发预测方面的应用价值展开分析探讨,以此来为宫颈癌术后复发的临床诊断提供参考.方法 选择我院2012年12月-2013年12月所收治的100例宫颈癌术后局部复发患者作为研究对象,利用随机数字法将其均分成两组,对照组的患者给予鳞状上皮癌抗原(SCC)检测,观察组患者给予HPV检测,比较两组患者的预测准确性.结果 观察组患者的阳性率为94％,对照组患者的阳性率为64％,两组患者的预测准确性存在明显差异,具有统计学意义(P＜0.05).结论 在对宫颈癌患者进行术后随访时,HPV可以有效地预测患者在术后是否存在局部复发的情况,其具有较高的临床应用价值,在宫颈癌患者的术后随访过程中可以大力推广并普及使用.     

多重PCR法和HPV分型基因芯片法在高危型人乳头瘤病毒感染检测中的应用

目的评价高危型人乳头瘤病毒（HPV）多重核酸扩增荧光检测法和HPV分型基因芯片检测法在HPV感染女性患者标本基因分型的临床应用效果。方法应用13种高危型HPV多重PCR荧光检测法对在深圳市人民医院进行宫颈癌筛查的653例疑似HPV感染女患者的宫颈细胞样本进行检测,并与HPV分型基因芯片检测法的检测结果进行比较,2种方法检测13种高危型HPV不一致的样本经序列分析方法进一步验证。结果13种高危型HPV多重PCR荧光检测法检测HPV阳性样本,阳性检出率为21．5％（140／653）;用HPV分型基因芯片检测法验证,与13种高危型HPV多重PCR荧光检测法一致的阳性样本占20．4％（133／653）,总一致率为98．2％。2种方法的检测结果具有高度一致性（kappa值一o．945）;用HPV分型基因芯片检测法检出：HPV单一型别感染占59．4％（79／133）,主要的高危HPV型别为HPV16、52、39、68、33和59型,6种高危型占总数的87．3％（69／79）,其中HPV16和HPV52为主要感染,占44．9％。结论高危型HPV多重PCR荧光检测法和HPV分型基因芯片检测法在13种高危型HPV的检结果具有高度一致性;多重PCR荧光检测法覆盖主要的13种高危型HPV,分型基因芯片检测法可进行具体的单一型别分型。2种检测方法的联合应用,对宫颈癌筛查和预防具有较高的临床应用价值,同时可为HPV分子流行病学和HPV疫苗的应用研究提供依据。     

深圳市妇女宫颈癌组织中人乳头瘤病毒16型E6、E7致癌基因的序列分析

目的 检测深圳市宫颈癌患者癌组织中HPV16型E6、E7致癌基因的序列.方法 采用PCR技术扩增1例在深圳生活17年宫颈癌HPV16型阳性患者组织中HPV16 E6、E7基因,对其进行克隆、构建重组质粒并进行核酸限制性内切酶鉴定及DNA测序分析.结果 成功地克隆了HPV16 E6、E7基因,测序分析表明克隆的HPV16 E6、E7基因与GenBank收录的德国标准株相比完全一致.结论 深圳市妇女宫颈癌组织中HPV16型E6、E7致癌基因的序列与德国标准株相同.     

Cervical Cancer Prevention and High-Risk HPV Self-Sampling Awareness and Acceptability among Women Living with HIV: A Qualitative Investigation from the Patients’ and Providers’ Perspectives

Routine cervical cancer screening is important for women living with HIV (WLH) due to the greater incidence and persistence of high-risk HPV (HR-HPV) infection. HR-HPV self-sampling has been proposed to overcome barriers to in-office cervical cancer screening in underserved populations. However, little is known about baseline knowledge of HR-HPV and the acceptability of HR-HPV self-sampling among WLH. This paper describes WLH’s experiences and needs regarding cervical cancer screening, specifically HR-HPV self-sampling, and seeks to reconcile their experiences with the views of their providers. In total, 10 providers and 39 WLH participated in semi-structured interviews and group discussions, respectively. Knowledge of cervical cancer and HR-HPV was generally limited among WLH; when present, it was often due to personal experience of or proximity to someone affected by cervical cancer. Most WLH were not familiar with HR-HPV self-sampling but, despite some of the providers’ skepticism, expressed their willingness to participate in a mail-based HR-HPV self-sampling intervention and highlighted convenience, ease of use, and affordability as facilitators to the uptake of HR-HPV self-sampling. The experiences identified can be used to guide patient-centered communication aimed at improving cervical cancer knowledge and to inform interventions, such as HR-HPV self-sampling, designed to increase cervical cancer screening among under-screened WLH.     

不同病程尖锐湿疣患者HPV感染与T细胞亚群表达情况的关系

目的探讨不同病程尖锐湿疣(CA)患者人乳头状瘤病毒(HPV)感染与T细胞亚群表达情况的关系。方法选择2012年1月—2015年2月我院收治的151例CA患者,根据病程长短分为A、B组,A组病程≤3个月,B组病程3个月,A组97例,B组54例,另选同期体检健康的51例作为对照组。采集3组患者外周血,并使用流式细胞术检测T淋巴细胞亚群的表达情况。取脱落疣体细胞,并使用核酸快速杂交分型法检测HPV-DNA。结果 B组患者CD4~+、CD4~+/CD8~+水平显著低于A组和对照组,CD8~+水平显著高于对照组及A组患者,差异均有统计学意义(P0.05)。HPV阳性患者151例,其中包括单纯HPV6/11~+、HPV16/18~+型感染各45例、49例,HPV6/11、16/18~+混合型感染57例。HPV6/11~+、HPV16/18~+、(HPV6/11~+、16/18~+)各组患者CD3~+、CD4~+、CD4~+/CD8~+水平均低于健康组,差异有统计学意义(P0.05);而3组间CD3~+、CD4~+、CD8~+、CD4~+/CD8~+水平比较,差异均无统计学意义(P0.05)。结论病程3个月以上CA患者细胞免疫功能明显降低,且各型HPV感染患者细胞免疫功能差异无统计学意义。     

子宫颈细胞学检查未见异常的HR-HPV亚型感染妇女的CIN2,3风险分析

目的:探讨子宫颈细胞学检查未见异常的HPV高危亚型感染者的管理模式。方法:收集2010年1月至2012年12月在北京大学第一医院妇产科门诊同时行宫颈细胞学检查及HPV DNA分型检测的妇女的资料,分析初次检出细胞学未见异常者的HPV高危亚型16、18、31、33感染者,其检出CIN2及以上病变的风险以及与感染亚型的相关性。结果:993例细胞学检查未见异常但HPV16、18、31、33型阳性者中,共检出CIN1 76例(7.7%),CIN2 50例(50/993,5.0%),CIN3 27例(27/993,2.7%);其中HPV16(+)感染者532例(532/993,53.6%),检出CIN2 34例(34/532,6.4%),CIN3 21例(21/532,3.9%);HPV18(+)HPV16(-)感染者142例(142/993,14.3%),检出CIN2 2例(2/142,1.4%),CIN3 1例(1/142,0.7%);HPV31(+)HPV16\18(-)感染者137例(137/993,13.8%),检出CIN2 9例(9/137,6.6%),CIN3 2例(2/137,1.5%);HPV33(+)HPV16\18(-)感染者182例(182/993,18.3%),检出CIN2 5例(5/182,2.7%),CIN3 3例(3/182,1.6%)。按是否检出CIN2+进行Logistic回归分析,发现HPV16型感染与CIN2+有相关性[OR值=2.353(95%CI 1.004~5.516),P=0.049]。结论:对筛查中初次检出宫颈细胞学未见异常,但HPV高危亚型感染者应予以重视,对于HPV16、18型感染者建议立即行阴道镜检查。     

Progressive increase of human papillomavirus carriage rates in potentially malignant and malignant oral disorders with increasing malignant potential

Introduction:  We investigated the potential role of human papillomaviruses (HPVs) in potentially malignant oral disorders, oral leukoplakia (OL) and oral lichen planus (OLP), and in oral squamous cell cancer (OSCC) in an Eastern Hungarian population with a high incidence of OSCC.
Methods:  Excised tumor samples (65 OSCC patients) and exfoliated cells from potentially malignant lesions (from 44 and 119 patients with OL and OLP, respectively) as well as from healthy controls (72 individuals) were analysed. OLPs were classified based on clinical appearance, 61 patients had erosive–atrophic lesions (associated with higher malignancy risk, EA-OLP) and 58 had non-erosive non-atrophic lesions (with lower risk of becoming malignant, non-EA-OLP), respectively. Exfoliated cells collected from apparently healthy mucosa accompanied each lesion sample. HPV was detected by MY/GP polymerase chain reaction (PCR) and genotyped by restriction analysis of amplimers. Copy numbers in lesions were determined using real-time PCR. Prevalence rates, copy number distributions, and association with risk factors and diseases were analysed using chi-square test, t -test, and logistic regression, respectively.
Results:  We detected HPVs significantly more frequently in lesions than in controls ( P  ≤ 0.001 in all comparisons). HPV prevalence increased gradually with increasing severity of lesions (32.8, 40.9, and 47.7% in OLP, OL, and OSCC, respectively). Copy number distribution patterns roughly corresponded to prevalence rates, but OLP and OL were comparable. HPV prevalence differed significantly between EA-OLP and non-EA-OLP groups (42.6 vs. 22.4%); EA-OLP group showed a prevalence similar to that found in OL.
Conclusion:  HPVs may be involved in the development or progression of not only OSCC but also of potentially malignant oral lesions.