1, common errors in human papillomavirus detection
Myth 1: Detection of low-risk human papillomavirus has clinical value.
Clinically, we often see that patients' HPV test reports contain low-risk HPV. In fact, it is a misunderstanding to detect low-risk human papillomavirus, and it is mistakenly believed that low-risk human papillomavirus has the same cancer risk as high-risk human papillomavirus. 2015-1-26 China Food and Drug Administration (CFDA) issued the "Guiding Principles for Detection and Genotyping of Human Papillomavirus (HPV) and Technical Review of Reagents", which clarified the type range of HPV detection in China-only for/kloc-0 for the anticipated purpose related to cervical cancer. According to the research results of the World Health Organization (WHO), the International Agency for Research on Cancer (IARC) and other international organizations, it is suggested that HPV 16, 18, 3 1, 33, 35, 39, 45, 5 1, 52, 56. Genotypes 26, 53, 66, 73 and 82 * * * are classified as medium risk types, and all these genotypes only need to be tested for the expected use related to cervical cancer for the above 18 human papillomavirus genotype; At the same time, it is pointed out that low-risk human papillomavirus is generally related to condyloma acuminatum or low-grade squamous intraepithelial lesions, and the clinical value of detection is not clear. Therefore, it is a misunderstanding that the detection of low-risk human papillomavirus can not play a role in cervical cancer screening.
Myth 2: The purpose of human papillomavirus detection is to find out whether there is virus or not.
80% of women may be infected with human papillomavirus all their lives, and most of them are too cute to be removed by their own immune system, so they will not have pathological changes, which means that infection is not equal to pathological changes. Therefore, human papillomavirus testing is used to find patients with cervical lesions [such as cervical intraepithelial neoplasia (CIN)2+], not to find out whether there is virus! At present, the detection technology of human papillomavirus can not meet the clinical needs well. The ideal detection method of human papillomavirus needs high clinical sensitivity and specificity. Clinical sensitivity is different from analytical sensitivity. The former requires a long time for large samples to obtain clinical verification tests, while the analytical sensitivity refers to the lowest number of copies or titers of human papillomavirus that can be detected by the detection method under laboratory conditions. The former aims to find patients clinically, and the latter aims to find out whether human papillomavirus exists. Therefore, a good clinical human papillomavirus detection method can not only ensure a very high detection rate for CIN2+ patients, but also minimize the high false positive rate caused by the inability to determine the clinical detection threshold (cutoff value) without clinical verification. In the guide "Performance Requirements for Detection of Human Papillomavirus as a Diagnostic Reagent in vitro", CFDA clearly pointed out that if a human papillomavirus detection technology needs to be approved, it must have a certain critical value, which is the boundary between positive and negative clinical trials. In the 20 13 who guidelines for screening and management of cervical cancer, there are specific suggestions for setting the critical value of human papillomavirus detection, suggesting that the critical value of high-risk human papillomavirus detection should be ≥ 1.0 ng/L, however, even the positive predictive value of FDA-certified high-risk human papillomavirus detection technology is not high enough (4.3% ~ 20./kloc. In clinical practice, the use of HPVDNA detection methods without clinical verification test evaluation is easy to lead to overdiagnosis, trigger a series of social problems and increase medical expenses.
Myth 3: The higher the quantitative detection value of human papillomavirus, the more serious the lesion.
At present, there is no quantitative detection method for human papillomavirus in all clinical detection methods; Judging from the existing detection methods, it is difficult to trace back or repeat, which is the fundamental reason why quantitative detection cannot be realized. Hybrid capture (HC2) human papillomavirus detection technology uses the relative light unit/cutoff clinical threshold (RLU/ carbon monoxide, relative light tuning/cutoff) to detect high-risk human papillomavirus. Many clinicians mistakenly believe that the higher the RLU/ CO value, the more serious the lesion, and the lower the RLU/ CO value, the lighter the lesion. In fact, as long as human papillomavirus is positive (RLU/CO≥ 1.0), CIN and cervical cancer can be caused regardless of RLU/CO value. A study of 349 cases of ASC-US HC2 positive cytology found that the median RLU/ CO of normal cytology, CIN 1 and CIN2+ were 42.68, 146.45 and 156.43, respectively, and the confidence intervals of the three groups overlapped greatly. The conclusion is that RLU/CO value is the same as CIN. It should be noted that the RLU/CO value in this study is the sum of the human papillomavirus viral load of the subjects, that is, if multiple subtypes are infected, the RLU/CO value represents the sum of the viral loads of all positive subtypes. For women infected with only one subtype of human papillomavirus (such as HPV 16), the higher the measured value, the higher the risk of CIN2+ (HR:1.34,95% CI1.10 ~1.64 In a word, there is no absolute correspondence between the detection value of human papillomavirus and the severity of the disease.
Myth 4: The detection results of different human papillomavirus detection technologies should be the same.
In fact, there are many products for detecting human papillomavirus in clinic, and the detection results of different products will be different due to different target gene fragments, methods and subtypes of human papillomavirus. At present, * * * four human papillomavirus detection technologies have passed the FDA's screening certification for cervical cancer, namely HC2, Cervista, Cobas and Aptima, in 2003, 2009, 201April and 201June, respectively. HC2 detected 13 high-risk human papillomavirus (HPV 16, 18, 3 1, 33, 35, 39, 45, 5 1, 52, 56, 52. That is, E 1, E2, E4, E5, E6, E7, L 1, L2, LCR * * 9 gene fragments, because HC2 detection does not need primary amplification, it is little affected by cross-contamination and sample collection factors, which may sometimes affect PCR detection and results; Cervista uses Invader, a signal amplification method for detecting specific nucleotide sequences, to detect L 1, E6 and E7 gene fragments of 14 high-risk human papillomavirus (13 and 66 subtypes); Cobas uses the target amplification method of polymerase chain reaction (PCR) to detect the above-mentioned 14 high-risk HPVL 1 gene fragment, which is the only human papillomavirus detection technology approved by FDA for cervical cancer screening in the United States. Aptima used the target amplification method of transcription-mediated amplification (TMA) to detect E6 and E7mRNA fragments of the above-mentioned 14 high-risk human papillomavirus. Therefore, even if the FDA-certified human papillomavirus detection technology has different target gene fragments, methods and subtypes, the results may be different.
Studies have shown that 65% ~ 75% of cervical cancer is caused by HPV 16 and 18 subtypes, and other high-risk types account for 25% ~ 35%. Therefore, human papillomavirus subtypes 16 and 18 have higher cancer risk than other subtypes, which is also an important reason why the FDA approved cobas HPV 16, 18 and non-18 typing detection methods for primary screening of women over 25 years old. It is noteworthy that, unlike the first three HPVDNA detection technologies, the detection target of Aptima is HPVE6?
2. The infection process of human papillomavirus
So how does human papillomavirus infect the cervix? Human papillomavirus (HPV) is an epithelial virus with a diameter of about 50-60 nm, and there are about 150 subtypes, of which 35 can infect female reproductive tract. According to the risk of virus and tumor, human papillomavirus can be divided into low-risk type and high-risk type, and HPV 16 and 18 are the most dangerous. The process of human papillomavirus infection is: firstly, the virus binds to heparan sulfate protein receptor (HSPG) on the basement membrane, and then the susceptible site of L2 is exposed and digested by N- precursor protein invertase. At the edge of infected epithelium, the unexposed area of L 1 binds to an unknown second receptor, and early inclusion bodies are formed in 2-4 hours by phagocytosis, and late inclusion bodies are formed in 3- 12 hours. Viruses.
Human papillomavirus (HPV) infects squamous epithelial basal layer through mucosal minimally invasive surface. After the virus removes its shell, DNA enters the host nucleus, and then the E6/E7 oncogene is first expressed, which accelerates the division of infected cells. Human papillomavirus DNA moved up with splinter cell, and E 1/E2/4/5 gene was expressed in turn with the division of infected cells. E 1/E2 is related to virus gene replication, and E4 protein is related to the last stage of virus gene. When the infected cells enter the terminal differentiation stage, the virus coat protein L 1 and L2 genes begin to express, and the newly assembled virus falls off with the dead epithelial cells. With the extension of human papillomavirus infection time, the proportion of DNA integration increases, and the risk of pathological changes increases.