Infection with Human papillomaviruses (HPVs) leads to the development of a wide-range of cancers, accounting for 5% of all human cancers. A prominent example is cervical cancer, one of the leading causes of cancer death in women worldwide. It has been well established that tumor development and progression induced by HPV infection is driven by the sustained expression of two oncogenes E6 and E7. The expression of E6 and E7 not only inhibits the tumor suppressors p53 and Rb, but also alters additional signalling pathways that may be equally important for transformation. Among these pathways, the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signalling cascade plays a very important role in HPV-induced carcinogenesis by acting through multiple cellular and molecular events. In this review, we summarize the frequent amplification of PI3K/Akt/mTOR signals in HPV-induced cancers and discuss how HPV oncogenes E6/E7/E5 activate the PI3K/Akt/mTOR signalling pathway to modulate tumor initiation and progression and affect patient outcome. Improvement of our understanding of the mechanism by which the PI3K/Akt/mTOR signalling pathway contributes to the immortalization and carcinogenesis of HPV-transduced cells will assist in devising novel strategies for preventing and treating HPV-induced cancers. Human papillomaviruses (HPVs) are non-enveloped, epitheliotropic, circular double-stranded DNA viruses ,2. HPV iGenomic instability is a hallmark of carcinogenesis and recognised as an important factor in the accumulation of mutated genes required for carcinogenesis . EndogenIt has been well established that HPV E6 and E7 oncogenes inactivate two tumor suppressors (p53 and pRb) in virus-infected cells. Molecular and cell biology approaches have revealed that alterations of additional signalling pathways are equally important for transformation of HPV oncogene-transduced cells . It is nHPVs whether they are low-risk and high-risk are epitheliotropic. Infection with two HPVs may be latent or active . The lat. In cells with impaired p53 activity, DNA damage repair requires the activation of p38MAPK along with MAPKAP kinase 2 (MK2) [It has been established that the cellular DNA damage response (DDR) is activated during the HPV life cycle . This ac 2 (MK2) . In HPV- 2 (MK2) . Thus, iRas and PI3KCA genes) to stimulate host cell survival and proliferation, leading to carcinogenesis through activation and alteration of the components of the PI3K/Akt/mTOR pathway [Active HPV infection which is also known as abortive infection leads to induction of cancer including benign and malignant neoplasms . In the  pathway ,48-53  . Sixty sAkt phosphorylation is required for a BRCT  domain-containing protein TopBP1 to interact with other transcription factors, especially E2F1. E2F1 functions as a tumor suppressor to induce apoptosis ,67. PhosMammalian genomes contain three Akt genes, Akt1, Akt2 and Akt3. Akt1 encodes the principal Akt isoform that regulates apoptosis . HPVs ma(Ser2448) and p70S6K(Thr389), indicating the constitutive activation and overexpression of the mTOR pathway in HSIL and SCC [mTOR kinase acts as a cellular rheostat that integrates the signals from a variety of cellular signalling pathways to sense growth factor, nutrient availability and energy status. Recently, it has been reported that activation of Akt /mTOR can be detected within several minutes following exposure of human keratinocytes to HPV16 pseudovirions . mTOR ac and SCC . All the(Ser2448) and p70S6K(Thr389) correlates with overexpression of the upstream signal transducer EGFR, increased cell cycles and mitotic indices [mTOR is a crucial metabolic sensor in the growth factor receptor (GFR) pathway, which integrates growth factor signals in cells. The increased nuclear translocation of p-mTOR indices . The act indices . All the indices .A high level of p-mTOR can serve as an independent prognostic marker to predict poor response to chemotherapy and survival of cervical cancer patients . ConcurrRecently, several review papers have provided comprehensive summaries of the biological and biochemical activities of three HPV oncoproteins: E6, E7 and E5 -88. HereHPV E6 oncoproteins are the key players in HPV-induced cancers. The E6 oncoproteins from high-risk mucosotrophic HPVs (α-HPVs) target not only P53, but also a range of host-cell proteins for proteasome-mediated degradation, resulting in alteration of multiple cellular and molecular events -101. A gaa83V) variant is also linked to invasive tumours. The E6aa83V variant activates PI3K signalling pathway and strengthens the possibility of the existence of Ras-independent mechanisms to recreate signalling through classical Ras effector pathways [HPV E6 variants (E6*) can act as an adaptor molecule linking a ubiquitin ligase to target proteins, which contain class 1 PDZ domains and are involved in cell junction stability and signalling . E6* propathways . The varE6 proteins of three HPVs , and BPV1 interact with acidic LxxLL motifs of transcriptional coregulator MAML1 to target many host proteins such as the mammalian target of rapamycin complex 1 (mTORC1) to delay keratinocyte differentiation -111. The(Thr389) and p-S6(Ser235/236) in 140 cervical cancer and 161 oesophageal cancer specimens has revealed that both p-S6K and p-S6 were detected significantly more frequently in the HPV16-infected cervical cancer specimens than those in the HPV16-negative specimens [Generally, β-HPV E6 proteins interact with fewer cellular proteins as is also observed for the α-HPV E6 . This ispecimens . HPV16 Epecimens ,115. Altpecimens . HPV16 Epecimens .NHERF-1 is a molecular pathway organizer that plays an important role in a number of cellular processes including signal transduction, cellular transformation and recruitment of membrane, cytoplasmic, and cytoskeletal signalling proteins into functional complexes . HPV16 EHPV E7 protein is responsible for pRb disruption in HPV-induced carcinogenesis. E7 binds to and inactivates pRb to disturb the normal cell division process, allowing the cells to grow out of control and unhindered and thus become cancerous. Clinically, decreased Rb expression is consistently associated with increased CIN grade in the HPV-infected woman’s cervices. It has been reported that HPV E7 significantly up-regulates Akt activity in differentiated keratinocytes, which depends on the ability of E7 binding to and inactivating the proteins of pRb family  . Up-regCip1 and p27Kip1 [Cip1 binds to the cyclin E/CDK2 complex to maintain Rb in a phosphorylated state [Cip1-dependent cell cycle arrest [Cip1-mediated growth arrest [Cip1 or derepressing p21Cip1-associated CDK2 complexes by sustaining Akt activity [Kip1 and may in turn regulate tumor metastasis/aggressiveness through the PI3K/ Akt pathway [Kip1 to block cyclin E-associated kinase and to inhibit transcription of cyclin A in vitro [Through the PI3K/Akt signalling pathway, HPV E7 oncoprotein inhibits the functions of two cyclin-dependent kinase inhibitors, p21 p27Kip1 ,121. As ed state . In the e arrest . In contactivity ,124. P27activity . HPV 16  pathway . E7 alsoin vitro .. Several studies have reported that activation of Akt induced by HPV E7 expression plays a crucial role in immune resistance [Apoptosis as a normal process of cellular self-destruction or suicide is one of the major contributors to the development of a normal immune system, which serves a protective role in our bodies. In response to oncogenic insults, normal human cells execute a defence response that culminates in apoptosis . In HPV sistance -128. Duesistance . The E7-sistance . Treatmesistance ,130.Furthermore, normal human diploid fibroblasts expressing the HPV16 E7 oncoprotein are predisposed to apoptosis when they are deprived of growth factors such as IGF-1 in serum-starved medium . The apoin vitro cultures resemble in vivo epidermal development when they enter a differentiation program [in vitro proceed to cell differentiation with downregulation of proliferation markers including K14 and K5 and upregulation of differentiation markers such as involucrin and K10 [in vitro cultures induces phosphorylation of AKT on threonine 308 and serine 473 to significantly inhibit cell differentiation and cause hyperproliferation [As mentioned above, keratinocytes are the host cells of HPV infection. In normal epithelial tissues, cell division and proliferation of keratinocytes are confined to the basal layer, where mitogenic signals are balanced by survival signals transmitted through PI3K/Akt pathway . Once in program . We and  and K10 ,135,136.feration ,17. It hferation . The HPVferation . Either feration . The HPVferation , but alsferation .HPV E5 gene encodes an 83-amino acid, membrane-bound protein, which plays an important role in early cervical carcinogenesis by regulating several cellular pathways -141. HPVHPV16 E5 induces the anchorage-independent growth of murine fibroblasts by overexpressing EGFR . HPV16 EHPV E5 may act as a survival factor as the E5-expressing cells in human keratinocyte culture exhibit a significant reduction in UVB-irradiation induced apoptosis . A genomIt is well documented that induction and progression of tumours by HPV infection are driven by the continuing expression of E6 and E7 oncogenes that degrade and inactivate p53 and pRb, respectively ,153. How. Nontheless, whether the PI3K/Akt pathway can play a role in HPV-induced carcinogenesis independent of E6/E7 proteins remains questioned. Thus, more detailed studies are required to improve our current understanding of the full spectrum of mechanisms underlying HPV-induced carcinogenesis.Generally, it is impossible that attachment of HPV pseudovirions and other viruses to the surface of cells activates the PI3K/Akt signalling pathway leading to the development of cancerous cells. However, several other published studies appear to suggest that PI3K signalling pathway plays a critical role in cellular entry of pseudovirions of HPV  and otheRecently, the PI3K/Akt/mTOR pathway has been identified as an important signalling pathway to tightly modulate many cellular events including the physiological activities of mitogenic or oncogenic factors, leading to the genesis of many human cancers. Published studies have shown that expression of HPV E6/E7 oncoproteins induces HPV transformed cells to be cancerous not only causing degradation and destabilization of p53 and pRb, but also altering multiple cellular and molecular events through activation of the PI3K/Akt/mTOR signalling pathway. The PI3K/Akt/mTOR signalling pathway in HPV-infected cells is activated through both mutation of the pathway components and activation of upstream signalling molecules. Activation of this pathway contributes to genetic instability, deregulation of proliferation, resistance to apoptosis, and changes in metabolism characteristics, eventually leading to the malignant transformation of the infected cells. This signalling pathway may potentially represents both a great therapeutic opportunity and a practical challenge for treating HPV-induced cancers. Thus, further understanding of the molecular mechanisms by which HPV infection activates the PI3K/Akt/mTOR signalling pathway and the biological roles of this pathway in HPV-induced carcinogenesis will improve the disease prevention, patient care, and surveillance strategies for HPV-positive cancers. We suggest that one important research direction will be to devise the novel biomarker-driven therapeutic strategies to target the PI3K/Akt/mTOR pathway in HPV-associated cancers with a specific molecular profile and evaluate the efficacy of the potential therapeutic agents.


Nepal has one of the highest cervical cancer rates in South Asia. Only a few studies in populations from urban areas have investigated type specific distribution of human papillomavirus (HPV) in Nepali women. Data on high-risk HPV (HR-HPV) types are not currently available for rural populations in Nepal. We aimed to assess the distribution of HR- HPV among rural Nepali women while assessing self-collected and clinician-collected cervico-vaginal specimens as sample collection methods for HPV screening.Study participants were recruited during a health camp conducted by Nepal Fertility Care Center in Achham District of rural far western Nepal. Women of reproductive age completed a socio-demographic and clinical questionnaire, and provided two specimens; one cervical-vaginal specimen using a self-collection method and another cervical specimen collected by health camp auxiliary nurse midwives during a pelvic examination. All samples were tested for 14 different HR-HPV mRNA and also specific for HPV16/18/45 mRNA.Of 261 women with both clinician- and self-collected cervical samples, 25 tested positive for HR-HPV, resulting in an overall HR-HPV prevalence of 9.6% . The overall Kappa value assessing agreement between clinician- and self-collected tests was 0.62 (95% CI: 0.43–0.81), indicating a “good” level of agreement. Abnormal cytology was reported for 8 women. One woman identified with squamous cell carcinoma (SCC), and 7 women with high grade squamous intraepithelial lesions (HSIL). Seven of the 8 women tested positive for HR-HPV (87.5%) in clinician-collected samples and 6 in self-collected samples (75.0%).This is the first study to assess HR-HPV among rural Nepali women. Self-collected sampling methods should be the subject of additional research in Nepal for screening HR-HPV, associated with pre-cancer lesions and cancer, in women in rural areas with limited access to health services. The World Health Organization (WHO) Information Centre on Human Papillomavirus (HPV) and Cervical Cancer states that the incidence of cervical cancer in Nepal is 24.2 per 100,000, making Nepal a country with one of the highest cervical cancer rates in South Asia To date, information on HPV in Nepal has been limited to study samples drawn from populations in selected urban areas in the central region of the country, with an emphasis on women with invasive cervical cancer There is considerable variation in the limited number of cervico-vaginal and cervical cancer reports across Nepal, with most reports originating from hospital-based registries because there is no national cancer registry The introduction of self-sampling for HPV detection in Nepal could help mitigate the problems of insufficient access to cervical screening. HPV self sampling has been shown to have high rates of agreement with physician collected HPV samples The regional differences in the risk profile for HPV infections and cervical cancer within Nepal necessitate the need for assessing the prevalence and type-specific distribution of HR-HPV infections in different regions of the country. Currently, there are no estimates of HPV prevalence or distribution in Nepal’s rural populations. This is the first study to assess the distribution of HR-HPV and the type-specific distribution of HR-HPV16/18/45 among women in rural far western Nepal. Our study also compares the self-collection of cervico-vaginal specimens with clinician-collected specimens as a method of sample collection for HPV screening in a resource-limited setting.The Institutional Review Board at the University of Alabama at Birmingham and Ethics Review Board at the Nepal Health Research Council approved this study.2 and only 36 of its 75 VDCs are connected by roads. According to the 2011 Nepal Demographic Health Survey, the population of females in Achham District was 141,643, which included 34,204 adolescent girls (aged 10–19), 60,937 women of reproductive age (aged 15–49) and 21,143 seniors (aged 60 years and above) This study was conducted in the Sanphebagar Village Development Committee (VDC) within the Achham District. Achham is one of the most remote districts in far western Nepal. Due to poor and mountainous road conditions, it takes over 13 hours to drive the 500 km from Kathmandu to Mangalsen, the Achham district headquarter. Achham district covers an area of 1,680 kmth, 2013 by Nepal Fertility Care Center (NFCC). Established in 1988, NFCC is a Nepali NGO with the primary objective of complementing and supplementing the Government of Nepal’s national reproductive health programs Women of reproductive age were recruited from the Sanphebagar VDC, approximately 25 km from Mangalsen, during a one day health camp conducted on July 5A questionnaire to assess socioeconomic, clinical, reproductive health, and migration related factors was developed for this study. The questionnaire was first developed in English, then translated into Nepali, and then translated back to English independently for quality control. NFCC staff assisted in the development of the questionnaire in order to insure that the survey questions were culturally sensitive and appropriate. In order to help assure the validity and comprehensiveness of the survey instrument, questions were based on English translations of the Demographic and Health Survey A dialect of the Nepali language known as “Dotyali” is spoken in far western Nepal. However, the term “Dotyali” is a macro-term used to describe a wide variety of local dialects in far western Nepal Each study participant was asked to provide two specimens during her health camp visit. One specimen was self-collected, using the APTIMA Cervical Specimen Collection and Transport (CSCT) kit , and another specimen was collected by a health camp ANM during pelvic examination. While waiting for their pelvic examination, women were given instructions on self-collection of a cervico-vaginal sample. A health staff member, centrally trained to give instructions on self-sampling procedures, instructed the women to insert a cytobrush  into the vagina as far as possible, rotate it 5 times in each direction, then swirl the cytobrush in the APTIMA specimen transport medium . Health camp ANMs collected clinician cervical specimens by inserting a cytobrush into the cervical canal and rotating 3–5 times, withdrawing and then rotating firmly around the full circumference of the transformation zone. Cytobrushes were then swirled in ThinPrep (TP) PreservCyt medium .Both clinician- and self-collected cervico-vaginal samples were transported to the Hologic/Gen-Probe, Inc. laboratory in San Diego for HPV testing. Laboratory testing of HPV was performed using a generic APTIMA HR-HPV mRNA (APTIMA HPV)  and then specific genotyping using APTIMA HPV16 18/45 Genotype  Assays The mean, standard deviation, median, inter-quartile range, and percentages were calculated for self-reported demographic, socioeconomic, and behavioral characteristics. An exact version of McNemar chi-square test was used to test for differences in HPV prevalence in clinician vs. self-collected specimens. Positive and negative agreement percentages and Kappa statistics values were used to describe agreement in detecting HR-HPV between clinician- vs. self-collected samples. We report point estimates for prevalence, % concordance, and Kappa statistics and associated 95% confidence intervals (95% CI). SAS version 9.2  was used to perform all statistical analyses.In total, 389 women registered for the health camp. Three hundred and sixty three women completed a survey; clinician-collected specimens were available from 278 women while self-collected specimens were available from 300 women . Eight oThe median age of the participants was 33 years (IQR: [26–40]) and the median age at first marriage was 17.5 years (IQR 15–19). The median number of children per woman was 3.0 (IQR: [2.0–4.0]) and 56% of women currently used some form of contraception. Over half of the study participants reported having heard of cervical cancer (58.1%), while only 16% reported having heard of HPV .Twenty women tested positive for HPV in clinician-collected samples while 17 women tested positive for HPV in self-collected samples and 25 out of 261 individuals tested positive for HR-HPV in either sampling method, resulting in an over-all HR-HPV prevalence of 9.6% (95% CI 6.3–13.8). Five individuals who tested positive for HR-HPV tested positive for HPV16, resulting in an overall prevalence of 1.9%  for HPV16. Only one individual tested positive for HPV18/45. Thus, among those with HR-HPV, only 24% (6/25) were either type HPV16 or HPV18/45. Five individuals tested positive for HR-HPV in the self-collected specimens, but did not test positive for HR-HPV in the clinician-collected samples. Eight individuals tested positive in clinician-collected samples tests but not in the self-collected samples.The non-stratified Kappa value assessing agreement between the HR-HPV test results using clinician- and self-collected specimens was 0.62 (95% CI 0.43–0.81), indicating a “good” level of agreement . The oveThinPrep (TP)  cervical cytology readings based on clinician-collected samples were available for 278 women. High grade cytological abnormality was reported for 8 women, with one woman identified with squamous cell carcinoma (SCC), and 7 women with high grade squamous intraepithelial lesions (HSIL). Of the 8 women with either SCC or HSIL, 7 of them tested positive for HR-HPV (87.5%) in clinician-collected samples and 6 in self-collected samples (75%). However, 4 of the 7 women who tested positive for HR-HPV tested positive for HPV16 or HPV 18/45 in clinician-collected samples versus 3 in the self-collected samples . SixteenTo the best of our knowledge, this is the first HR-HPV prevalence study conducted among rural women from far western Nepal. Of 261 women with both clinician- and self-collected cervico-vaginal specimens who were tested for 14 HR-HPV types, 25 women tested positive with an overall HR-HPV prevalence of 9.6% (95% CI 6.3–13.8). A non-stratified Kappa value of 0.62 indicated “good” agreement between self- and clinician-collected HPV samples. The cytology of 8 women indicated either SCC or HSIL, of which 7 women who had clinician-collected samples tested positive for HR-HPV and 6 women who had self-collected samples tested positive for HR-HPV.Our study’s prevalence of HR-HPV (9.6%) is higher than the prevalence of HR-HPV (6.1%) found in Bharatpur, Nepal Only one woman tested positive for HPV18/45 , yet HPV genotypes 18 and 45 represented approximately 20% of HPV strains linked to cervical abnormalities in studies conducted by IARC and Bhusal The number of women with an abnormal cytology reading was low. However, these abnormalities pose a significant health risk to women given the dearth of reproductive health care access in rural Nepal. Despite the official Nepali government protocol of cervical screening using visual inspection with acetic acid (VIA), a paltry number of women actually undergo cervical screening each year Non-stratified kappa values assessing the testing agreement between clinician- and self-collected specimens was 0.62 (95% CI 0.43–0.81), indicating a good level of agreement between the two sampling methods. Several studies conducted in rural areas of developing countries which have limited access to cervical screening have also found the results of HPV testing using self-collected samples comparable to the results of HPV testing using clinician-collected samples Only a few studies have assessed the utility of testing for HPV using self-collected samples in less-developed countries This cross-sectional study could not assess lifetime exposure to HPV, nor could it measure HPV persistence, which precedes and predicts the development of cervical precancerous lesions. However, while our study was not longitudinal, HPV prevalence is correlated with the risk of developing cervical cancer In summary, our study assessed the prevalence of HR-HPV among women in a rural region of far western Nepal. Furthermore, our study demonstrated that the self-collected cervical sample for HPV testing is a viable method with a high level of concordance with clinician-collected cervical samples. Self-sampling, while not as sensitive as clinician-collected specimens for detecting prevalent HR-HPV, associated with cervical pre-cancer or cancer


Epstein Barr virus (EBV) and human papillomavirus (HPV) can co-exist in pharyngeal and cervical malignancies. However, the natural history and factors associated with persistent HPV infection among HIV-infected men who have sex with men (MSM) are unclear.131 HIV-infected MSM were followed for 48 weeks and screened for multiple co-infections, including seminal EBV DNA and high risk (HR)-HPV messenger RNA (mRNA) at several sites . Primary analysis tested if seminal EBV shedding was associated with increased prevalence of HR-HPV at baseline using univariate tests and multivariable logistic regression. In participants with detectable anal HR-HPV at baseline, we tested if presence of seminal EBV shedding at baseline was also predictive of reduced HR-HPV clearance by log-rank test (over 48 weeks of follow-up).N = 54/121); pharynx 3.8 % (N = 5/131); semen 7.1 % (N = 7/98). Seminal EBV shedding was present in 28 % of participants and was associated with more than double the prevalence of detectable anal HR-HPV mRNA . In participants with detectable anal HR-HPV at baseline, we found increased persistence of HR-HPV over 48 weeks of follow-up (measured as time to first negative HR-HPV test in the EBV shedding group (p < 0.01).Baseline prevalence of HR-HPV was: anal 44 % (Seminal EBV shedding was associated with an increased risk of having detectable anal HR-HPV in a cohort of HIV-infected MSM on suppressive ART. Future studies should examine if co-infection with EBV and HR-HPV may act synergistically in pathogenesis of anal cancer in HIV-infected individuals. Human Papillomaviruses (HPV) are sexually transmitted and can infect the genital and anal areas, mouth, and throat . Over 10Another common viral infection found in MSM and in the general population is Epstein Barr virus (EBV), which is the cause of infectious mononucleosis . After pThe studies were conducted with appropriate written consent and were approved by the Human Research Protections Program at the University of California San Diego, Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, the University of Southern California and the BioMedical Research Alliance of New York.A total of 179 participants were prospectively enrolled and followed as part of the California Collaborative Treatment Group (CCTG) 592 study, which was an internet-based behavioral intervention study of HIV-infected MSM at high-risk for sexually transmitted infections. At baseline, 131 participants were receiving ART with HIV RNA <500 copies/ml in blood plasma, and were included in this analysis. As part of the study protocol, blood and semen samples were collected at baseline , and anaReal-time PCR was used to measure levels of HIV RNA and different human herpesviruses (HHV) in semen at baseline  .N = 52) and pharyngeal (N = 5) samples tested positive for HR-HPV.Levels of mRNA from the E6/E7 oncogenes for 14 HR-HPV types (16/18/31/33/35/39/45/51/52/56/58/59/66/68) were measured by Aptima HPV Assay  in semen (at baseline), anus and pharynx (baseline and every 12 weeks). Additionally, Aptima HPV 16 18/45 Gentoyping Assay was run on a subset of anal  or Mann Whitney U test . Test of correlation between continuous variables was done with Spearman rank correlation. For the outcome of HR-HPV clearance, we used log-rank test for time to first and second consecutive negative HPV test. The association of EBV and HR-HPV prevalence was analyzed by logistic regression including any factor that was independently associated with HPV considering behavioral factors , blood and seminal plasma HIV levels, current and nadir CD4 T and CD8 T cell count, and genital shedding of other HHV.Statistical analyses were performed using SAS (version 9.2). For this post-hoc analysis, viral load variables were transformed to logarithm-base ten values. The primary analysis tested if the presence of detectable genital EBV at baseline was associated with increased prevalence of HR-HPV infection (at baseline) and reduced clearance of HR-HPV DNA over the following 48 weeks. Continuous variables were tested for normality with the Shapiro-Wilk test, and if not normally distributed, comparisons were performed using nonparametric tests or values were dichotomized. Comparisons between groups  were performed using Fisher-exact test , The majority of study participants (84.0 %) had <50 HIV RNA copies/ml in blood plasma. The median CD4 T cell count was 604 cells/μL IQR: 414–761). Characteristics of this cohort have been previously described ,16, and 4–761. ChAt baseline, 61 (46.6 %) individuals had detectable HR-HPV mRNA at any site for at least one of the 14 HR genotypes tested. HR-HPV mRNA was most commonly detected from anal swabs with a baseline prevalence of 45.0 % . Using the HR-HPV genotype specific assay on anal mucosa samples (for HPV 16 and 18/45), we observed that HPV 16 was detected more frequently than genotypes 18/45 . Overall, at baseline HR-HPV mRNA was not frequently detected in the pharynx (3.9 % [N = 5/127]), or in seminal secretion (7.1 % [N = 7/98]). None of the 5 HR-HPV positive samples collected from the pharyngeal mucosa were positive for 16 or 18/45 genotypes. Presence of HR-HPV in semen and pharynx was not associated with anal HR-HPV and only 2 individuals had detectable HR-HPV at multiple concurrent mucosal sites. Including all analyzed longitudinal anal swabs for all individuals over 48 weeks of follow-up we found that 85/127 subjects (66.9 %) presented detectable anal HR-HPV and 12/131 (9.2 %) presented detectable pharyngeal HR-HPV (semen samples were only taken at baseline).At baseline, 84 64.1 %) had detectable HHV DNA in their seminal plasma including: EBV (27.5 %), HSV-1/2 (2.3 %), CMV (51.9 %), HHV-6 (6.9 %), HHV-7 (8.4 %), HHV-8 (3.1 %)  . Among participants with detectable anal HR-HPV (any genotype) there was no statistical difference for genotypes 16 and 18/45 between EBV shedders and non- shedders. Also, having detectable seminal EBV was not associated with significantly increased HR-HPV infection in pharynx , and in semen  compared to undetectable EBV. Notably, the increased risk of HR-HPV infection associated with EBV presented a similar effect size in semen compared to the anal but this was not statistically significant . None of the other factors (except for detectable EBV DNA and lower CD4+ T cells count) was associated with increased HR-HPV mRNA detection. The association of EBV shedding with anal HR-HPV had an adjusted odd ratio of 3.99 (1.62-9.81) when including CD4+ count in the model , except week 48 where sample size was reduced due to loss to follow-up (P = 0.15). Among participants with detectable anal HR-HPV at baseline we found increased persistence of HR-HPV in the EBV shedding group over 48 weeks of follow-up .Overall prevalence of HR-HPV infection did not change over the course of 48 weeks of follow up  when including CD4+ count in the model (see Table We found that presence of seminal EBV shedding at baseline was the strongest factor associated with detectable baseline HR-HPV infection of anal mucosa. The only other factor associated with detectable HR-HPV mRNA in the univariate analysis was lower CD4 3.99 1.6–9.81 wheDifferently from previous reports, no association was observed between presence of HR-HPV infection and number of sexual partners, unprotected receptive sex or use of illegal drugs , 18.Among participants with detectable HR-HPV at baseline, we observed that EBV shedding was associated with significantly reduced likelihood of HR-HPV clearing during the following 48 weeks. This is in line with a previous report suggesting that EBV might be associated with higher risk of HR-HPV integration into the human genome , which iThis exploratory study has several limitations. Despite the strong association, our study only included a limited number of HIV-infected men and these results should be confirmed as part of a larger prospective cohort study including both HIV infected and uninfected individuals. Also, the observed association between EBV shedding and HR-HPV infection might be confounded by an unmeasured factor. In particular, there could be a common HIV-related immune defect that could make individuals susceptible to both EBV reactivation and HR-HPV infection. However, it is important to note, that none of the other measured HHV was associated with presence of detectable HR-HPV. Also, the effect of EBV shedding at baseline did diminish over time and by 48 weeks the prevalence of HR-HPV at the anal site was no longer statistically significant between groups. Since EBV seminal replication is intermittent, future studies should collect longitudinal samples to investigate if this observed effect is sustained over time. We also recognize that, since HR-HPV mRNA was detected in anal mucosa, local EBV shedding in seminal plasma may not be directly associated with increased HR-HPV acquisition but be a proxy of EBV replication in the surrounding ano-genital area. Future studies should measure EBV DNA in several anatomic locations to support our observation.In summary, we found that EBV shedding was associated with almost double prevalence of anal HR-HPV infection compared to non EBV-shedding. Further investigation is needed to confirm this observation and explore the causal pathways of how these two viruses might interact by evading immune responses and whether this interaction extends to carcinogenesis.


Analysing human papillomavirus (HPV) viral load is important in determining the risk of developing cervical cancer (CC); most knowledge to date regarding HPV viral load and cervical lesions has been related to HPV-16. This study evaluated the association between the viral load of the six most prevalent high-risk viral types in Colombia and cervical intraepithelial neoplasia (CIN) frequency.a) were estimated for evaluating the association between each viral type’s DNA load and the risk of cervical lesions occurring.114 women without CIN and 59 women having CIN confirmed by colposcopy, all of them positive by conventional PCR for HPV infection in the initial screening, were included in the study. Samples were tested for six high-risk HPV types to determine viral copy number by real-time PCR. Crude and adjusted odds ratios  compared to those having high HPV-16 load . High viral load in HPV-31 patients was associated with lower CIN frequency .The highest viral loads were identified for HPV-33 in CIN patients and for HPV-31 in patients without lesions ; 9.41 HPV copies, 2.58 IQR). Lesions were more frequent in HPV-16 patients having a low viral load (3.53 ORAn association between HPV DNA load and CIN frequency was seen to be type-specific and may have depended on the duration of infection. This analysis has provided information for understanding the effect of HPV DNA load on cervical lesion development. The main factor for developing cervical cancer (CC) lies in persistent infection by at least one viral type of high-risk human papillomavirus (HR-HPV). Fifteen types of HR-HPV have been described, 99.7% being associated with cases of CC and/or cervical intraepithelial neoplasia (CIN) -3. HowevResearchers have thus become interested in HPV viral load. Its association with infection duration has already been described ,7. PriorAs HPV-16 is the viral type most associated with cases of CC 50%–70%) ,5, most 0%–70% ,5The real-time polymerase chain reaction (RT-PCR) has been widely used and described in detecting and typing HPV, as well as quantifying a broad range of viral copies and normalising viral load according to the amount of human DNA, having high reproducibility, sensitivity, specificity and yield ,17. It wThe present study was thus aimed at using RT-PCR for determining the association between HPV viral load and the presence of CIN for six HR-HPV types, which have been previously reported as having the greatest prevalence in Colombia . It was Women eligible for the present study were voluntarily attending their cervical screening consultations between April 2007 and March 2010 in three Colombian regions . Bogotá  has the highest percentage of inhabitants, being mainly an urban population. Girardot is a city located in the Cundinamarca department which has focused its economy on the tourist sector due to its climate and infrastructure. The city of Chaparral (Tolima department) was included in the study as it is located in Colombia’s coffee-growing region and is also known for ecotourism. Girardot and Chaparral were grouped together in the “other city” category to improve the quality of the present study’s statistical analysis.Homo sapiens hydroxymethylbilane synthase (HMBS) gene (Gene ID: 3145) by RT-PCR and those having an insufficient sample for analysis  Ethics Committee.Genomic DNA from cervical samples  taken from HR-HPV 16, 18, 31, 33, 45 and 58 patients, which had been previously confirmed by conventional PCR  , was extHMBS were synthesised according to a study published by Moberg et al. . Ab. Ab15]. Normalised viral load formulaSample size was calculated using the difference of proportions test for high viral load between women having and without cervical lesions (0.42 and 0.052 respectively) ,30; 0.05Analysis was based on type-specific HPV infection rather than on individual women, taking into account that multiple infection is common in the Colombian population .Categorical variable differences between groups were assessed by Chi-squared test or Fisher’s test, as appropriate, using a 0.05 significance level. Median and interquartile ranges (IQR) were used for quantitative variables, according to the data distribution.U test or Kruskal Wallis test, depending on the number of groups to be compared. Both absolute HPV DNA load and normalised HPV DNA load were analysed. Absolute viral load was categorised according to percentile distribution in both groups of patients as follows: negative ≤ 0, low 0 < VL ≤105 HPV copies and high >105 HPV copies .HPV DNA load distribution between women according to colposcopy and biopsy results was analysed by the Mann–Whitney Considering that women with CIN were paired with women without CIN by age and date of entering the study, conditional logistic regression was used for assessing the association between the HPV DNA load for each viral type and cervical lesion frequency according to colposcopy results. This analysis was not done taking the presence of biopsy-defined cervical lesions as outcome, as histology results were not available for all patients included in the study. Crude odds ratio (OR) and adjusted OR with their 95% confidence intervals (CI) were estimated, taking control variables into account, such as origin, ethnicity, age on starting to have sexual relations and the number of infecting HPV types. Hypothesis testing involved a two-tailed test (0.05 significance); STATA 10 was used for all statistical analysis.HMBS gene could not be amplified. This meant that 114 women were classified as negative for intraepithelial lesions  and 59 women having CIN identified by colposcopy (56 women having CIN 1 and 3 having CIN 2+) were included in the analysis  of the women had confirmation of CIN 1 by biopsy (only one woman with CIN 2+ was found). Two of the CIN 2+ women detected by colposcopy had CIN 1 by biopsy.Regarding women with CIN, median age was 40 years old (14 years IQR) and 41.5 years old (13 years IQR) in women without CIN. Most women participating in the study came from the city of Girardot ; 76.19% (n = 80) of these women were negative for lesions. 95.95% of the women in the study were mestizos (n = 166) and the remaining percentage (4.05%) was made up of indigenous, white and black women. The distribution of socio-demographic characteristics and risk factors associated with CC and the detection of HPV infection was compared between both groups (those with CIN and those without it), significant differences being found regarding origin (p < 0.05) Table .Table 2Overall, 91.91% (n = 159) of the sample proved positive for the detection of HPV by RT-PCR, i.e. 93.22% (n = 55) of women with CIN (92.86% positive from the group having CIN 1 and 100% positive from the group having CIN 2+) and 91.23% (n = 104) of women without lesions. 79.24% (n = 126) of all infected women were infected by more than one viral type; this was observed in 81.82% (n = 45) of women with CIN and 77.88% (n = 81) of women negative for lesions. Simultaneous infection was more frequent concerning 2 high-risk viral types in women without lesions  and 3 types in women with cervical lesions . The most frequently encountered viral types were HPV-18 and HPV-16 in multiple infections, in both groups.The type-specific distribution revealed HPV-18 as being most frequent in both groups (69.49% in women having CIN and 66.66% in women without CIN), followed by HPV-16 (57.63%) and HPV-45 (38.98%) in women having lesions and HPV-16 (45.61%), HPV-31 (45.61%) and HPV-45 (38.60%) in women proving negative for lesions. HPV-33 had the lowest infection frequency in both groups.Higher high viral load was recorded concerning HPV-18, HPV-16 and HPV-33 infection in women with CIN, whilst high viral load was most frequent in HPV-31, HPV-45 and HPV-58 infection in women without lesions (Table 8 HPV copies). The highest absolute viral load was detected for HPV-31 in women with CIN (1022 HPV copies) and highest normalised viral load for HPV-33 in women without CIN. No statistically significant differences were observed regarding viral load distribution  for each HR-HPV type in either group of patients.Figures The three patients having CIN 2+ were positive for HR-HPV; HPV-18 and HPV-31 were detected in two of them, whilst the other one was positive for HPV-18, HPV-16 and HPV-45. Even though women having CIN 2+ had a higher viral load  than women having CIN 1, the differences in viral load distribution were not statistically significant. However, normalised viral load for HPV-31 was greater in women negative for cervical lesion and having CIN 1 compared to women having CIN 2+ .The distribution of viral load was also analysed for each HR-HPV type, according to biopsy result. Similar results were found to those with colposcopy ; and for some types  higher normalised viral loads; however, the differences were not statistically significant due to the amount of women analysed Table .Table 4a, 1.16–10.74 95%CI; 2.63 ORa, 1.09–6.36 95%CI, respectively). It was also found that CIN frequency was lower in women having HPV-31 and high viral load . No significant associations were obtained for the other viral types with the presence of CIN (Table Crude and adjusted odds ratios (OR) were calculated for estimating the magnitude of absolute viral load association with CIN for each viral type. The conditional logistic regression model revealed that HPV-16 infection was significantly associated with greater frequency regarding cervical lesions. However, lesions occurred more frequently in the group of women having low viral load for HPV-16 (0 < VL ≤ 5.86 HPV copies) than in women having a high load (>5.86 HPV copies), (3.53 OR22 HPV copies) this provided the best approach for this study [This study involved using RT-PCR; this enabled type-specific evaluation of the viral load of the most frequently occurring oncogenic types in Colombia   for deteis study ,16,18,33More HPV infections were found in women having CIN in our sample, amongst whom all women having CIN 2+ were HPV positive. The foregoing was consistent with the fact that almost 99.7% of CC cases are associated with HPV . PreviouMultiple infection frequency has been variable (16.3%–55%) in previous reports concerning women having lesions ; up to 3HPV-18 and HPV-16 occurred most frequently in the present study, followed by HPV-45 and HPV-58. Differences concerning type-specific prevalence have been reported according to geographic and demographic factors ,35. It iAbsolute viral load was highest in women having CIN compared to women without lesions determined by both colposcopy and biopsy; an increase in the viral load was observed for HPV-18 and HPV-33 proportional to the degree of injury. The foregoing was consistent with previous studies which have revealed the effect of viral load on developing CC. Most HPV-16 studies have found that viral load has increased in relation to the degree of cervical lesion severity ,15,16,42et al., [cf 20 copies/cell; p = 0.0003). It has been found recently that low viral load was characteristic of intermittently detected persistent infection [An association between viral load and cervical lesion frequency (as assessed by colposcopy) was observed in this study just for HPV-16 and HPV-31. The present study’s results highlighted the fact that women having low HPV-16 load (<5.86 HPV copies) had higher cervical lesion frequency. Such results agreed with those from a study by Manawapat, Stubenrauch et al.,  which shnfection . Reducednfection ,45-47. Anfection .Contrary to our findings regarding HPV-16 viral load, the present study found that a high HPV-31 load (>5.14 HPV copies) was associated with lower cervical lesion frequency. As mentioned previously regarding HPV-16 results, it has been shown that viral load has been greater in transitory infections regarding patients having persistent infection . This aget al., found that high HPV-16, HPV-31 and HPV-18/45 viral load increased the risk of developing carcinoma in situ (CIS) [Regarding the other viral types , no association was found between viral load and cervical lesion frequency; such result was supported by data from other authors ,42,49,50tu (CIS) .6 copies/million cells threshold, this being highly specific for grade 2 diagnosis [6 copies as cut-off point for ensuring analysis quality.The pertinent literature gives different cut-off points when categorising viral load, depending on the quantification technique used )  and distiagnosis . Taking It is worth stressing that this technique managed to detect a broad range of viral load, even after stratifying by colposcopy result and viral type. However, this hampered establishing viral load cut-off points to enable identifying women at greater risk of developing cervical lesions; previous studies have also experienced such difficulty ,16,33.This work’s value lies in it being a study where a reproducible, sensitive and specific technique (i.e. RT- PCR) was used for detecting and quantifying viral load  not just for one viral type but for the 6 most frequently occurring high-risk HPV types described to date in Colombia. Besides, this is the first study carried out in Colombia which has included women from regions having high HPV infection prevalence and which was aimed at evaluating the association between HPV viral load and cervical lesion frequency.This study’s results were obtained from a single evaluation of HPV viral load; this means that predicting the risk of lesion progression and developing CC later on cannot be ascertained from this. However, it can be stated that our results were consistent with some findings reported in longitudinal studies ,43,44,48A significant association was found in this study, low HPV-16 and high HPV-31 viral loads were associated with higher CIN frequency; this might have been related to infection duration and immune system response. HPV infection’s effect on developing CC is influenced by viral load, meaning that measuring load could improve the predictive value of HPV detection; however, the scope of quantification depends on the viral type being detected. These findings support the idea of quantifying viral load (as a type-specific marker of CC), coupled to cytology, for improving and strengthening CC screening programmes. This would lead to identifying HPV positive women at greater risk of developing cervical lesions, as well as identifying women as yet lacking cervical anomalies for predicting the beginnings of neoplasia.


We aimed to assess any factors associated with dysplasia regression and with HPV clearance in a cohort of HIV+ patients, with particular focus on cART and gender.Asymptomatic HIV+ patients of the San Paolo Infectious Disease (SPID) cohort who underwent anoscopy/gynaecological evaluation were enrolled. Anal/cervical brushing were analyzed for: HPV-PCR detection/genotyping (HR-HPV), cytologic abnormalities (Bethesda System 2001: LSIL-HSIL). Demographics and HIV-related parameters were evaluated at baseline. Activated CD8+/CD38+ lymphocytes were measured (flow citometry). Patients were examined at baseline (T0) and at 12–18 months visit (T1). HPV clearance was defined as negativisation of HPV at T1; SIL regression (SIL-R) and progression (SIL-P) were defined as change from HSIL/LSIL to a lower-grade/absence of dysplasia and as change from absence of HSIL/LSIL to a higher-grade dysplasia at T1, respectively. Mann Whitney test, Chi-square test and multivariate logistic regression were used.A total of 189 patients were examined, 60 (32%) were women. One hundred fifty patients (79%) were HPV+, 113 (75%) harboured HR-HPV; 103 (68%) showed LSIL/HSIL at T0 (32% of women and 65% of men) . No differences in demographics and HIV-related markers were found between patients with SIL-P  and patients with SIL-R . HPV+ patients who cleared HPV  were found to be more frequently female, heterosexual infected, more frequently on cART and with lower Log10 HIV-RNA and lower levels of CD8+/CD38+ % compared with HPV persistence group .No differences in PI exposure were found between the two groups (p=.08). Interestingly, also when only HR-HPV were considered, clearance was associated with exposure to cART . In multivariate analysis, heterosexuals  were independently associated to HPV clearance, whereas CD8+/CD38+%  were predictive of HPV persistence.Close follow-up of HPV and SIL should be promoted particularly in men and in untreated individuals. We cannot exclude behavioural variables linked to risky sex and reinfection.


The human papillomavirus (HPV) family comprises more than 170 different types that preferentially infect the mucosa of the genitals, upper-respiratory tract, or the skin. The ‘high-risk HPV type’, a sub-group of mucosal HPVs, is the cause of approximately 5% of all human cancers, which corresponds to one-third of all virus-induced tumours. Within the high-risk group, HPV16 is the most oncogenic type, being responsible for approximatively 50% of all worldwide cervical cancers. Many studies suggest that, in addition to the high-risk mucosal HPV types, certain cutaneous HPVs also have a role in the development of non-melanoma skin cancer (NMSC).Functional studies on the HPV early gene products showed that E6 and E7 play a key role in carcinogenesis. These two proteins use multiple mechanisms to evade host immune surveillance, allowing viral persistence, and to deregulate cell cycle and apoptosis control, thus facilitating the accumulation of DNA damage and ultimately cellular transformation.The demonstration that high-risk HPV types are the etiological agents of cervical cancer allowed the implementation in the clinical routine of novel screening strategies for cervical lesions, as well as the development of a very efficient prophylactic vaccine. Because of these remarkable achievements, there is no doubt that in the coming decades we will witness a dramatic reduction of cervical cancer incidence worldwide. Papillomaviridae family that display a distinct tropism for mucosal or cutaneous squamous epithelia. Until now, more than 170 HPV types have been isolated and characterized [The human papillomaviruses (HPV) consist of a heterogeneous group of capsid-enclosed dsDNA viruses from the Based on the homologous nucleotide sequence of the major capsid protein L1, a HPV phylogenetic tree has been generated that groups the different HPV types in genera . The alpWhile the involvement of HPV in causing benign warts was already known, the first evidence of association between human cancer and certain HPV types was proposed thirty years ago by zur Hausen and colleagues . SubsequEpidermodysplasia verruciformis (EV) [The first indication for the oncogenic potential of beta-HPV types came from their isolation in the skin of patients suffering from a rare genetic disorder called mis (EV) , 19. EV mis (EV) . Throughmis (EV) . HoweverDetermination of beta-HPV viral load by quantitative PCR revealed that only a minority of skin cancer cells contained viral DNA , 22. A pIn the last three decades, cervical cancer incidence has been constantly decreasing in high-resource countries, because of the introduction of national cervical cancer screening programmes. In contrast, in low-resource countries, because of the lack of screening programmes, cervical cancer still remains a serious health problem being the first or second cancer in women . The conBecause of this constrain of the HPV detection assays, the interpretation of HPV-positive data require a particular attention and further analyses. It is now clear that additional markers need to be considered to improve the positive predictive value of the HPV detection assays. Many studies suggest that analysis of viral load , 41, HPVINK4a (p16) has been described as a surrogate biomarker of HPV-induced cellular transformation [et al, in this issue).Recently, over-expressed p16ormation . A furthRemarkably, earlier this year a landscape of cervical carcinomas genomic alterations has revealed previously unknown somatic mutations, suggesting that novel therapeutic strategies and/or biomarkers may soon be considered using these as indicators in the pathogenesis and/or treatment of this disease .The genome of all HPV types consists of a circular double stranded DNA filament of about 8 kb, and encodes approximately eight genes. According to protein expression during the viral cycle two functional genome regions have been identified: (i) a coding region containing the early genes, E1, E2, E4, E5, E6, and E7 and (ii) a region containing two late genes, the major (L1) and minor (L2) capsid proteins. In addition, the HPV genome has a non-coding region, termed long control region (LCR), which includes most of the regulatory elements involved in viral DNA replication and transcription . HPV E6 The majority of sexually active women are infected by mucosal HPV types during their lifetime. Most of these infections remain asymptomatic and are eliminated by the immune system in 6–18 months. Only in a minority of women, HPV infection persists and after a period of latency evolve into low and/or high-grade CINs, which may still regress or progress to an invasive cervical carcinoma . NotablyBiological studies have been mainly focused on HPV16 and HPV18, since they are the most frequent types detected in cervical cancer worldwide. These studies have clearly demonstrated that E5, E6, and E7 are directly involved in promoting cellular transformation and altering pathways related to the immune response, as well as cellular transformation , 56 by tThe most characterised activity of HPV16 E6 is its ability to degrade the tumour suppressor protein p53 via the proteasome pathway. The p53 is a transcription factor that is activated in response to stress or DNA damage and positively regulates the expression of genes involved in the control of cell cycle arrest or apoptosis. E6 interacts with a 100 kDa cellular protein, E6AP (E6 associated protein), which functions as an ubiquitin protein ligase (E3). The E6/E6AP complex then binds p53, which becomes very rapidly ubiquitinated and, as a consequence, is targeted to proteasomes for degradation. Since the major role of p53 is to safeguard the integrity of the genome by inducing cell cycle arrest or apoptosis, cells expressing HPV16 E6 show chromosomal instability, which greatly increases the probability that HPV-infected cells will evolve towards malignancy. So far, a vast number of E6 and E7 cellular targets have been identified , 58. ManSince 2006 two prophylactic HPV vaccines, Gardasil and Cervarix, are commercially available. They are based on virus-like particles (VLPs) assembled from recombinant main capsid protein L1 produced in eukaryotic systems . Both vaSeveral clinical trials have evaluated the safety and effectiveness of these two products during a follow-up period of seven to eight years. A recent clinical study aiming at directly comparing the two vaccines has clearly demonstrated that so far both products showed high efficacy in preventing the development of cervical pre-malignant lesions, inducing a robust response in immunised individuals and providing protection for at least five years , 61, 62.Although the introduction into the market of this first generation of HPV vaccines represents an important clinical goal in cancer prevention, several additional aspects need to be solved or improved to optimise vaccine efficacy.HPV-types specificity represents one of the major limitations. Indeed, vaccine efficacy is addressed only to the selected HPV types without any remarkable cross-protection against additional oncogenic HPV types. Hence, both Gardasil and Cervarix do not cover 20–30% of HPV associated cancers. Another important aspect is the high cost of these vaccines, which prevent their widespread application in the vast majority of human population from low and middle resources where cervical cancer is highly prevalent. Modifications of the VLPs assembly chain based on less expensive packaging protocols are currently under evaluation to reduce production costs. In addition, ongoing studies are currently focused on further improving vaccine immunogenicity testing novel antigenic components, such as adjuvants and immune system boosters. This will contribute to obtain a longer-life protection from viral infection and to reduce the number of shots needed for a complete protection . A recenBoth L1 vaccines have no therapeutic effect in already infected women. Since in the majority of pre-malignant and malignant cervical lesions, E6 and E7 are the only HPV genes expressed, most therapeutic vaccines are focused on the stimulation of an immune-response against these two viral proteins . RecentlThe demonstration by epidemiological and biological studies that high-risk HPV infection is associated with human carcinogenesis represents a very important milestone in cancer research. The establishment of such an association led to the development of efficient preventive strategies that has had, and will continue to have a profound impact on public health and overall wealth by reducing the incidence of pre-malignant and malignant cervical lesions. In addition, biological studies on HPV proteins did not only clarify the molecular mechanisms of HPV proteins, but also contributed to our understanding of the fundamental cellular pathways involved in the life of a normal cell.


Colorectal cancer (CRC) is a major burden of public health and healthcare worldwide. Microbiota has been suggested in promoting chronic inflammation in the intestine which, in turn, promotes tumor development. This study focuses on possible correlations of human papillomavirus (HPV) infection with proinflammatory Stat3 signaling activities and the resulting levels of its downstream proinflammatory cytokine IL-17 in CRC patients.HPV was examined using HPV Genotyping Chip technology and constitutively active Stat3 (p-Stat3) and IL-17 levels were tested using immunohistochemistry (IHC) in paraffin-embedded cancerous and adjacent normal tissues (ANT) from a cohort of 95 CRC patients. Correlation analyses were performed between HPV infection and clinicopathological characteristics, Stat3 activities and IL-17 levels among these CRC patients.Three major findings were observed: (1) HPV infection existed in a high rate of CRC cases , of which 45 cases  were high-risk HPV16-positive and only one case was HPV53-positive. (2) HPV infection correlated with poorer clinical stages (III+IV) of CRC. (3) HPV infection strongly correlated with both constitutively higher Stat3 activities (P<0.01) and higher IL-17 levels (P<0.01) only in CRC tissues but not in ANT tissues.HPV infection is common in CRC patients suggesting potentially preventive effectiveness of HPV vaccination among high-risk young individuals. We have for the first time revealed a tri-lateral relationship among HPV infection, constitutive Stat3 activity and IL-17 level, whose collaborative act may orchestrate a proinflammatory microenvironment in the colorectum that, in turn, may promote carcinogenesis and possibly facilitate progression of CRC. Colorectal cancer (CRC) is the third most common cancer globally following lung cancer and breast cancer, with incidence, mortality and 5-year prevalence accounting for 9.7%, 8.5% and 10.9%, respectively, among all cancers according to GLOBOCAN 2012 .The current paradigm is that most solid tumors including CRC are linked to chronic inflammation, directly or indirectly . Among m+ T cells under the induction of TGF-β. Recent studies have suggested that IL-17 plays a dual role in serving either as a promoter or antitumor factor depending upon cancer models [Due to various inflammatory cells and cytokines present in tumor microenvironment, tumors have been referred to as “wounds that do not heal” . Signal r models . In CRC,r models ,13,14. Cr models .Bacterial infections have been shown to collaborate with Stat3 signaling pathway to induce CRC . It is nThis study was approved by the Institutional Ethics Review Board (IERB) at our First Affiliated Hospital of Shihezi University School of Medicine (IERB No. SHZ2010LL02). The IERB waived the need of patients’ consent due to anonymous analyses of the data and standard university hospital guidelines in accordance with the Declaration of Helsinki including confidentiality and anonymity were followed in the handling and publication of patients’ tissues.A total of 235 surgically resected and paraffin-embedded archival human tissues was obtained, including 95 CRC tissues and 90 paired ANT tissues, and 50 colorectal adenoma (CRA) tissues from the Department of Pathology, the First Affiliated Hospital of Shihezi University School of Medicine dated from 2005 to 2007. All selected tissues were based on histopathological diagnoses and reconfirmed for diagnoses before experimentation. These patients were originally selected because of both their available clinical information and sufficient tissue size for experimental analyses (including repeats). CRC patients were aged from 33 to 85 years (with a median age of 66 years) including 43 cases of colon cancer (with 41 paired ANTs available) and 52 cases of rectal cancer (with 49 paired ANTs available). The 5 CRC patients without paired ANTs remained in the study in order to gain statistical power in clinical analyses with several sub-groupings. These CRC patients did not receive preoperative chemotherapy or chemo-radiotherapy and their clinicopathological characteristics were summarized.Due to consumption of tissues in prior studies and required large size of tissues for DNA extraction and subsequent HPV testing, only 50 paired ANT tissues were available that satisfied the selection criterion of 5 cm away from the edge of cancer. For DNA extraction and HPV testing, therefore, we were only able to test these available 50 paired ANT tissues which were randomly distributed among paired CRC tissues. In addition, we also tested 50 unrelated CRA tissues, a common non-cancerous disease of the digestive system, for HPV infection to investigate whether there was any difference between CRC and CRA, of which CRA tissues served as a different control group in terms of HPV infection.http://www.yanengbio.com/en/home2.asp) for HPV testing. The quality and integrity of extracted DNA was tested by PCR using β-actin gene as an internal control (forward primer: 5’-CTTAGTTGCGTTACACCCTT-3’ and reverse primer: 5’- TGTCACCTTCACCGTTCC-3’). A 155 bp PCR product of β-actin gene was detected by electrophoresis on 2% agarose gel.Genomic DNA was extracted from tissues using a DNA extraction kit following manufacturer’s protocols . DNA specimens were then coded without identification of cancer or non-cancerous diagnoses and sent to a specialty DNA genotyping laboratory  Single-use disposables were employed in DNA extraction and subsequent HPV testing; (2) From the first batch of extracted DNA and HPV test results, we selected one negative HPV control (an ANT sample with large size of tissue) and one positive HPV control (a CRC sample with large size of tissue) to be included in every batch of DNA extraction (20 samples per batch); (3) In each batch of DNA extraction, equal numbers of CRC tissues (10 per batch) and ANT or CRA tissues (10 per batch) were included for DNA extraction; and (4) These control DNAs were tested for HPV together with the same batch of DNA extracts. Throughout the entire DNA extraction and HPV testing, we noticed two batches of DNA extracts among which the positive control did not show positivity for HPV and these two batches of 40 tissues were then re-extracted for DNA and re-tested for HPV until the positive HPV control was positive for HPV test. However, we did not observe the negative HPV control turned to be positive through DNA extraction and HPV testing.HPV genotyping was performed using HPV Genotyping Kit, a clinical diagnostic product of Yaneng Bioscience Co.  in the Specialty HPV Genotyping Laboratory within Yaneng Bioscience Co. which had been chosen by the International Agency for Research on Cancer  to perform the Prevalence Surveys of HPV Infection and Cervical Neoplasia in China. PCR reaction mixture (25 μl) contained 5 μl of genomic DNA, 20 μl of PCR reaction buffer. Cycling conditions were as follows: Initial denaturation at 95°C for 5 min, then 40 cycles at 94°C for 30 s, 42°C for 90 s, 72°C for 30 s, and a final extension at 72°C for 5 min. A positive HPV control DNA was placed on each hybridization blot . The revParaffin-embedded tissues were sectioned in 4-μm slices and IHC analyses were carried out using streptavidin peroxidase method . Briefly, tissue slides were dewaxed and endogenous peroxidase was blocked by 3% hydrogen peroxide. After blocking with normal goat serum, primary rabbit anti-human polyclonal antibodies against IL-17  and p-Stat3  were applied onto the tissue slides following the procedures as described previously .Positive IHC stains were defined as yellow-brown color according to the manufacturer’s demonstrative slides . IHC sta2 test and t-test were adopted for variance analysis and correlation was analyzed using Spearman rank correlation method. Differences with P<0.05 were considered statistically significant.Statistical analyses were performed using the SPSS statistical software package (version 17.0). χThroughout the entire DNA extraction and HPV testing, we noticed two batches of DNA extracts among which the positive control did not show positivity for HPV and these two batches of 40 tissues were then re-extracted for DNA and re-tested for HPV until the positive HPV control was positive for HPV test. However, we did not observe the negative HPV control turned to be positive through DNA extraction and HPV testing, suggesting an appropriate management of cross-contamination during HPV testing. HPV infection was detected in 46/95 of CRC cases  which was much higher than ANT  and colorectal adenomas , respectively . Among 42 = 5.9, P = 0.015). Among these 50 paired cases, 6 cases were HPV16-positive in both CRC and ANT tissues ; 14 cases were HPV-negative in both CRC and ANT tissues ; 21 cases were HPV-positive in CRC but HPV-negative in ANT tissues ; and the remaining 9 cases were HPV16-positive in ANT tissues but HPV16–negative in CRC tissues . In terms of HPV infection, no correlation was found between the 50 CRC tissues and their paired ANT tissues . The above observations indicated a fact that HPV infection was a common event in Chinese CRC patients (48.4%), and CRC tissues had a higher frequency of HPV infection than both their paired ANT tissues  as well as unpaired ANT tissues  no low-risk HPV subtypes were detected in all of our tissues tested; (2) among all tissues originated from ANT and CRA, we only detected a single HR-HPV16 subtype . In addiP values . Because clinical stages III+IV were defined as having positive metastasis either to lymph node or to other distant organs, the above correlations suggested a possible role of HPV infection in the progression and metastasis of CRC. Among the 50 paired ANT tissues, we failed to observe any correlations between HPV infection and clinicopathological characteristics as shown in To understand whether HPV infection would affect patients’ clinicopathological characteristics, we investigated possible impacts of HPV infection on a number of clinicopathological characteristics including age, gender, tumor location, histological grading, lymphatic metastasis, clinical staging  among otP<0.001). The differences were particularly obvious for strong active p-Stat3 . For IL-17, on the other hand, this phenomenon (see P<0.001).As shown in enon see  was striP = 0.001). Very similarly, IL-17 levels were also higher in CRC tissues than in ANT tissues . These observations, from a different angle, were supporting the observations as presented in P<0.001) and IL-17 (P<0.001), again, in keeping with the observations as shown in P = 0.07) or IL-17 (P = 0.25).To investigate general differences between CRC and ANT for their Stat3 activities and IL-17 levels, we compared pooled IHC scores from all individuals tested for p-Stat3 and IL-17 using t-test. As shown in It was a puzzling phenomenon that we failed to observe any correlations of either p-Stat3 activities or IL-17 levels with clinicopathological characteristics among these CRC patients including cell differentiation, metastasis, clinical staging among others analyzed . FurtherP<0.01); between HPV infection and IL-17 levels ; and between p-Stat3 and IL-17 levels . However, such correlations among HPV infection, p-Stat3 levels and IL-17 levels were not existent in ANT tissues .To investigate whether HPV infection would play a role in orchestrating a tumor environment with heightened active Stat3 and IL-17 levels, we conducted bivariate correlational analyses. In CRC, as shown in The current paradigm is that the immune system and the microbiota mutually interact to maintain homeostasis in the human intestine. However, components of the microbiota may alter this balance and promote chronic inflammation, further fueling tumor development . Among tHigh-risk HPV (HR-HPV) infection is the major cause for cervical cancer and HPV vaccination is effective in preventing cervical cancer . InteresIn addition, we have also tested HPV infections in other cancers and found quite different frequencies of HPV infection. For example, 10.5% of HPV infection were in breast cancer patients (manuscript submitted); 48.4% in CRC patients (this study); and 90.4% in cervical cancer patients (manuscript in preparation). Our observed frequencies of HPV infection in these cancers are generally in agreement with most published reports, suggesting the quality and reliability of the HPV genotyping method used here.To the best of our knowledge, this is the highest prevalence of HPV infection observed in Chinese CRC, suggestive of potential preventive HPV vaccination  among hiP = 0.028), but not with any other clinicopathological features including tumor sites (colon vs rectum)  . NeverthIndeed, constitutively activated Stat3 (p-Stat3) is significantly increased in CRC tissues than in adjacent normal tissues or ANT , especiaTaken the above together, in a virus-mediated inflammation, Stat3 signaling pathway may also play a crucial and central role as bacteria do in carcinogenesis ,11. FurtP<0.01) and with IL-17 levels . Furthermore, it is absolutely relevant that a correlation also exists between p-Stat3 and IL-17 again in CRC , not in ANT  and there are 23 HPV-specific probes shown. PC is a positive control (filled blue round dot) placed on every chip membrane. (A) Shows a CRC tissue which is positively hybridized for HPV16 subtype (blue round dot in the square 16). (B) Shows a CRC tissue which is negative for all HPV probe hybridizations.(TIF)Click here for additional data file.


Anal human papillomavirus (HPV) infection and its related diseases are relatively common in men who have sex with men (MSM), especially in those HIV positive. In China, molecular epidemiology of anal HPV infection among HIV-positive MSM has been sparsely studied.A cross-sectional study was conducted among HIV-positive MSM in Xi’an, China between April and July 2014. Anal swabs were collected for HPV genotyping.+ T cell level.A total of 195 HIV-positive MSM were included in this study. HPV genotyping showed that 99.0% (191/193) of participants were positive for at least one of the targeted 37 HPV genotypes. 183 (94.8%) of them were infected with multiple high-risk types and 154 (79.8%) of them with low-risk HPV types. HPV 18 was the most frequently identified high-risk type, followed by HPV 16 and HPV 51. As for low-risk types, HPV11, HPV 6 and HPV 81 were most commonly observe. High-risk HPV infection was found to be associated with the status of antiretroviral therapy (ART), the distribution of low-risk types was observed to be varied by CD4Almost all HIV-positive MSM were anal HPV infected in our study. It is highly recommended to consider regular active screening and preventive intervention of HPV infection among this high risk population. Human papillomavirus (HPV) infection is one of the most common sexually transmitted infections worldwide, representing a significant health problem due to its high prevalence and transmissibility . The humOur previously studies reported around 60% HIV negatives and 90% HIV positives were anal HPV infected in MSM from China . To imprThe study was approved by the Ethics Committees of the Institute of Pathogen Biology, Chinese Academy of Medical Sciences & Peking Union Medical College. Written informed consent was obtained from each study participant before the interview and testing.The study was conducted in the Eighth People’s Hospital in Xi’an from April to July 2014. Study participants were recruited through a local nongovernment organization (Xi’an Tongkang Volunteers Workstation). Multiple methods were used for recruitment including website advertisements, distributing flyers with study-related information at MSM frequented venues , and eligible study participants were also encouraged to refer their peers to attend the study. Those eligible participants were HIV-seropositive males, at least 18 years old, ever had homosexual behaviors, willing to provide anal swabs and blood for the test, and physically able and willing to provide written informed consent. Study participants who were tested HPV positive were informed by study personnel confidentially and referred to treatment at the Institute of STD/AIDS Prevention and Treatment, Xi’an District Center for Disease Prevention and Control and the STD/AIDS clinic of Xi’an eighth hospital.+ T cell counts and HPV genotypes were collected for blood test and anal swabs test, respectively.Self-reported socio-demographic characteristics , antiretroviral therapy (ART) status and sexual behaviors in the past 6 months data were collected through one-to-one interviews by the trained interviewer in a separate room using a standardized questionnaire. Each study participant was assigned a unique code that was used to link the questionnaire and specimens. Personal contact information, which was blinded to researchers, was kept by the Xi’an Tongkang Volunteers Workstation for test results feedback and data validation. CD4+ T cell counts (BD FACSCountsystem). Trained personnel collected anal samples by rotating a saline water moistened nylon flocked swab in the anal canal for about 2 minutes. The swab was then kept in 3 mL of sample transport medium for Hybribio HPV DNA Test. Hybribio HPV DNA Test is based on flow-through hybridization to identify HPV types. The denatured DNA was placed into sample wells containing specific probes on the Hybrimem HPV-37 membrane to determine HPV types. The final results were detected by colorimetric change on the membrane under direct visualization. Positive and negative controls were included in the GenoArray test kit in every PCR analysis as well as during the hybridization process for quality. Mixtures of different specific probes can be used in the same well of a 42-well plate format allowing for multiplex analysis. HPV were classified as low-risk (LR) type and high-risk (HR) type according to the kit instruction [Blood samples were collected for testing CD4truction ,13. Hybr+ T cell level  or ART status (started or not), subjects were classified into two categories, respectively. Differences between HPV-types in these variables were assessed with Pearson chi-square test. Frequency distributions are presented for qualitative variables, medians, and inter-quartile ranges (IQRs) are presented for quantitative variables with asymmetrical distributions. The general description of the principal variables including central tendency and dispersion  for the quantitative variables was performed. To identify potential variables related with HPV infection, univariate analysis were performed using Pearson’s chi-square test. All variables with p-values < 0.05 in univariate analysis were entered into the unconditional multiple logistic regression analyses, and the associations were then assessed by means of odds ratios (OR) and 95% confidence intervals. Age, ART status and CD4 level were fixed in the multiple logistic regression models.Questionnaires and laboratory results were double entered and compared with EpiData software . After cleaning, the data was analyzed using SPSS . The characteristics of study population were showed by age, education, marriage status, self-reported sexual orientation, ever had sex with women, age at the first homosexual act, whether anal sex is a regular sex behavior or not. Based on CD4+ T cell counts were available for 193 (99.0%) and 170 (87.2%), respectively. The major characteristics of the study participants are listed in + T cell count was 394 cells/μL . A vast majority (86.2%) of them ever had started ART.A total of 195 eligible HIV-positive MSM were enrolled. HPV genotyping results and current CD4As shown in p = 0.019 and p = 0.044, respectively). ART status, role of sexual behaviors and un-preferred respective sexual behavior were associated with HR-HPV infection , CD4+ T cell level (p = 0.037) and having a fixed sexual partner (p = 0.019) were related with LR-HPV infection in univariate analysis. In multiple logistic regression analysis, as shown in +T cell level was found to be associated with significantly increased rate of LR-HPV infection  and started ART was associated with decreased risk of HR-HPV infection .As shown in + T cell counts (+ T cell level (p = 0.015). As shown in the + T cell, HPV 18, HPV 51, and HPV 16 were the most frequently identified HR-HPV types, but HPV 16, HPV 18, and HPV 33 were found to be the most frequently identified types among subjects with higher CD4+ T cell counts ≥ 394 cells/μL.HPV types distribution stratified by median CD4l counts  showed tWhen stratified by ART status, HPV 11, HPV 6, and HPV 81 were the most frequently identified types among LR-HPV types regardless of ART status. As for HR-HPV types group, HPV 18, HPV 16, and HPV 51 were most frequently detected among those who had started ART. But for those who had not started ART, HPV 18, HPV 52, and HPV 51 were most frequently identified. No significant different distribution of LR-HPV types was found by ART status as shown in + T cell level higher than 394 cells/μL may be more susceptive to infection by LR-HPV than those with less than that and ART might be a protective factor for HR-HPV infection.The present study aims to investigate the HPV genotype distribution among HIV-infected MSM in China. Overall, we found a high prevalence of any-HPV 99.0%), HR-HPV (94.8%), and LR-HPV (79.8%) infection in our study participants. The prevalence of any-HPV in the present study was close to a meta-analysis (96.2%), Lei Gao et al.(96.0%) and Xj Zhang et al. (96.6%) reported 9.0%, HR-,14, but Anal cancer is a relatively rare disease compared to cervical cancer. Its annual incidence is less than 2 in 100,000 persons in the general population worldwide, but is high among well-defined populations such as MSM, particularly HIV-infected MSM, in whom the incidence is up to 80-fold higher than that in the general population ,21. A laThe presence of LR-HPVs, in particular HPV 6 and HPV 11 has been reported for HPV-related invasive anal carcinoma as well ,28. The + T cell level were more likely to be infected by HPV. However, by applying the Bruzzi method [+ T cell counts drop below 200 cells/μL 6–7 years prior to anal cancer diagnosis would prevent 20% of anal cancer cases, this estimate could rise to 49% or 79% by keeping CD4+ T cell counts higher than 350 cells/μL or 500 cells/μL, respectively. A cohort study from Swiss addressing the relationship between CD4+ T cell level and anal cancer risk suggested a rough estimate of the fraction of anal cancer [In our study, we obtained a remarkable finding that HIV-positive MSM with higher CD4i method , it is el cancer . The conl cancer ,41. If tl cancer –43.Several limitations of this study should be kept in mind. Firstly, potential recall bias on sensitive questions could not be excluded completely because our questionnaires were interviewer-administered. Secondly, in terms of difficult sample acquirement and the potential limitation of enrollment methods, our study participants might not represent the general HIV-positive MSM in Xi’an City. Thirdly, due to the relative small sample size, the power of our study for association analysis is limited. Last but not least, cross-sectional study design has its limitation on association analysis. Therefore, our results need confirmation by further large-scale case-control studies or prospective studies.In conclusion, high prevalence of anal HPV infection was observed among HIV-positive MSM in our investigation. Further studies are needed to estimate the risk of HPV related cancers and to explore strategies accordingly for intervention in this population. Developing targeted vaccination based on the most prevalent carcinogenic HPV types should be considered for different population in different areas.


Many Taiwanese women (43.8%) did not participate in regular cervical screening in 2011. An alternative to cervical screening, self-sampling for human papillomavirus (HPV), has been available at no cost under Taiwan’s National Health Insurance since 2010, but the extent and likelihood of HPV self-sampling were unknown.A cross-sectional study was performed to explore determinants of women’s likelihood of HPV self-sampling. Data were collected by questionnaire from a convenience sample of 500 women attending hospital gynecologic clinics in central Taiwan from June to October 2012. Data were analyzed by descriptive statistics, chi-square test, and logistic regression.Of 500 respondents, 297 (59.4%) had heard of HPV; of these 297 women, 69 (23%) had self-sampled for HPV. Among the 297women who had heard of HPV, 234 (78.8%) considered cost a priority for HPV self-sampling. Likelihood of HPV self-sampling was determined by previous Pap testing, high perceived risk of cervical cancer, willingness to self-sample for HPV, high HPV knowledge, and cost as a priority consideration.Outreach efforts to increase the acceptability of self-sampling for HPV testing rates should target women who have had a Pap test, perceive themselves at high risk for cervical cancer, are willing to self-sample for HPV, have a high level of HPV knowledge, and for whom the cost of self-sampling covered by health insurance is a priority.The online version of this article (doi:10.1186/s12905-014-0139-0) contains supplementary material, which is available to authorized users. Cervical cancer is the second most common cancer in women worldwide, with over 529,000 new cases and 275,000 deaths in 2010 -7. A potCervical cancer was the seventh leading cause of cancer mortality among Taiwanese women in 2012 . Taiwan’To explore the likelihood of self-sampling for HPV, we reviewed the literature on factors impacting uptake of HPV vaccination, Pap test, and cervical cancer screening because these factors may also affect uptake of HPV self-sampling. For example, women’s willingness to collect their own HPV samples is crucial  because This cross-sectional study was conducted between June and October, 2012. A convenience sample was recruited from women seeking obstetrical and gynecological health care but not cervical screening at the gynecologic clinics of a regional hospital in central Taiwan. Women were excluded if they reported a history of cervical cancer because these women received different cervical screening recommendations . After tData were collected by questionnaire from June to October 2012. Women respondents were first asked if they had heard of HPV. If a woman responded “yes”, she was asked three HPV-related questions: her perceived risk of cervical cancer (high/low), her priority considerations for HPV self-sampling , and if she had ever self-sampled for HPV (yes/no). If a woman had not heard of HPV, she skipped the first three questions (see “Determinants of the likelihood of self-sampling for HPV” below). All women, regardless of having heard of HPV, were then asked about their willingness to self-sample for HPV  and extent of HPV knowledge. HPV knowledge was measured by 21 true/false items drawn from previous research . PercentDemographic variables included age range , education , annual household income , marital status , and residential area . A related clinical variable was previous Pap testing (yes/no).All the data were entered and analyzed using SPSS version 14.0 for Windows . Data were analyzed by descriptive statistics, the chi-square test, and multivariate logistic regression. All women were asked about their willingness to self-sample for HPV and extent of HPV knowledge because we wanted to determine whether having heard of HPV was associated with willingness to self-sample and HPV knowledge.n = 216, 43.2%). A majority of the women was married , had at least a college education , had an annual household income of < NT$1,000,000 , and lived in a city . The majority of women had had Pap testing  and had heard of HPV . For details, see Table The 500 women who completed our questionnaire were 18–65 years old, with the largest proportion 30–39 years old  and perceived that they had a low risk of cervical cancer . Regarding priority considerations for HPV self-sampling, a majority  was concerned about cost. Two-thirds of these women were willing to self-sample for HPV , and the largest proportion had a moderate level of HPV knowledge . For details, see Table Among the 297 women who had heard of HPV, a majority had had Pap testing  were used as the cutoff for selecting determinants in multivariate logistic regression analysis for the likelihood of women’s self-sampling for HPV.If a woman had not heard of HPV, her data were not included in the analysis of determinants of women’s likelihood to self-sample for HPV because not having heard of HPV indicates unawareness of HPV . AssociaAmong the 297 women who had heard of HPV, 69 (23%) had self-sampled for HPV . This result implies that the odds of a woman self-sampling for HPV were related to these five independent variables, i.e., previous Pap testing, perceived risk of cervical cancer, willingness to self-sample, extent of HPV knowledge, and priority considerations for HPV self-sampling. This model also passed tests of goodness of fit and of collinearity. Regarding goodness of fit, the final model passed the Hosmer and Lemeshow test and was not significant (p = 0.945). Each variable also passed tests for collinearity, with all tolerance scores >0.95, which exceeded the suggested criterion of 0.1 [2) and 32.9% (Nagelkerke R2) of variance in the likelihood of self-sampling for HPV.The results of multivariate logistic regression analyses are presented as factors predicting the likelihood of self-sampling for HPV Table . The resn of 0.1 , and alln of 0.1 . This moThe odds ratios in Table Another strong determinant of women’s likelihood of self-sampling for HPV was priority considerations for HPV self-sampling Table . Women wOther determinants that significantly contributed to the model were extent of HPV knowledge, willingness to self-sample, and perceived risk of cervical cancer Table . Women wThis study contributes to knowledge about the factors related to the likelihood of a clinical sample of Taiwanese women to self-sample for HPV. The majority of our women respondents (59.4%) had heard of HPV, and most of these women (97.3%) understood that HPV causes cervical cancer. However, the remaining 40.6% of our sample had not heard of HPV and did not understand its link to cervical cancer. Although a majority of women had heard of HPV and had moderate-to-high levels of HPV knowledge, they misunderstood important information about the virus, such as condom use only partially protects against transmission of HPV and HPV cannot be transmitted by exchange of bodily fluids . Our results echo those of a study with US college students, most of whom had heard of HPV and had moderate HPV knowledge but did not understand important HPV facts related to cervical cancer . In addin = 331) and those who had heard of HPV (n = 297), suggesting that some women who had a Pap test had not heard of HPV. This may have been due to their doctors not explaining that HPV has been linked to the cytological changes that the Pap test detects or the women not understanding or remembering their doctor’s explanation.Women’s likelihood of self-sampling for HPV was most strongly predicted by previous Pap testing, consistent with the strongest determinant of intent to undergo future Pap smear screening being prior screening . This reAnother strong determinant of women’s likelihood of self-sampling for HPV was cost priority consideration. Cost is a highly negative predictor of self-sampling for HPV. This finding is similar to other studies that vaccine’s cost would impede vaccination ,28,29.Three other determinants of women’s likelihood of self-sampling for HPV were extent of HPV knowledge, willingness to self-sample for HPV and perceived risk of cervical cancer. Similar to a previous report that Turkish women’s acceptance of Kato’s device as an alternative to the Pap smear was related to knowledge of HPV and cancer . This reOur findings are consistent with previous findings that willingness to self-sample for HPV was a determinant of future HPV self-sampling . WillingThe results of our study are subject to some limitations. First, our results are based on data from a gynecological clinic sample recruited from central Taiwan, and only 69 respondents had ever self-sampled for HPV. Thus, the results may not be generalizable to all women in Taiwan. Second, not having heard of HPV does not mean that a woman has no opinion of her risk of cervical cancer or did not self-sample for HPV. Further research should ask women about their perceived risk of cervical cancer and whether they have ever self-sampled for HPV, regardless of whether they have heard of HPV. Third, determinants of the likelihood of HPV self-sampling were examined only among women who had heard of HPV. More research is needed to find effective methods to disseminate information about HPV to increase the proportion of women aware of HPV and to improve their understanding of the link between HPV and cervical cancer. More accurate and complete knowledge about HPV may increase cervical screening rates among women who do not regularly participate in screening by increasing their willingness to self-sample for HPV as reported for native women in Canada .Taiwanese women’s likelihood of HPV self-sampling was predicted by having had a previous Pap test, perceiving a high risk of cervical cancer, willingness to self-sample for HPV, a high level of HPV knowledge, and a cost priority consideration. Therefore, outreach efforts to increase the acceptability of self-sampling for HPV should disseminate information about HPV to increase the rate of women who have heard of HPV and target women who not only have had a Pap test, but also perceive themselves at high risk for cervical cancer, are willing to self-sample for HPV, and have a high level of HPV knowledge. Another important concern for HPV self-sampling is that its cost is covered by health insurance. These findings can be used by policy makers to plan appropriate activities and strategies to educate women about HPV self-sampling for cervical screening, as well as to target women who are likely to accept HPV self-sampling, and in turn to increase regular cervical screening participation in Taiwan.