Is there variation in the diagnostic test completion rate and in the time to diagnostic workup among veterans younger than 50 years with iron-deficiency anemia (IDA) and/or hematochezia?In this cohort study of veterans with IDA and/or hematochezia, diagnostic testing rates were low. Diagnostic testing was less likely among female, Black, and Hispanic veterans with IDA and Hispanic veterans with hematochezia.This study suggests that optimizing timely follow-up may help improve early age–onset colorectal cancer–related outcomes and reduce sex-based and race and ethnicity–based disparities. To date, the diagnostic test completion rate and the time to diagnostic endoscopy or colonoscopy among adults with iron-deficiency anemia (IDA) and/or hematochezia have not been well characterized.To evaluate the diagnostic test completion rate and the time to diagnostic testing among veterans younger than 50 years with IDA and/or hematochezia.This cohort study was conducted within the Veterans Health Administration between October 1, 1999, and December 31, 2019, among US veterans aged 18 to 49 years from 2 separate cohorts: those with a diagnosis of IDA (n = 59 169) and those with a diagnosis of hematochezia (n = 189 185). Statistical analysis was conducted from August 2021 to August 2023.Diagnostic testing factors included age, sex, race and ethnicity, Veterans Health Administration geographic region, and hemoglobin test value (IDA cohort only).Primary outcomes of diagnostic testing were (1) bidirectional endoscopy after diagnosis of IDA and (2) colonoscopy or sigmoidoscopy after diagnosis of hematochezia. The association between diagnostic testing factors and diagnostic test completion was examined using Poisson models.There were 59 169 veterans with a diagnosis of IDA , 189 185 veterans with a diagnosis of hematochezia , and 2287 veterans with IDA and hematochezia . The cumulative 2-year diagnostic workup completion rate was 22%  among veterans with IDA and 40%  among veterans with hematochezia. Veterans with IDA were mostly aged 40 to 49 years (37 719 [63.7%]) and disproportionately Black (24 480 [41.4%]). Women with IDA  had a lower likelihood of diagnostic test completion compared with men with IDA. Black  and Hispanic  veterans with IDA were less likely to receive diagnostic testing compared with White veterans with IDA. Veterans with hematochezia were mostly White (105 341 [55.7%]). Among veterans with hematochezia, those aged 30 to 49 years were more likely to receive diagnostic testing than adults younger than 30 years of age . Hispanic veterans with hematochezia were less likely to receive diagnostic testing compared with White veterans with hematochezia .In the cohorts of veterans younger than 50 years with IDA and/or hematochezia, the diagnostic test completion rate was low. Follow-up was less likely among female, Black, and Hispanic veterans with IDA and Hispanic veterans with hematochezia. Optimizing timely follow-up across social and demographic groups may contribute to improving colorectal cancer outcomes and mitigate disparities. This cohort study evaluates the rate of diagnostic endoscopy or colonoscopy and the time to testing among veterans younger than 50 years with iron-deficiency anemia (IDA) and hematochezia. Accordingly, our goal was to examine the receipt of guideline-concordant workup for IDA and hematochezia and the factors associated with any diagnostic workup for adults younger than 50 years with IDA and/or hematochezia.Published guidelines recommend that all men and postmenopausal women with IDA undergo bidirectional endoscopy—both esophagogastroduodenoscopy and colonoscopy.21 To identify study data, we used several Department of Veterans Affairs (VA) electronic health record data resources, including the VA Corporate Data Warehouse, the VHA Vital Status file, and the National Death Index. We used separate IDA and hematochezia cohorts based on a prior study that examined these CRC symptoms separately.14 We excluded veterans with EAOCRC or inflammatory bowel disease diagnoses prior to diagnosis of IDA or hematochezia. Reporting on study design, analyses, and results followed the guidelines outlined by the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) reporting guideline and statement specific to cohort studies. The study was approved by the Veterans Affairs San Diego and University of California San Diego institutional review boards and was granted a waiver of informed consent by both institutional review board committees because the data are retrospective from a previously defined cohort with data collected under waivers of consent and Health Insurance Portability and Accountability Act authorization.We conducted a retrospective cohort study among US veterans aged 18 to 49 years receiving care between October 1, 1999, and December 31, 2019, in the Veterans Health Administration (VHA).14 Iron-deficiency anemia was identified by laboratory diagnosis using World Health Organization criteria: a hemoglobin test identifying anemia  with a follow-up iron test within 3 months indicating iron deficiency .18 Veterans with menorrhagia or hysterectomy prior to or within 30 days of diagnosis of IDA were also excluded. In addition, we excluded veterans based on any International Classification of Diseases, Ninth Revision (ICD-9) and International Statistical Classification of Diseases and Related Health Problems, Tenth Revision (ICD-10) diagnosis codes for IDA prior to the date of hemoglobin blood test.The IDA analytic cohort included veterans aged 18 to 49 years with a diagnosis of IDA, derived from a previously derived cohort of veterans who had at least 1 blood test measuring hemoglobin level conducted within the VHA.Current Procedural Terminology code for an office visit initiating care within the VHA.14 Hematochezia was identified by ICD-9 codes (569.3 and 578.1) or ICD-10 codes (K62.5 and K92.1). We excluded veterans based on any diagnosis of hematochezia prior to the date of first VHA office visit.The hematochezia analytic cohort included veterans aged 18 to 49 years with a diagnosis of hematochezia, derived from a larger cohort of adults who had at least 1 We also developed a cohort of adults with both IDA and hematochezia as a secondary analysis to examine factors associated with diagnostic workup when individuals had both diagnoses. We defined joint exposure of IDA and hematochezia as diagnoses within 60 days of each other.Our primary outcome was the time to diagnostic testing. For veterans with IDA, diagnostic test completion was defined as bidirectional endoscopy. To account for potential variation in practice where only the first endoscopic test performed would yield diagnostic resolution, we counted the first date of esophagogastroduodenoscopy or colonoscopy completion. For veterans with hematochezia, completed diagnostic testing was defined as either colonoscopy or sigmoidoscopy. In the secondary analysis of veterans with both IDA and hematochezia, diagnostic test completion was measured as completion of sigmoidoscopy or colonoscopy.22 and hemoglobin test value (measured in grams per deciliter) in the IDA cohort only. Inclusion of race and ethnicity as a variable in our study was to measure whether there were differences in time to diagnostic testing across race and ethnic groups.Candidate factors for timely diagnostic testing evaluated included age, measured continuously and categorically , sex, patient-reported race and ethnicity , US Census–defined region of VHA care ,P values were from 2-sided tests and results were deemed statistically significant at P < .05. We used R, version 4.0.2  to perform our analyses.23Statistical analysis was conducted from August 2021 to August 2023. We conducted survival analyses to examine the association between candidate factors and time to diagnostic testing, including measurement of cumulative diagnostic test completion rate at 60 days, 180 days, and 2 years, based on a Kaplan-Meier estimation. Additional estimates of the cumulative diagnostic test completion rate were calculated by age group, sex, race and ethnicity, and census-defined geographic region. Follow-up was defined as the time from IDA and/or hematochezia diagnosis to 1 of the following: (1) endoscopy workup (outcome), (2) death, (3) end of 2 years of follow-up (730 days), or (4) December 31, 2019. The choice to end follow-up at a maximum of 2 years was based on the theory that diagnostic testing after this time would no longer correspond to the diagnosis of IDA and/or hematochezia. The proportional hazards assumption was tested by examining the correlation between time and scaled Schoenfeld residuals for all variables. Because the proportional hazards assumption was violated in all models, Poisson regression models were used to calculate rate ratios (RRs) and corresponding 95% CIs. Sensitivity analyses were performed excluding sigmoidoscopy as a recommended diagnostic examination for veterans with both IDA and hematochezia. All Of 3 728 118 veterans aged 18 to 49 years with at least 1 encounter in the VHA health system between 1999 and 2019, 59 169 had an IDA diagnosis , 189 185 had a hematochezia diagnosis , and 2287 had both an IDA and hematochezia diagnosis  . The oveOf the 59 169 veterans with IDA, 37 719 (63.7%) were aged 40 to 49 years, 28 667 (48.4%) were women, 24 480 (41.4%) were classified as Black, and 4161 (7.0%) were classified as Hispanic . The estAmong the 189 185 veterans with hematochezia, most were aged 40 to 49 years (106 730 [56.4%]) and men (163 690 [86.5%]) and were composed of 105 341 White (55.7%), 44 939 Black (23.8%), and 17 137 Hispanic (9.1%) veterans . The estThere were 2287 adults with a diagnosis of both IDA and hematochezia . VeteranAmong veterans aged 18 to 49 years with a diagnosis of IDA and/or hematochezia, the cumulative proportion receiving guideline-recommended diagnostic testing after 2 years was low, at 22% for those with IDA and 40% for those with hematochezia. Furthermore, receipt of diagnostic testing within 60 days of diagnosis of IDA or hematochezia occurred for only 7% of those with IDA and 22% of those with hematochezia. After IDA diagnosis, men were more likely to receive diagnostic testing compared with women, and the likelihood of workup increased with age. After diagnosis of hematochezia, adults aged 30 to 49 years were more likely to receive diagnostic testing compared with adults younger than 30 years. Black veterans were less likely to receive diagnostic testing after an IDA diagnosis, and Hispanic veterans were less likely to receive diagnostic testing after an IDA or hematochezia diagnosis, compared with White veterans. Given that both IDA and hematochezia have been shown to increase EAOCRC risk, our findings suggest that there are significant opportunities to improve EAOCRC outcomes, including sex-based and race and ethnicity–based disparities, by promoting diagnostic testing after IDA and hematochezia diagnosis.24 Although the risk of incident and fatal EAOCRC is lower among women than men,26 the markedly lower likelihood of diagnostic testing is surprising, given that the population with a diagnosis of IDA had a nearly equal amount of men and women. Iron-deficiency anemia is commonly attributed to menorrhagia in premenopausal women, but our study found the disparity in workup persisted even after removal of women with diagnosed cases of menorrhagia or prior hysterectomy. It is plausible that clinicians are more likely to attribute iron deficiency in women to disorders of menstruation. Nevertheless, more research is necessary to uncover why this sex-based disparity in diagnostic follow-up exists and whether substantially lower rates of endoscopic follow-up for women are clinically appropriate.Men were more likely than women to receive diagnostic testing after an IDA diagnosis, which aligns with prior research.2 Recent evidence indicates that, despite greater increases in EAOCRC incidence among White adults compared with Black adults over a 15-year period, White adults had higher relative survival.27 Differences between Black and White adults have been postulated to be associated with differences in risk factor burden, access to health care, and follow-up patterns, such as for abnormal results from stool tests performed for CRC screening, as well as structural racism. Our findings of variation in diagnostic testing after IDA and hematochezia diagnosis suggest that there may be specific clinical scenarios amenable for interventions that can reduce disparities in CRC outcomes for Black vs White adults.Our study found that Black veterans with IDA were less likely to receive diagnostic testing than White veterans. Black veterans of all ages have been shown to have higher CRC incidence and mortality and a more advanced stage of CRC at presentation.29 In addition, Hispanic adults have similar or worse EAOCRC-related survival compared with White adults.31 Our study findings indicate that effective strategies for timely diagnostic testing may help address these differences.Hispanic veterans with IDA and/or hematochezia were also found to have a lower likelihood of receiving diagnostic testing compared with White veterans. Recent evidence has shown that EAOCRC incidence is increasing rapidly among Hispanic adults, particularly for regional or distant-stage disease.33 Similarly, Hispanic veterans were found to have higher rates of diagnostic colonoscopy completion after abnormal stool blood test results compared with non-Hispanic adults.34 These findings in diagnostic follow-up after abnormal stool blood test results contradict the diagnostic testing findings in our study, in which Black veterans and Hispanic veterans had a lower likelihood of receiving diagnostic testing. Other studies in community health settings have found no significant association between race and ethnicity and diagnostic colonoscopy workup for adults with positive stool blood test results.35 Although these studies highlight the variation in findings regarding race and ethnicity–specific follow-up patterns, there is still limited evidence focusing on follow-up after potential signs or symptoms for CRC.29To date, there are few data on race and ethnicity–specific diagnostic follow-up patterns for CRC-related conditions, particularly for adults with IDA and/or hematochezia, to our knowledge. Prior studies that examined colonoscopy follow-up after positive stool blood test results found that Black veterans were more likely to receive follow-up than White veterans, including 1 study among adults aged 45 to 50 years.37 It is also plausible that racial and ethnic differences could be due to variations in adherence to clinician follow-up recommendations potentially in response to experiences of structural racism in health care. Explicit and implicit bias by health care professionals and health care systems in the assessment of and management recommendations for Black or Hispanic adults vs White adults with IDA and/or hematochezia have been documented in the literature.40 Nevertheless, our results suggest that more research is necessary to understand whether race and ethnicity may be associated with the response of clinicians or the health care system to these diagnoses. Reasons for differences in diagnostic follow-up for IDA and hematochezia, as well as interventions to address these differences, require further study and could help to address racial and ethnic disparities in CRC outcomes.Our study was conducted within the VHA, where all participants had no barriers associated with insurance coverage, yet disparities still were found. The observed disparities may be associated with individual , interpersonal , organizational, community, or societal influences.Our study has some limitations. Given the limitations in capturing community health data, we were unable to ascertain endoscopy uptake claims outside of the VHA, which could underestimate diagnostic testing completion rates. Nonetheless, we do not expect these ascertainment issues to explain the between-group variation in testing rates. In addition, our study was unable to examine specific individual-, clinician-, or system-related factors, such as distance to care and interpersonal bias, that may contribute to the observed findings. Future research should focus on whether these factors are associated with disparities in diagnostic test completion.Our study also has some strengths. One key strength was that the absolute numbers of women in our IDA (n = 28 667) and hematochezia (n = 25 495) cohorts were substantial, and, to our knowledge, they represent one of the largest observations of diagnostic testing outcomes among women younger than 50 years presenting with IDA or hematochezia.This cohort study found that the rates of diagnostic testing with endoscopy among veterans aged 18 to 49 years with IDA and/or hematochezia are low and vary significantly across demographic groups. Black veterans with IDA and Hispanic veterans with IDA and/or hematochezia were less likely than White veterans to receive diagnostic testing for EAOCRC. Furthermore, women with IDA were less likely than men to receive diagnostic testing. Optimizing diagnostic test completion among individuals with IDA and/or hematochezia may help improve early detection of EAOCRC and contribute to reducing EAOCRC-related disparities.


Blue emissions in organic light-emitting diodes (OLEDs) are essential for their application in solid-state lighting and full-colour flat panel displays. On the other hand, high-power blue emitters are still uncommon, especially those that can achieve the Commission Internationale de l’Eclairage  coordinates of  in the National Television System Committee (NTSC) blue standard and have high external quantum efficiencies (EQE) of more than 5% because their molecular design presents an enormous challenge. Therefore, creating effective, stable, pure, and deep blue fluorescent materials is vital. Here, it is addressed how useful blue fluorescent Zn (II) complexes are for making organic light-emitting diodes (OLEDs). Utilizing Zn (II) complexes is appealing because of their favourable luminous characteristics, acceptance and mobility, and affordability. This mini-review article aims to provide an overview of Zn (II) complexes that emit blue fluorescent light and have been reported since 2018, while highlighting the unique qualities that make them appropriate OLED materials. Metal complexes with small organic ligands have attracted a lot of attention in organic light-emitting diode (OLED) applications because of their exceptional electroluminescent properties ,7,8,9,102), zinc nitrate (Zn (NO3)2), zinc acetate hydrate (Zn (CH3CO2)2·2H2O), zinc acetate Zn (OAc)2, and zinc nitrate hexahydrate (Zn (NO3)2·6H2O are common sources of metal ions used in the coordination chemistry of Zn (II) complexes imidazole, and the delocalized π-π* transition from the 2-substituted phenolate to the phenanthroimidazole ring, respectively  and film state (447 nm) at room temperature with quantum yields 68% (in solution) and 21% (thin film). The authors further reported that the preparation of the ZNL16 complex with a configuration of ITO|HATCN|TAPC|TCTA|ZnL16|TmPyPB|LiF|Al provides good blue emission in an undoped device with a maximum current efficiency of 3.79 cd/A at a low input voltage of 3 V, a maximum power efficiency of 3.97 lm/W, and a maximum brightness of 1153 cd/m2, with an external quantum efficiency of 2.47% at CIE coordinates  of 0.17 and 0.18. The data obtained recommended ZnL16 as a good blue emitter and host material phenyl)-3-(9-ethyl-9H-carbazol-3-yl)propan-1,3-dione L17 with zinc acetate dihydrate (THF) using redistilled triethylamine (Et3N) in tetrahydrofuran solvent /LiF(1 nm)|Al configuration utilizing three distinct guest-doping concentrations of 1 weight (wt)%, 3 wt%, and 5 wt% of ZnL17 complex. The 3% doping device demonstrated deep-blue electroluminescence and attained the greatest electroluminescent performances among them. At maximum voltage of 8 V, this deep-blue device produced a CIE value of , which is quite near to the NTSC standard blue value   see . The 3 wZnL18 and ZnL19) with pyrazolone-based azomethine ligands and used them for OLED applications. The reaction of the ligands 5-methyl-4-methyl)-2-(p-tolyl)-2,4-dihydro-3H-pyrazol-3-one (L18) and 5-methyl-4-((4-Butoxyphenyl)imino)methyl)-2-(p-tolyl)-2,4-dihydro-3H-pyrazol-3-one (L19) with zinc acetate dihydrate in an ethanol solution followed by recrystallization from ethanol: methylene chloride (2:1) provided tetradentate Zn (II) complexes ZnL18 and ZnL19 in overall yields of 70 and 80%, respectively . Both complexes were reported to have effective thermal stability at high decomposition temperatures greater than 350 °C, and both complexes were amorphous in nature with no glass transition temperature, as measured by TGA and DSC. The UV–Vis absorption and photoluminescence properties of ZnL18 and ZnL19 were reported both in DMF solution and in the solid state at room temperature. The redox potential of Zn (II) complexes measured by the cyclic voltammetry analysis was reported to have HOMO energy values of −5.54 and −5.46 eV and LUMO energy levels of −2.73 and −2.60 eV. The UV–Vis absorption of ZnL18 and ZnL19 in solutions was found to emit a blue emission at 424 and 432 nm, respectively, while in the solid state, a red shift emission at 466 and 468 nm for ZnL18 and ZnL19 was observed  complexes were further processed in solution with doped and undoped devices that constructed light-emitting diodes in a multilayer architecture with device configurations of ITO|PEDOT:PSS| coordinates were recorded for a ZnL19 complex-based device. The deep blue emission exhibited by the ZnL19 complex is aligned with the NTSC colour coordinates, meeting the high colour saturation requirements in display technology.In 2022, Solanki et al.  synthesiectively . Only thrved see . The qualm/W see . On the lm/w see . The CIEZnL20, which is indispensable in OLEDs. Applications of the ZnL20 complex, such as anti-counterfeiting and latent finger printing (LFP), have also been described due to the strong cyan-blue luminescence emission with high quantum efficiency. The ZnL17 complex was reported to be synthesized as a white crystalline solid from the reaction of the bisimidazolyl phenol (L20) ligand with zinc acetate using methanol as a solvent . According to DFT calculations, the first absorption maximum occurs at about 344 nm, which corresponds to the S0-S1 transition  technology. It was reported that LFPs can be better visualized using ZnL20 under UV light as the standard reference material, indicating that the ZnL20 produced is considered to be an excellent means of visualizing LFPs. The authors demonstrated the utilization of Zn-BDI as an emissive material to construct efficient, high-luminance blue OLEDs.In 2021, Kempegowda and co-workers  reported solvent . The comtion see . The ass3.8% see . It was OLEDs have made remarkable strides in the last decade, driven by the massive flat panel and solid-state lighting markets. Recent research efforts in OLED materials have focused on developing efficient, stable, and pure blue emitters to capture the much higher performance of lower-energy colours. Researchers have been able to overcome hurdles in modifying the structure and electronic properties of Zn (II) complexes, which has had a substantial impact on their photophysical properties thanks to the discovery of new synthetic pathways. The creation of efficient and resilient blue emitters for OLED applications is made possible by careful ligand selection. The perfect candidates for Zn (II) complexes for commercial applications should show high stability in devices with more than 90% PLQYs and CIE coordinate values  conforming to the values of , corresponding to the Internationale de l’Eclairage in the National Standard Blue Standard of the Television System Committee (NTSC). Such ideal Zn (II) complex candidates have been described in this review, and one of them has been reported by Solanki and co-workers . In addiIn the future, understanding the relationship between structure, electrical properties, and other photophysical aspects will be crucial for the development of next-generation Zn (II) complexes for OLEDs. In addition, the standardization of photophysical data would help to understand the complex systems already described.


Human papillomavirus (HPV) is recognized as being related to a wide variety of known cancers: cervical, oropharyngeal, anal, vaginal, penile, and skin. For some of these cancers, rigorous algorithms for screening, therapeutical interventions, and follow-up procedures have been established. Vaccination using the nonvalent anti-HPV vaccine, which prevents infection regarding the most frequently involved high-risk HPV types  and low-risk HPV types (6 and 11), has also extensively prevented, controlled, and even eradicated HPV infections. Still, even with all of these multidisciplinary interventions, the burden of HPV cancers is still high worldwide. The circulating DNA of HPV-induced cancers is thought to be an adequate biomarker for optimizing the control of these virus-related cancers. We analyzed the literature published in the last 5 years regarding ctDNA and four of the above-mentioned cancers. The most frequently used assay for ctDNA detection was the droplet digital PCR assay, used for the management of therapy in the late stages of cancer. ctDNA could not be used for early detection in any of the studied cancers. The OPSCCs were the most frequent cancers analyzed via ctDNA assays. Larger, properly designed cohort studies might establish the clinical utility of this biomarker. The burden of cancer cases attributable to HPV infection is very high, as this infection causes almost 5% of all new cancer cases worldwide. Annually, in women, HPV infections cause 530,000 cancer cases located in the cervix, 18,000 in the anus, 8500 in the vulva, 12,000 in the vagina, and 5500 in the oropharynx. In men, HPV infections cause 17,000 cases of anal cancer, 13,000 cases of penile cancer, and 24,000 cases of oropharyngeal cancer annually [The natural history of HPV-related cancers remains partially understood, with cervical cancer being the most analyzed HPV-related cancer. Recently, in 2022, the International Agency for Research on Cancer (IARC) published a handbook on cervical cancer screening, which presents and summarizes the published evidence on screening methods and practice through narrative and systematic reviews, as well as quality assessments. Additionally, it describes the natural history of cervical cancer, from a normal cervix to HPV infection, the progression to pre-cancerous squamous lesions, and an invasion leading to squamous cell carcinoma (SCC) . The majThe relative importance of carcinogenic human papillomavirus (HPV) types has been established by IARC groups; HPV 16 is uniquely carcinogenic, with a 62.4% attributable etiological fraction (the percentage of cancer caused by HPV 16), followed by HPV 18 and HPV 45, which are relatively important for cancers, especially adenocarcinomas when compared to precancers, with attributable etiological fractions of 15.3% and 4.8%, respectively. For clinical use, commercial HPV screening assays often detect a pool of carcinogenic (or high-risk) HPV types; the 14 types most commonly included in current HPV tests are HPV 16, HPV 18, HPV 45, HPV 33, HPV 58, HPV 31, HPV 52, HPV 35, HPV 59, HPV 39, HPV 68, HPV 51, and HPV 56 [The natural history of HPV, from oral HPV infection to HPV-related oropharyngeal cancer, is not as well understood as cervical cancer, with some data remaining unknown. There are, however, accepted risk factors: oral sex, being a male, and being a current smoker; these are factors that favor the transition from HPV exposure to oral HPV infection. Regarding persistence risk factors, we can add older age and immunosuppression to those previously mentioned. The signs of HPV can appear within 1–2 years, and progression to oropharyngeal cancer can take up to several decades, although no reliable precursors or progression risks have been identified yet. There is a great need in the field for large natural history studies in diverse populations to characterize the steps from the acquisition of oral HPV infection to its progression into HPV-positive oropharyngeal cancer .The natural history of HPV in men became more well-known after the HIM study of Giuliano AR et al. was published in The Lancet . We founSkin cancer is thought to be etiologically connected to cutaneous beta HPV types, especially non-melanoma skin cancer. By using transgenic mice, Viarisio D et al. demonstrated that beta HPV 38 E6 and E7 oncoproteins act as promoter and progression factors in multi-stage skin carcinogenesis . A revieThe above data underline the high burden of HPV-related infections and cancers and the fact that these represent an important health problem worldwide. Screening and vaccination are important interventions that could lead to a major reduction in HPV-related diseases. Given the fact that so many anatomically different regions can be affected, it is difficult to obtain an ideal assay to screen for all the above-mentioned cancers. Still, in recent years, many research groups worldwide have tried to analyze the suitability of ctDNA detection techniques for different HPV-related cancers.In 2010, Pantel K et al. presented a clinical application: circulating tumor cells in the peripheral blood and disseminated tumor cells. The authors of this paper mentioned the possibility of detecting these markers with ultrasensitive methods for the prognosis of cancer patients, monitoring systemic anticancer therapies, and identifying therapeutic targets . Since tAim: in this narrative review, we aimed to investigate the clinical utility of ctDNA assays for HPV-related oropharyngeal, cervical, anal, and skin cancers.Literature search and study selection: A systematic search of the PubMed and the EMBASE databases was carried out for all the published studies in the last 5 (2018–2023) years, using the following search algorithm: circulating DNA HPV AND cervical cancer, circulating DNA HPV AND oropharyngeal cancer, circulating DNA HPV AND anal cancer, and circulating DNA HPV AND skin cancer. Our systematic analysis identified 55 articles for oropharyngeal squamous cancer (OPSCC), 41 for cervical cancer (CC), 15 for anal cancer (AC), and 2 for skin cancer (SC). After a careful analysis of all these articles by two different team members, we excluded the review papers and only analyzed studies that were related to our research topic, giving 17 studies for OPSCC, 6 for CC, 7 for AC, and 1 study for SC . The abon = 15), followed by real-time multiplex PCR (n = 2). Three papers used commercial ddPCR assays , and the other authors used in-house optimized real-time PCR assays. Most studies collected blood and preserved plasma before and after the therapeutic intervention, which was either surgery alone or surgery with radio-chemotherapy . A total of 12 of these studies were performed in the USA, two in Japan, and the rest in Europe. The main assay used for ctDNA HPV evaluation was digital droplet PCR (ddPCR)  in its first prospective clinical trials, showing that quantitative cell-free HPV-DNA (cfHPV-DNA) levels could help to identify patient candidates for therapy de-escalation in HPV-related OPSCC . The secThe authors of the studies mentioned in We identified six studies that evaluated ctDNA in cervical carcinoma (CC). Three of these studies were performed in Europe, one in Canada, and one in Thailand. Five studies used ddPCR assays for ctDNA HPV evaluation, and the research group from IARC used the E7-MPG bead-based HPV genotyping assay, based on Luminex technology. All studies analyzed the blood from patients at different stages of HPV cervical infection .The authors of the below-mentioned studies performed cohort studies, case–control studies, and prospective multi-center studies. The number of CC cases analyzed for ct DNA HPV in these studies was between 23 and 206 cases. The sensitivity of the ctDNA assay was between 30.77 and 100.00%, and specificity was 100% in all the studies below.We identified seven studies that evaluated ctDNA in anal carcinoma (AC). Two of these studies were performed in the USA, one in Brazil, and the rest were carried out in Europe. Five studies used a ddPCR assay for ctDNA HPV evaluation, three used NGS, one used in situ hybridization, and one used flow cytometry. Plasma samples were analyzed in all seven of these papers .The authors of the below-mentioned studies performed cohort studies, case–control studies, and prospective multi-center studies. The number of AC cases analyzed for ct DNA HPV in these studies was between 2 and 288 cases. The sensitivity of the ctDNA assay reached 100% for both sensitivity and specificity in detecting ctDNA in anal cancers.Péré H et al., 2021, presented a very interesting case of a patient who died from HPV 16-induced high-grade anal intraepithelial SCC with vertebral HPV 16-positive metastasis. By using the Capture-HPV method and then next-generation sequencing (NGS), the authors found that HPV 16 was detected episomally in all tested samples: anal biopsy samples, plasma samples, and vertebral metastasis biopsy. The availability of the above-mentioned technologies could be used in future prospective studies to anticipate overall and progression-free survival and to offer patients optimized and personalized treatment .For skin cancer, we identified only one paper published in the past 5 years regarding blood testing for HPV in relation to cutaneous cancer. Hampras SS et al. conducted a skin screening study by collecting blood, eyebrow hair, and skin swabs, which were analyzed for viral DNA using a bead-based multiplex PCR-Luminex assay. The authors found that skin infection with gamma HPV infections could increase the chances of cutaneous malignancy development by recruiting immunosuppressive lymphocytes into the skin .In this study, we have evaluated the clinical utility of ctDNA in HPV-related cancers. We found that the most frequently used assay for this biomarker evaluation was ddPCR, which was both optimized in-house and commercially developed, followed by NGS, real-time PCR, and in situ hybridization assays. This subject is of interest because other authors have tried to analyze the test parameters of the ddPCR assay ,48,49, mThe OPSCC was the most frequent type of HPV-related cancer, which was evaluated by ctDNA HPV in our study approach. One reason could be that, for OPSCC, even though it is known that performing oral sex is a risk factor for oral HPV infection, there are no recognized HPV-induced precancer lesions like there are for CIN2/3 or AIN2/3. Older age, male sex, current smoking, and immunosuppression could be considered risk factors for oral HPV persistence, but there is no recognized precursor from oral HPV persistence to oropharyngeal cancer . Researc+ TIL counts, age, T-stage, and E2 expression) [All the above studies show how many research scenarios have been developed by clinicians in collaboration with laboratory doctors, with the aim of therapy optimization for OPSCC patients. Over the years, researchers have been pre-occupied with finding the best selection of HPV-driven OPSCC cases, for example, the strict algorithm used in the HPV AHEAD studies, which combined DNA HPV testing with mRNA HPV analysis and p16 positivity ,53,54,55ression)  by analyression)  or analyression) ,59. All ression) ,61.The specificity in the OPSCC studies that we analyzed varied between 88.9 and 100%. The detected sensitivity of ctDNA HPV for OPSCC was between 56 and 100%. The lowest sensitivity was identified for 25 OPSCC Asian patients who were p16-positive. The highest sensitivity was reported by the authors who tested American patients using a ddPCR assay. Generally, a single study evaluates over 1000 samples. Some analyzed studies had between 12 and 200 samples evaluated for ct DNA in OPSCC patients. For the period analyzed (2018–2023), the highest number of studies was found for OPSCC. This could be explained by the lack of a classical screening method, which is available for CCs. There is a need for the identification of an optimal biomarker for the early detection and monitoring of OPSCC HPV-related patients.A recent meta-analysis, which included 10 studies with a total of 457 HPV-positive HNSCC, identified a pooled sensitivity and specificity of 0.65 (95% CI) to represent the diagnostic performance of cfHPV-DNA. Campo F et al. concluded that cfHPV-DNA could be used as a biomarker for monitoring treatment response during the clinical trials of de-escalation therapy .Another review supports ctHPV assessment in the early detection of disease progression and at any stage of managing a patient with HPV-related OPSCC .However, the diagnostic value of cfDNA as a biomarker, especially ct HPV DNA in OPSCC, is under clarification. As the vast majority of OPSCC studies are from the USA, ethnicity does seem to play a role, but rather the logistics and the experience of the researchers using ddPCR for ct DNA evaluation do not. Future studies on HPV-induced OPSCC cases should be cost-efficient and follow ethical study designs that could further advance the secondary prevention of HPV-induced OPSCC. It is important to identify pre-cancerous lesions and to optimally manage biomarker-positive patients and HPV-DNA-positive OPSCC patients.The natural history of cervical carcinoma is very well known, starting from the normal cervix to pre-cancerous lesions and leading to carcinoma  . We idenOver the years, the screening of cervical cancer and pre-cancerous lesions has been optimized by very well-known groups of researchers ,66,67,68The specificity in the analyzed CC studies reached 100% in two papers. The sensitivity of ctDNA from CC was low, between 30.77 and 74.7%. Some studies mentioned that they had ”high sensitivity”, without any precise specification. Only two studies tested more than 200 patients. The other studies had a very low number of CC patients tested for ctDNA (between 23–39 patients). One possible explanation for this low number of tested patients is that in developed countries, the CC prevalence has decreased thanks to well-implemented organized screenings. Therefore, it is more time-consuming to perform larger studies. On the other hand, in low-income countries, where the prevalence of CC is high, ctDNA-related technologies from body fluids are still in development and may not be feasible for application yet. The presence of HPV-ctDNA is associated with unfavorable outcomes in cervical neoplasia.A recent meta-analysis identified the low sensitivity (0.36) and high specificity (0.96) of cHPV DNA as being diagnostic for cervical cancer. The authors concluded that along with further optimization of the analyses to achieve higher sensitivity, liquid biopsies are promising diagnostic and prognostic biomarkers for the detection of CC .We should also be aware of the effects and harms of cervical cancer screening, which should be considered in accordance with the principle of evidence-based decision-making, which states that before implementing an intervention , the balance of clinical benefits, harms, and costs should be evaluated .The epidemiology of HPV infection in men can be described by the results of the HIM study conducted by Giuliano AR et al., which found that the age-specific prevalence of any genital HPV infection among men was detected to be high among all age groups: above 50% . Anal caStudies on the clinical application of ctDNA detection for anal cancer had similar designs to the research on OPSCC and cervical cancer. Some authors tried to use this marker to optimally monitor this category of patients, and they expressed the intention to conduct studies on larger cohorts to identify if there is any association between the screening markers of anal cancer and HPV . Other rThe specificity of ct DNA HPV (AC-related) in the analyzed studies varied between 67 and 100%. The sensitivity had the same values but varied among different studies, between 67 and 100%. Lee JY et al., 2020, identified SE = 100%, SP = 100%, PPV = 100%, and NPV = 100% in a study on 33 HPV-positive AC by using a novel NGS assay. None of the analyzed studies reached 100 patients (limits: 2–88 patients) ,44,45,46A recent meta-analysis identified a high sensitivity (0.95) and specificity (1.0) of cHPV DNA for the diagnosis of AC. Additionally, the authors considered that cHPV DNA was useful in the prediction of treatment response or progression-free survival in AC and in post-treatment detection . A revieAs it is currently not known which technique (ddPCR vs. NGS) is more sensitive, larger studies are required, including clinical trials, to assess the clinical utility of ct DNA HPV.The proper diagnosis, treatment, and follow-up of HPV-induced anal cancer is a subject of interest, as there are many guidelines from different research groups ,79,80 thSkin cancer development is initiated by ultraviolet (UV) radiation, together with beta HPV types, which augment UV-induced DNA mutations . Bandoli® RSC ccfDNA Plasma Kit  [® MiniElute® ccfDNA Mini Kit  and diluted it at a ratio of 1:40 according to the manufacturer’s instructions [Extraction methods for cell-free DNA and ctDNA from blood samples could be a logistical challenge for some laboratories. In the case of OPSCC studies, Akashi et al. collected a volume of 10 mL of blood, in addition to samples, during routine medical practice. The blood was processed via centrifugation using specific parameters. The cell-free DNA solution was extracted from 1 mL aliquots of plasma using the MaxwellWI, USA) . Tanaka WI, USA) . Haring ructions .® collection tubes . These samples were stored at room temperature (6–37 °C) for up to 14 days before DNA extraction, as per the manufacturer’s instructions [By using cervical cancer samples, Bønløkke et al. extracted ccfHPV DNA from 30 mL of full-blood samples collected from each participant in Cell-Free DNA BCTructions . Jeannotructions .Lefèvre et al., in their anal cancer study, used a Chemagic 360 robot  and a 1304 cfDNA purification kit (PerkinElmer) to purify DNA from 4 mL of thawed plasma, to which 191 base pair (bp) spike-in DNA was added . AnotherThese selected methods show that there are already different commercially available kits for extracting ctDNA or cfDNA, which could favor, to some degree, the large applicability of this assay.One major limitation of detecting HPV using ctDNA assays is that it cannot be used to monitor disease progression from low- and high-grade lesions and carcinomas, especially in ASCUS , LGSILs , and HGSILs . Moreover, due to the costs of these methods, it is unlikely that HPV ctDNA assays can be used on women from low-income countries. The same limitation can be mentioned for pre-cancerous anal lesions.In this comprehensive analysis of HPV-related cancers, we have summarized the known natural histories for each of these types of cancer and the available guidelines for screening. We identified ddPCR as the most frequently used assay for detecting and quantifying specific, high-risk HPV types. Further clinical optimization of this assay is still needed in terms of sensitivity, specificity, and positive and negative predictive values. HPV-driven OPSCC was the most commonly studied type of cancer using the ddPCR methods.None of the studies used ctDNA for the early detection of any of the analyzed HPV-induced cancers. The most useful clinical applications in the analyzed studies were to assay ctDNA as a biomarker before and after surgical intervention, to de-escalate CRT therapy, and to better optimize surgical interventions. Larger cohort studies are necessary to confirm the clinical utility of ctDNA in HPV-related cancers.


Oropharyngeal cancer, a subset of head and neck cancer, is increasingly recognized as a unique clinical entity primarily influenced by high-risk human papillomavirus (HPV) infections, particularly HPV-16. This review delves into the viral life cycle of HPV-16 and its interactions with host cells, with a specific focus on the crucial roles played by the viral oncoproteins E6 and E7. These oncoproteins drive cellular proliferation by targeting critical tumor suppressor proteins like p53 and Rb, resulting in uncontrolled cell growth and genomic instability. Furthermore, the significance of epigenetic modifications induced by HPV-16 and their implications is important for cancer progression. This comprehensive review provides valuable insights into the intricate molecular landscape of HPV-induced oropharyngeal cancer, shedding light on the development of targeted therapies and preventive strategies for this emerging global health concern.Video AbstractThe online version contains supplementary material available at 10.1186/s12964-023-01365-0. In the domain of head and neck squamous cell carcinoma (SCC), oropharyngeal squamous cell carcinoma (OPSCC), often referred to as throat or tonsil cancer, is distinguished by its involvement of the base and posterior third of the tonsils, tongue, soft palate, as well as the posterior and lateral pharyngeal walls . A troubHPV, as a DNA virus with a diameter of about 55 nm, lacking an envelope is known for its tropism to epithelial cells. It contains a double-stranded DNA molecule, which encompasses about 8,000 base pairs (8 kbp) . The virpapillomaviridiae have been identified, exhibiting a minimum of 10% nucleotide variation within the capsid gene (L1) [Currently, more than 200 different genotypes of ene (L1) . Severalene (L1) , 12. Furene (L1) , 14. On ene (L1) . Severalene (L1) . HPV is ene (L1) . E6 alsoene (L1) –11. Otheene (L1) .Current HPV detection methods have undergone significant advancements, leading to improved accuracy and efficiency. These methods encompass Nucleic Acid Amplification Tests (NAATs), including PCR and digital PCR, recognized as the gold standard, with a focus on quantifying viral load. Hybrid Capture Assays, Next-Generation Sequencing (NGS), and the combination of Pap smear and HPV DNA testing for heightened sensitivity are prominent trends. Additionally, immunohistochemistry identifies viral oncoproteins in tissue samples, while the development of point-of-care and saliva-based tests aims to increase accessibility. Ongoing research explores novel biomarkers such as microRNAs and DNA methylation patterns for enhanced early detection. Quantitative PCR and HPV genotyping, achieved through multiplex PCR assays and DNA microarrays, continue to play a crucial role in assessing risks associated with HPV infection, ultimately contributing to the prevention and early diagnosis of HPV-related cancers , 11, 17.Since the beginning of the twenty-first century, a new type of head and neck cancer, HPV + OPSCC, has emerged . Among aBased on the reports, in sub-Saharan Africa, the incidence of HPV-positive OPSCC seems to be relatively low, with only a limited number of reported cases so far, despite the region having a high prevalence of HPV-associated cervical cancer . In a stIn the past, there has been a robust correlation has existed between head and neck SCCs, including those affecting the oropharynx, and individuals with an extended record of substantial smoking and alcohol intake , 26. PriThe evidence concerning the role of tobacco and alcohol in oral HPV infection and HPV-related OPSCCs remains inconclusive . While cHPV-related OPSCCs are strongly linked to sexual behaviors, which are correlated with the disease . NumerouHPV-positive OPSCCs represent a distinct subgroup that tends to be diagnosed at a younger age, typically several years earlier than HPV-negative tumors . DespiteHPV-related OPSCCs have a strong association with sexual behaviors, which are linked to the disease . MultiplBoth non-HPV-related and HPV-related HNSCCs show a higher incidence in males, with a ratio of about 3 to 1 . In the There have been indications that non-sexual HPV transmission may be feasible through actions like kissing, transmission during childbirth, and transmission during laser surgery , 74. OraEnhancing cancer prevention is crucial to align with shifting societal norms. Presently, no early screening techniques exist for OPSCC, emphasizing the significance of widespread vaccination initiatives for prevention . The effNationwide vaccination initiatives that encompass boys have been expanded in several countries, including Australia, Italy, Germany, Austria, New Zealand, UK, and the USA . DespiteHPVs have genetic material composed of circular double-stranded DNA, which is approximately 8000 base pairs long . Among tThe intricate relationship between HPV-16's productive life cycle and the differentiation of keratinocytes within stratified mucosal epithelia contributes to the process of carcinogenesis , 89. ThiThe field of understanding how HPV interacts with cellular differentiation pathways has recently witnessed significant activity. In the context of HPV-induced carcinogenesis, a pivotal phase is marked by the activation of two viral early genes: HPV oncogenes E6 and E7 , 92. TheThe physical state of the HPV genome holds clinical importance in individuals with HPV + OPSCC . An examFor an extended period, the disruption of Rb function has been acknowledged as a vital oncogenic trait stemming from HPV-induced epigenetic reshaping , 100. ThE6 and E7-mediated rewiring of cell-cycle control suggests that the increase in p16 expression observed in HPV + cancers is a consequence of E7-induced KDM6B, rather than Rb inhibition, which has been widely believed . FurtherThe biological profile of oropharyngeal cancer associated with HPV infection involves the degradation of p53, the inactivation of the Rb pathway, and an increase in p16 expression. On the other hand, tobacco-related oropharyngeal cancer is characterized by mutations in p53 and a decrease in CDKN2A expression. In contrast, HPV-positive oropharyngeal cancer demonstrates enhanced sensitivity to both chemotherapy and radiation therapies when compared to HPV-negative forms of the disease. The detection of HPV-16 can be regarded as a prognostic indicator linked to better overall and disease-free survival outcomes. However, its utility as a predictive marker remains to be firmly established. Numerous inquiries into the natural progression of oral HPV infection are currently subjects of ongoing research. Concerning disease management, it is acknowledged that HPV-positive oropharyngeal cancer represents a unique category within head and neck SCC, characterized by a more favorable prognosis. Individuals diagnosed with HPV-positive oropharyngeal cancer typically belong to a younger age group and generally exhibit better overall health. In forthcoming clinical trials, categorizing patients with head and neck cancer based on their HPV status is advisable. Regardless of the chosen treatment strategy, there exists an opportunity to investigate less aggressive treatment approaches that preserve survival rates while minimizing the potential for severe side effects.


Cervical cancer is preventable with vaccination and early detection and treatment programs. However, for these programs to work as intended, stigma related to HPV and cervical cancer must be understood and addressed. We explored pre-existing stigma associated with HPV and cervical cancer in the public healthcare system and community of a low-resource setting prior to implementation of an HPV screen-and-treat program.This study conducted thematic analysis of data collected during implementation of a novel HPV screen-and-treat system for cervical cancer early detection and treatment in Iquitos, Peru. We included 35 semi-structured interviews , eight focus groups (70 community women), one workshop , 210 counseling observations (with 20 nurse-midwives), and a document review. We used the Socio-Ecological Model to organize the analysis.We identified three main themes: 1. the implication that women are to blame for their HPV infection through characterizations of being easy or promiscuous, 2. the implication that men are to blame for women’s HPV infections through being considered careless or unfaithful, 3. HPV is shameful, embarrassing, and something that should be hidden from others. Consequently, in some cases, women refrained from getting screened for HPV. These themes were seen at the individual level among women, relationship level among women, men, and family members, community level among healthcare staff, and societal level within components of cervical cancer guidelines and male chauvinism.Cervical cancer early detection and treatment programs in limited resource settings must address stigma entrenched throughout the entire healthcare system and community in order to sustainably and successfully implement and scale-up new programs. Interventions to tackle this stigma can incorporate messages about HPV infections and latency to lessen the focus on the influence of sexual behavior on HPV acquisition, and instead, promote screening and treatment as paramount preventative measures. The large-scale elimination of cervical cancer is feasible through the implementation of human papillomavirus (HPV) vaccination for girls, screening eligible women, and providing treatment for those who need it , 2. CervStigmatization of women with HPV is a barrier to cervical cancer screening uptake , 4 and aIn LMICs, rates of cervical cancer exceed those of high-income countries (HICs)  . In the We did not originally intend to explore stigma related to HPV and cervical cancer, but the theme emerged so often among women, healthcare staff, and governmental institutions during Proyecto Precancer’s work and challenged programmatic success that we decided to explore this topic retrospectively. This qualitative study explores the perspectives of various stakeholders within the EDT system, including women, nurse-midwives, doctors, and cancer coordinators on HPV and cervical cancer stigma. The data were collected during Proyecto Precancer's involvement in Iquitos, prior to implementation of the novel screen-and-treat program and include interviews, focus groups, a workshop, counseling observations, and document reviews. Our objective is to understand the existing stigma before introducing the screen-and-treat program by identifying the influence of community members, health professionals, the healthcare system, and healthcare policies in perpetuating HPV and cervical cancer stigma. This research will provide an understanding of stigma in low-resource settings that must be addressed prior to implementation of novel EDT programs to ensure appropriate and sustainable implementation.This study was based in the largest public health network – the Micro Red Iquitos-Sur (MRIS) – in Iquitos, the capital city of the state of Loreto, located in the Peruvian rainforest (population 1.1 million). Iquitos is the largest city in the world that can only be accessed by boat or plane; there are no roads to Iquitos. The MRIS extends over urban, peri-urban, and rural areas, with much heterogeneity with regards to access to tertiary care. Specifically, for those in urban areas, access to a hospital could take 15 minutes on paved roads. However, for those living in peri-urban and rural areas, the main forms of transport are motorcycle taxis and boats for the river communities. The boats and motorcycle taxis frequently traveling on unpaved roads are often slow, meaning that accessing healthcare facilities, specifically tertiary care, can take hours for much of the population. Though the Amazon basin region of Peru is geographically unique from the mountain and coastal regions, this region accounts for the highest cervical cancer mortality and covers two-thirds of Peru’s territory ” (community women's focus group #8). In some cases, having HPV was associated with prostitution. In a focus group, when the interviewer asked what participants thought of women with HPV, one woman responded: “that they are prostitutes” (community women's focus group #2).On the individual and relationship levels, women associated HPV infection with behaviors such as having multiple sexual partners and engaging in what was perceived as irresponsible sexual conduct, particularly among younger or single women. One participant spoke about how younger, sexually active women could get HPV stating, “"You have to know the person you're with, I mean, what kind of person you're messing with" (community women's focus group #3). Other women stated that it is women’s responsibility to not have unprotected sex, even with a steady partner. During a discussion of a scenario where a woman with a partner finds out she has HPV, a participant responded: “We are not going to be trusting because we have a partner; it is better to be safe” (community women's focus group #5).Some women emphasized that it is a woman’s responsibility to know the type of person she has sex with in order to avoid HPV. One woman described this as, Sometimes, we don't know what kind of people we are dealing with" (woman with precancer/cancer's interview #11). In a separate counseling session, another nurse-midwife addressed what they perceived as risky behavior and advised the patient to question her partner’s sexual history asking, “How many partners has your partner had? Who is he?” (nurse-midwife observation #7).On the community level, these messages were similarly conveyed by health professionals. One woman reported she was told by a nurse-midwife during counseling that it is important to know the type of people she has sex with because, as the nurse-midwife told her, “The more sexual partners we have, the more likely we are to have HPV” (nurse-midwife observation #1). Another nurse-midwife listed risk factors for HPV during counseling as: “promiscuity, fewer children, early initiation of sexual activity” (nurse-midwife observation #7). In an interview, a nurse spoke about a patient with precancer as having poor sexual behavior: “We know that she is a young woman, that she is not very well behaved” (nurse interview #1).Health professionals also described women who have had multiple sexual partners as more likely to get HPV. During counseling about HPV, one nurse-midwife referred emphatically to having multiple sexual partners as an important risk factor: “Here, sexual activity starts at the age of 12, 13, 14 and even more [earlier] if we talk about the communities that are [on the] rivers” (nurse-midwife interview #6) and another stating:My concern is the beginning of sexual relations at an earlier age. Here we know that at 13, 14 years old they already have their first child or maybe their first partner. They arrive at 19 with more partners, even with more children. Those women are getting lost, they are going to arrive at 40 already with cancer (workshop #1).A number of health professionals expressed concern that women in Iquitos are more likely to develop cervical cancer at an early age due to early initiation of sexual activity with one nurse-midwife stating, “The discussion of women’s actions leading to their acquisition of HPV was also visible at the societal level in materials published by the Peruvian Ministry of Health. The 2017 National Clinical Practice Guidelines for Cervical Cancer Prevention and Control was created for health personnel to provide evidence-based recommendations for cervical cancer prevention. It specifies the following risk factors associated with HPV progression to cervical cancer: HPV type, genetic susceptibility , immunocompetency , tobacco use, and vitamin deficiency. Notably, a lack of screening is not mentioned as a risk factor for developing cervical cancer. Although not directly related to the development of cervical cancer, the list also mentions risk factors that significantly increase risk of HPV infection such as early initiation of sexual activity, having multiple sexual partners, and being with a partner who has multiple sexual partners .idiosincrasia de la poblacion”), low screening coverage, delay in returning results, high loss to follow-up, low number of treatment centers, and concentration of specialized care in urban settings. Notably, the National Plan does not identify the same risk factors specified in the Clinical Guidelines though it was published only six months later; instead, it only lists one vague risk factor for cervical cancer related to individual sexual behavior   has been promiscuous … I am paying the price" (community women's focus group #4). Another woman described how women can develop cervical cancer due to men’s infidelity: "Sometimes men are not faithful to you. You may think they are, but, in the end, you don't know. You can be healthy, and some time goes by, and you find out that you are sick" (community woman's interview #11). In some cases, these women had experienced infidelity or knew women with unfaithful partners: "My brother had sex with another woman. The woman infected him. And he went to his wife, and he infected her" (community women's focus group #4). Only a couple of women mentioned the possibility of infection through previous relationships: “Perhaps her partner has had other encounters before her” (community women's focus group #8).HPV infection was also commonly viewed as a sign of male infidelity. Women spoke about what their reactions would be if they were told that they had HPV, with one woman stating, “the partners, right?” but after thinking about it a bit more stated, “well, but in the end, she will get it anyway” and emphasized the importance of screening (field notes). During counseling for HPV, a nurse-midwife stated, “Your partner infected you during sexual relations” (nurse-midwife observation #3), and another spoke about when she used a megaphone to announce that “faithfulness” is one of the ways to avoid HPV (nurse-midwife interview #6). A doctor also discussed seeing a book titled Human Papillomavirus: The Virus of Infidelity displayed on a nurse-midwife's desk. The book was written by a nurse-midwife from Iquitos, and the doctor told this nurse-midwife, “You can't put that [book there] because it is not entirely true. You’re going to get in trouble” (doctor interview #2).The implication that men are to blame for HPV infection was also observed at the community level. When asked about the risks and main ways to prevent cervical cancer, one of the cancer authorities quickly answered: “The implication that men are to blame for HPV infection was also observed at the societal level in the 2017 National Clinical Practice Guidelines and 2019 Health Directive for the Prevention of Cervical Cancer which suggested a heightened importance of men’s sexual activity to explain women’s positive screening results. The Clinical Practice Guidelines and Health Directive included being with a partner who has multiple sexual partners and having a partner who has had or has many sexual partners, respectively, as risk factors for progression to cervical cancer. However, the 2017–2021 National Plan does not explicitly include this risk factor related to men’s sexual behavior, and the National Plan and Health Directive both include the absence of screening or inadequate screening first on the list of factors contributing to cervical cancer .Women often experienced the consequences of stigma related to abnormal cervical cancer screening results and HPV. This was observed across the individual and relationship level among community women who discussed how women are told to consider and considered abnormal cervical cancer screening results to be embarrassing, in some cases disgusting, and something that should be hidden from others. This was also seen at the community level among nurse-midwives who described abnormal cervical cancer screening results as embarrassing, and the societal level where male chauvinism resulted in secrecy around HPV. As a consequence of the stigma, women sometimes refrained from getting screened for cervical cancer.ugly” and felt “disgust” (community women's focus group #6). Similarly, women referred to stigma around gynecological symptoms they associated with abnormal cervical cancer screening results: “Sometimes they have discharge and do not want to go to the post [because] of shame,” (community women's focus group #6). Another woman described this as, “Sometimes they feel the symptom and are ashamed to talk, to go to a health center” (woman with precancer/cancer's interview #8).On the individual and relationship levels, women referred to HPV as ““Maybe she doesn´t want someone to find out, so she gets information through the internet” (community women's focus group #5) while another stated, “I think she prefers the internet because… she could suddenly feel shame, I don’t know, to go and to ask about this virus, no?” (community women's focus group #3).Women also expressed perceived feelings of embarrassment to seek information regarding HPV or HPV screening and fears of being perceived as having HPV. During discussions of scenarios about how women with HPV might seek more information, one woman described, that we have cancer” (community women's focus group #6). One of these women stated that she wanted to do this “to keep others from seeing [her drop off the HPV test]” (community women's focus group #6).Women were often reluctant to be seen dropping off HPV samples for fear of being judged. During a focus group, a few women discussed how if they were to do an HPV test, they would prefer to drop off the HPV samples at a secure location outside of the health center so that their neighbors would not think ““Because when they hear the name HPV, they would think that she is having a very liberal sex life, wouldn't they? … because the mother does not know how her daughter behaves in the street” (community women's focus group #3).Women also described perceptions that their families would think differently of them if they found out they were HPV positive. During a discussion of a scenario where a young woman with HPV tells her family about her HPV infection, participants described how the family would react: the first thing she said was that she did not want to be registered in the system because she was a little ashamed” (field notes).This was also seen at the community level among nurse-midwives who support the need for secrecy; according to a nurse-midwife, some colleagues who had positive cervical cancer screening results asked not to be logged on the registry system as they preferred discreet management. Our research team also observed this, with one researcher noting that when a nurse-midwife found out about her positive result, “Maybe your husband sees you are sick. He is not going to want a sick woman who is suffering. Instead of supporting you, what he will do is, “You know you are sick, and I have another one out there.” He will go to the other one. He will abandon you. (community women's focus group #4).Women's desire for secrecy regarding HPV was reinforced at the societal level, largely due to widespread male chauvinism. Women expressed concerns about men's ability to accept information about HPV and provide support if they became ill. During a discussion of scenarios where women with HPV tell their husbands about their HPV infections, one woman described:Maybe because they are afraid of being left,” and another adding, “The fear of feeling rejected” (community women's focus group #3). Another woman described how she felt a husband would react to finding out his wife has HPV: “Suddenly, he would leave you” (community women's focus group #1).During a focus group, other woman discussed why women would be afraid to tell their husbands that they have HPV with one stating, “In order to eliminate cancer, in order to protect so many women who sometimes die because of their negligence, I say, because of their negligence, because they do not take care of themselves… sometimes they feel the symptom and are ashamed to talk, to go to a health center. (woman with precancer/cancer's interview #8).The stigma around HPV was described by some women as a barrier to screening:She said that my father always told her that, why would she go for a check-up, why would she go for a check-up if she has nothing… he was always a little bit macho in that aspect, and my mother never went for a check-up. She says that she was afraid of having a Pap smear done… I asked her, “Why are you scared?” …"I don't know, but your dad never wanted me to do that." (woman with precancer/cancer's interview #11).Male chauvinism was also mentioned as a barrier to getting screened for cervical cancer by women and health professionals. One woman spoke about why her mother had never done a Pap test before she was diagnosed with cervical cancer and died from cervical cancer a few years later:Even in our region there is a lot of male chauvinism. How many times have I visited the homes of families where the husband came out and said, “My wife is not a prostitute, she cannot have a Pap test,” even another one told me - the patient - “Look, I do not do the Pap test because I only have one husband, because they [women with HPV] have a lot of men (workshop #1).A nurse-midwife also discussed stigma related to HPV as a barrier to screening in the workshop:We found pervasive stigma throughout the healthcare system and community that, in some cases, led women to avoid HPV screening. This stigma may, ultimately, facilitate women dying from a preventable cancer, and subtly or overtly, it is shaped by policymakers, experts, healthcare providers and community members who have formed perceptions about who is at risk for and who might develop cervical cancer due to women’s or their partners’ sexual behaviors. Throughout this study, we found that stigma emerged unprompted in approximately half of interviews, in all focus groups, the design workshop, and among a fifth of the nurse-midwives’ providing counseling.Stigma was associated with HPV through the implication that women are to blame for their HPV infection. This implied blame was attributed as either a direct consequence of their actions  or an indirect consequence . On the individual, relationship, and community levels, women, nurses, and nurse-midwives characterized women with HPV as promiscuous or irresponsible while on the societal level healthcare guidelines emphasized the influence of women’s sexual activity on HPV acquisition. Previous literature focusing on women’s perspectives has similarly found the implication that women are to blame for HPV infections due to having multiple sexual partners, promiscuity, prostitution, or a lack of cleanliness , 24, 25.We also found that women attributed HPV infections to men’s actions, through infidelity or carelessness. On the individual, relationship, and community levels, women, nurse-midwives, and cancer authorities characterized men as unfaithful, and on the societal level healthcare guidelines emphasized the influence of men’s sexual activity on women’s HPV acquisition. Previous studies in LMICs have found perceptions that HPV is caused by infidelity, though many of these studies discuss women being perceived as unfaithful , 12, 28;Additionally, we found attitudes of shame toward HPV. This was seen on the individual, relationship, and community levels expressed by women and nurse-midwives. The shame towards HPV, in addition to the fear of being stigmatized as being promiscuous or easy, lead both women and healthcare workers to keep their cervical cancer screening results secret. In previous research, women have described similar attitudes of perceiving HPV as shameful , 26 and Stigma pervades each level of the healthcare system and community, potentially intensifying as it moves through different levels. For instance, the presence of stigma among women may be exacerbated by healthcare professionals who, in turn, may have internalized stigmatized information from policies. It is important to recognize that the information flow is predominantly unidirectional; policymakers wield influence over healthcare workers and, subsequently, women, whereas women may exert limited impact on policymakers. While addressing stigma across all levels is essential, we must prioritize instigating change where it can have the greatest influence, starting with healthcare policies.We have seen that misinterpretation and over-emphasis of less-significant risk factors related to sexual behavior stigmatizes women (regardless of whether they even have any of these risk factors). While early sexual initiation and number of sexual partners have been identified in epidemiological studies as increasing the likelihood of testing positive for HPV or developing cervical cancer , 32, theFor HPV related stigma, this process must begin with developing national plans, directives, and clinical guidelines that consistently provide the same key messages about HPV infection and its latency and promote access to screening and treatment. Messaging on the importance of sexual activity on HPV acquisition in these documents is fallacious. The changes must include developing communication messages and cervical cancer screening forms that do not focus on or collect data on age of sexual initiation and number of male sexual partners. These materials can be developed in a participatory manner with key stakeholders, allowing them to internalize these key messages and feel ownership over new policies , 39. AwaThis process must also directly target healthcare providers who play a critical role in transmitting information about and attitudes toward HPV to women. Training can aim to improve knowledge among healthcare professionals about HPV infections and promoting access to screening, while specific guidelines and tools can be developed for health professionals that help reduce misinterpretation and allow consistency in messaging. A number of interventions using these methods have been shown to reduce HIV stigma among healthcare providers –43 and mReducing stigma among women and community members may also include providing counseling or developing an intervention that focuses on open discussion and improving knowledge at the community and individual level. In places with high male chauvinism or that are very conservative, it might be appropriate to include men in such interventions. In fact, addressing stigma through the lens of also addressing gender equality presents an opportunity for regional authorities in low-income settings to work toward the fifth Sustainable Development Goal while providing women with further autonomy and empowerment in screening and treatment of cervical cancer . Only a During implementation of the HPV screen-and-treat program, Proyecto Precancer provided counseling and training to nurse-midwives that aimed to address stigma and improve the overall care process for women. Future research will need to evaluate the impact of this work and any further interventions implemented to address stigma.This study utilizes a variety of data that were not specifically collected to examine stigma in the MRIS. As a result, it is not possible to determine the prevalence of stigma, and there is a possibility that additional stigma-related themes could emerge with further stigma-focused data collection. However, the themes and data in this paper are reflected in extensive observations made throughout years of Proyecto Precancer's fieldwork in a public health network of a low-income area in Peru and include the perspectives of multiple stakeholders, from community women to hospital-level doctors, that consistently highlight these themes. While the findings may not be generalizable to all countries or the whole country, they provide valuable information on the stigma faced by women in resource-limited settings and provide context for the scale-up and implementation of new EDT programs in these settings.Stigma is prevalent throughout this resource-limited healthcare system and community, with women, healthcare workers, and policy makers contributing to implied blame of women for their HPV infections and implied blame of men through accusations of infidelity. This stigma created feelings of shame and secrecy surrounding HPV that were present among women and nurse-midwives, and, in some cases, led women to avoid HPV screening, and ultimately, can facilitate women dying from this preventable cancer. Cervical cancer EDT programs in resource-limited settings must take into consideration and address the stigma experienced by women at the policy, health system, healthcare provider, community, and individual levels when implementing and scaling-up new EDT programs. Stigma reduction interventions should emphasize information on HPV infection and its latency over and beyond emphasis on the impact of sexual behavior on HPV acquisition to promote screening and treatment as the most important preventative measures.


The recent increase in high‐risk human papillomavirus (HR‐HPV)‐associated oral and oropharyngeal cancers has gained considerable importance due to their distinct clinical and molecular characteristics. However, the natural history of oral HPV from acquisition to persistence and malignant transformation is still unclear. The global prevalence of oral HPV infection in healthy individuals ranges from 0.67% to 35%, while 31%–38.5% in head and neck cancer (HNC). The persistence rate of oral HR‐HPV infection is 5.5% –12.8% globally. India has the highest HNC burden due to apparent differences in predisposing factors compared with the West. The prevalence of oral HPV in healthy individuals and its contribution to HNC is less evident in Indian studies. HR‐HPV‐associated HNC in this region accounts for 26%, with an active infection in 8%–15% of these tumors. There is a lack of concordance in the expression of p16 as a surrogate marker for HPV detection in HNC because of differences in behavioral risk factors. Due to a lack of evidence, treatment de‐escalation cannot be implemented despite the improved outcome of HPV‐associated oropharyngeal cancers. This review critically analyzes the existing literature on the dynamics of oral HPV infection and HPV‐associated HNC, identifying potential avenues for future research. A better understanding of the oncogenic role of HR‐HPV in HNC will help to formulate novel therapeutic approaches and is expected to have a significant public health impact as preventive strategies can be implemented. High‐risk human papillomavirus (HR‐HPV)‐associated oral and oropharyngeal cancer has recently gained clinical attention due to their distinct clinical and molecular characteristics. However, the natural history of oral HPV from acquisition to persistence and malignant transformation is still unclear. This review critically analyzes the literature on the dynamics of oral HPV infection and HPV‐associated head and neck cancer (HNC), identifying potential avenues for future research. Most HPV infections are transient and cleared by the host immune system without clinical symptoms. Both high‐risk and low‐risk‐HPV (LR‐HPV) exhibit restricted viral gene expression sufficient for basal cell genome maintenance and persistence.Integration of the HPV genome into human chromosomes and expression of E6/ E7 oncoproteins have been the classic mechanism for HPV‐mediated oncogenesis. However, extrachromosomal circular oncogenic DNA (ecDNA), including viral circular DNA (vcDNA), human ecDNA, or viral–human hybrid ecDNA, was observed as a dominant feature of HPV+ (HPV‐positive) OPSCC.HPV viral loads are lower in HNC, except in OPSCC, which exhibits higher loads, mainly in the episomal form.A recent study has reported an intact E2 gene in 68% of HPV16+ OPSCC along with a high viral load and E2/E6/E7 oncogene expression with improved clinical outcomes compared with patients with disrupted E2 gene.3The virus‐host interactions modulate the host genetic and epigenetic landscape that facilitates tumorigenesis. HPV‐positive tumors have exhibited a complete mutual exclusivity with driver mutations in TP53, CDKN2A, and TERT. HPV is associated with an increase in mutations attributed to the mutational signature 2, as evidenced by the overexpression of APOBEC3B.Epigenetic alterations like DNA hypermethylation are common in HPV‐related OPSCC. Checking for DNA methylation of a group of cellular or viral genes has been proposed as a second triage in cervical cancer screening programs. It helps to identify the premalignant lesions that could potentially progress (persistent infection) from transient infection.Viral miRNAs modulate cellular and viral gene expression, promoting carcinogenesis and evading the host's immune system. HPV16 encodes HPV16‐miR‐1, HPV16‐miR‐2, and HPV16‐miR‐3, which are known to target cellular genes promoting carcinogenesis.4Currently, the HPV tests approved by the United States Food and Drug Administration (FDA) are those intended for use in cervical cancer. There are no HPV diagnostic tests with regulatory approval for OPSCC. HPV testing options available for use in the clinical setting for HNC are laboratory‐developed tests (LDTs), which involve various techniques and platforms for the analysis of HPV DNA, HPV messenger RNA (mRNA), or the p16 protein.VLP) based enzyme‐linked immunosorbent assays (ELISA) are the standard serological assays used to measure the HPV‐specific antibody responses following vaccination.Next‐generation sequencing (NGS)‐based “capture HPV” is feasible on biopsies and ctDNA. It helps characterize HPV integration status and sites and could define prognostic subgroups in HPV+ OPSCC.TP53 mutations and high levels of EGFR expression, which is absent in HPV+ tumors.The widely used test in clinical practice is p16 (INK4A) detection by immunohistochemistry (IHC). p16 protein is a surrogate marker of transcriptionally active HR‐HPV infection with a sensitivity of 100%. It has demonstrated good agreement with HPV E6/E7 mRNA expression detected by RT‐PCR and RNA ISH.5The average global prevalence of oral mucosal HPV infection in healthy individuals is 4.9% (0%–24%).The oral mucosal HPV dynamics from infection to malignancy are still not understood and extrapolated mainly from cervical cancer studies. A study on oral HPV infection (HIM Study) in 1626 healthy men has shown that the mean duration of HPV and HPV16 persistence were 6.9 and 7.3 months, respectively, with clearance of most HPV infections (63%).6The average global prevalence of HR‐HPV DNA positivity in HNC is 45·8% (OPSCC), 24·2% (OSCC), and 22.1% .In the West, HR‐HPV+ OPSCC has significantly increased from 40∙5% before 2000 to 72∙2% between 2005 and 2009, indicating a time trend increase in incidence.The average prevalence rate of HPV DNA‐positive HNC in India is 26%, based on a review of PCR‐based studies, mainly focused on OSCC Table  Supplem. Complet7HPV is sexually acquired, and early sexual debut, a high number of sexual partners, including oral sex partners, and previous genital warts, provide an increased risk for HPV‐positive OPSCC.Oral HPV prevalence remains either stable or increases significantly with age. Bimodal peaks of HPV infection at 30 and 55 years for men and about 25 and 55 years for women might be due to increased sexual activities in the young and a decrease in immunity with age.Homosexual males (12.2%) had a higher oral HPV prevalence compared with heterosexual males (4.7%) and females (2.9%).8HPV‐associated HNC risk is highest for the tonsil (OR: 15.1), intermediate for the oropharynx (excluding tonsil) (OR: 4.3), and lowest for the oral and larynx (OR: 2.0).The relative risk of developing OSCC or OPSCC among women with cervical cancer is more significant, with a standardized incidence rate of 1.5 and 2.7, respectively.9NCT03811015; and NCT03452137).HPV‐related OPSCC has shown better outcomes with conventional treatment in more developed countries, with a 70% relative reduction in risk of death.The improved prognosis of patients with HPV‐associated HNC is mainly due to tumor radiosensitivity, active immune response to viral antigens, and the absence of field cancerization especially in nonsmokers. Besides, the lack of p53 mutations enables an intact apoptotic response to radiation and chemotherapy.IKZF3], ARHGAP26, and calcium voltage‐gated channel subunit α1 D [CACNA1D] genes are associated with poor prognosis, representing a promising prognostic biomarker. These genes are linked to the differential HPV biology among HPV+ OPSCC.RBL1, RBL2, UBE3A, EP300, and CREBBP, in the high‐risk group of HPV+ OPSCC patients are similar to those in HPV‐negative OPSCC cases. The expression of the E1^E4 splicing isoform is significantly higher among tumors with a better outcome. E1^E4 protein inhibits E6/E7 activity, prevents E2 degradation, and downregulates mitochondrial genes.Gene expression data reveal a subset of HPV+ OPSCC (high‐risk group) with a differential expression of 38 HPV‐correlated genes associated with poor outcomes. Among them, lower expression of IKAROS family zinc finger 3 [10The increasing incidence of HR‐HPV‐associated OPSCC warrants the identification of healthy individuals with a persistent oral HR‐HPV infection, as it helps to detect individuals at risk for OPSCC. HR‐HPV detection is simple as the method is noninvasive, cost‐effective, and serves as a biomarker for HPV+ HNC.NCT04199689) is underway to evaluate the safety and efficacy of Gardasil 9 vaccination to prevent persistent oral HR‐HPV infection, a surrogate endpoint for HPV‐related HNCs.Another effective prevention method is HPV vaccination, but its exact role in preventing HNC is still unclear. Cervarix, a bivalent vaccine against HPV16/18; Gardasil, a quadrivalent vaccine against HPV6/11/16/18; and a recent nonavalent vaccine against 9 HPV types (Gardasil 9) are available worldwide. The United States Advisory Committee on Immunization Practices (ACIP) recommends HPV vaccination for females and males aged from 9 to 26.Close, long‐term monitoring of HPV types helps detect evolutionary changes.11Despite an extensive literature on the role of HPV infection in HNC, most of these studies were based on identifying HPV DNA in the tumor. Nevertheless, active HPV infection is defined by detecting E6/E7 mRNA and p16 expression in tumors. Geographical variation in the contributory factors is also responsible for different HR‐HPV‐positive HNC frequencies. The prevalence of oral mucosal HPV infection in healthy individuals and their contribution to HNC remains unclear from Indian studies. Thus, it is crucial to understand the natural history of oral HPV infection in India, as this will significantly impact HPV prevention strategies in this region. The current data on oral HPV infection are based on PCR detection of HPV types that cause cervical cancer. However, sequencing studies have shown a complex oral HPV community in healthy individuals. This necessitates more research to understand the oncogenic role of other mucosal and cutaneous HPV genotypes apart from those found commonly in the cervical region. HR‐HPV‐associated OPSCC responds well to treatment with a superior survival rate. However, studies about their outcomes are limited for Indian patients and are necessary to refine current treatment regimens. Further research on HR‐HPV‐positive HNC will help understand their biology leading to more specific treatments. Table .Nandimandalam Venkata Vani: Conceptualization ; data curation ; formal analysis ; funding acquisition ; visualization ; writing – original draft (lead); writing – review and editing . Rajendran Madhanagopal: Conceptualization (supporting); data curation ; formal analysis (supporting); visualization (supporting); writing – original draft (supporting); writing – review and editing (supporting). Swaminathan Rajaraman: Conceptualization ; formal analysis ; funding acquisition ; supervision ; writing – review and editing . Trivadi Sundaram Ganesan: Conceptualization ; formal analysis ; funding acquisition ; supervision (lead); visualization (lead); writing – original draft (supporting); writing – review and editing (lead).The authors declare no potential conflicts of interest.Data S1Click here for additional data file.