Pathobiology of human papillomaviruses in human immunodeficiency virus – Infected persons

Author’s Accepted Manuscript Pathobiology of Human Papillomaviruses in Human Immunodeficiency Virus – Infected Persons Uma Krishnamurti, Elizabeth R. Unger

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To appear in: Seminars in Diagnostic Pathology Cite this article as: Uma Krishnamurti and Elizabeth R. Unger, Pathobiology of Human Papillomaviruses in Human Immunodeficiency Virus – Infected Persons, Seminars in Diagnostic Pathology, http://dx.doi.org/10.1053/j.semdp.2017.04.005 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting galley proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Pathobiology of Human Papillomaviruses in Human Immunodeficiency Virus – Infected Persons

Uma Krishnamurti MD, PhD Assistant Professor, Department of Pathology Emory University School of Medicine

Elizabeth R. Unger PhD, MD Division of High-Consequence Pathogens and Pathology Centers for Disease Control and Prevention

Disclaimer: The findings and conclusions in this report are those of the authors and do not necessarily represent the official position of the Centers for Disease Control and Prevention.

Correspondence to:

Elizabeth R. Unger Centers for Disease Control and Prevention 1600 Clifton Road, MS G41 Atlanta, GA 30329 eunger @cdc.gov 404-639-3533

Abstract There is a complex interrelationship between human papillomaviruses (HPV) and human immunodeficiency viruses (HIV) that has been recognized from the start of the HIV epidemic. Cervical cancer was used as a surveillance indicator for acquired immunodeficiency syndrome (AIDS) before definitive identification of the viral etiology of either condition were known. Careful epidemiologic studies combined with clinical and laboratory measures of HPV, HPV-associated disease, and HIV have helped us understand many aspects of the relationship between these two virus groups; however, questions remain. The histopathology associated with HPV is identical in HIV-positive and negative patients though the lesions are more frequent, with higher frequency of multiple HPV types, and persistent in HIV infected individuals. In this review we will briefly explain the pathobiology of HPV in HIV-infected persons and the potential impact of secondary (screening) and primary (vaccination) to reduce HPV-associated disease in those infected with HIV.

Introduction The complex interrelationship between human papillomaviruses (HPV) and human immunodeficiency viruses (HIV) has been recognized almost from the start of the HIV epidemic, now more than 35 years ago. Cervical cancer was used as a surveillance indicator for acquired immunodeficiency syndrome (AIDS) before definitive identification of the viral etiology of either condition. Our understanding of both viruses has evolved as laboratory methods for detection have changed. Careful epidemiologic studies combined with clinical and laboratory measures of HPV, HPV-associated disease, and HIV have clarified many aspects of the relationship between these two virus groups, but, perhaps not surprisingly, questions remain. Effective therapy for HIV infection and effective primary prevention for HPV are now available, and widespread implementation has the potential to impact the likelihood of disease associated with either virus. In the meantime, pathologists will still be confronted with HPV-associated pathology in patients infected with HIV. While HPV-associated pathology is histologically identical in HIV-positive and negative patients, the lesions are more frequent and persistent in HIV patients. The purpose of this review is to briefly explain the pathobiology of HPV in HIVinfected persons and the potential impact of secondary (screening) and primary (vaccination) to reduce HPV-associated disease in those infected with HIV.

Overview of HPV biology and natural history The abbreviation “HPV” masks the fact that this is not a single virus, but a large number of viruses in the family Papillomaviridae.1,2 All papillomaviruses (PV) are small double-stranded DNA viruses with a circular genome about 8 kb in length. These viruses characteristically infect epithelial surfaces causing epithelial proliferations that project from the surface, recognized as

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papillomas or warts. This characteristic is the source of their name. PV are however also commonly associated with latency or asymptomatic infection. PV are found in a wide variety of species, including birds, reptiles, and many mammals, but each is species-specific.3 Most HPV types are in three papillomavirus genera – alpha-, beta- and gammapapillomaviruses. HPV are classically identified as types based on nucleotide sequence and numbered in the order of their discovery. This approach was taken because HPV cannot be cultured by conventional techniques and antibody responses to natural infection are weak, making “serotypes” impractical.2 A new type requires more than 10% difference in sequence in the L1 gene. Currently more than 200 types are listed in the International HPV Reference Center and Papillomavirus Episteme (PaVE) database.4,5 The genomic organization of all HPV types is similar, with open reading frames (ORFs) identified for 2 late genes (L1 and L2) encoding capsid proteins and 5 early genes (E1, E2, E4, E6 and E7), Figure 1. (Note – there is no E3 as genes were named following homology to the bovine papillomaviruses and E3 ORF is missing in HPV). The protein products are transcribed from only one strand in polycistronic messages with a complex translational pattern. Given the limited number of viral proteins, the virus is dependent on host cell proteins for many functions. The E6 and E7 proteins interact with p53 and Rb host proteins (among many others), prolonging cell cycle proliferation. This allows viral replication to proceed in normally quiescent epithelial layers. HPV infection is non-lytic and confined to the epithelium. Virions are assembled in differentiated cells and are shed in desquamating cells. The combination of restriction to the epithelial compartment and little cellular damage shields the virus from the host immune system. Through the same mechanisms that E6 and E7 proteins use to support productive infection, these proteins also contribute to host cell genomic instability. HPV transformation can be viewed as

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an abortive viral infection .6 Genital HPV is the most common sexually acquired infection. Estimates in the US for the year 2008 indicate HPV contributes 71% of all prevalent and nearly 72% of incident sexually transmitted infections among persons of all ages (Figure 2).7 The natural history of HPV has been most carefully studied in the cervix, and forms the basis for current understanding of anogenital HPV infection and oncogenesis.8 Most infections are asymptomatic and the majority cleared within a few years of acquisition. About 40 HPV types are common in the genital tract, and 14 are included in clinical assays because of their oncogenic potential (HPV 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68). Persistent oncogenic HPV infection is a necessary, but not sufficient for development of cervical cancer. The time between infection and cancer development is long (decades). HPV is causally associated with a variable proportion of other anogenital cancers (anal, vulvar, vaginal, penile) and oropharyngeal cancers. Based on a study of archived tissues collected between the years of 1993 -2005 from 7 US population-based cancer registries, HPV was detected in 90.6% of cervical, 91.1% of anal, 75.0% of vaginal, 70.1% of oropharyngeal, 68.8% of vulvar and 63.3% of penile cancers.9 Benign lesions associated with genital HPV, condylomas (warts) and low grade epithelial abnormalities, are generally associated with other HPV types and occur within a shorter time-frame of infection.

Interaction of HIV and HPV Given the high prevalence of HPV and shared transmission through sexual contact, most HIVinfected persons will be co-infected with HPV. Studies have consistently found HIV-positive persons to have a higher prevalence of HPV and higher frequency of multiple type HPV

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infections compared to those without HIV at oral and anogenital sites. HIV and HPV interactions may favor these co-infections. HPV infection is initiated in the basal epithelial cells, so changes in epithelial integrity associated with cytokines produced by HIV-infected cells may increase the likelihood of HPV infection. Further, immune suppression may impair clearance of HPV once infection occurs. In addition, HIV proteins may directly interact with HPV proteins to enhance HPV transcription as HIV-infected cells can be demonstrated to co-localize in HPV-associated lesions. It is also possible that HPV facilitates HIV infection. While challenging to separate in epidemiologic studies, several studies have found the risk of HIV acquisition increased in women with cervical HPV as well as in men with anal HPV.10-12 Mechanisms by which HPV could facilitate HIV infection are not clear. An interesting hypothesis is that HPV could recruit inflammatory cells susceptible to HIV infection to the epithelium. This was suggested by two studies that found HIV acquisition was associated with HPV non-persistence (i.e., clearance), a situation in which T-cells are recruited.12

HIV/HPV related disease: Pathology view point Persons with HIV and AIDS have an elevated risk for a variety of cancers. Five different malignancies (cervical cancer, Kaposi’s sarcoma, Burkitt’s lymphoma, immunoblastic lymphoma, and primary lymphoma of brain) are included as Stage-3 AIDS defining opportunistic illnesses in HIV infection. Non-AIDS defining cancers are also increased in HIV patients and these include various HPV-related cancers. A meta-analysis of 7 studies involving 444,172 HIV patients, calculated the standardized incidence ratio of cancers at various sites in these patients compared to the general population as 5.82 (95% C.I -2.98-11.3) for the cervix,

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6.45 (95% C.I 4.07-10.2) for the vulva and vagina, 28.75 (95% C.I 21.6-38.3) for the anus, and as 2.32 (95% C.I 1.65-3.25) for the oropharynx & larynx.13 Interestingly, despite the increased infection with multiple HPV types, the types detected in invasive cancers have not been found to differ between HIV-positive and negative patients.14 With the introduction of effective HIV therapies, HIV patients are living longer. The impact of anti-retroviral therapy has been seen in the marked decline in the incidence of Kaposi’s sarcoma and lymphomas, however the impact on HPV-associated cancers in the HIV population has been less clear.10 As a result, the non-AIDS defining cancers are increasing in this population.15 This increasing cancer burden means that pathologists will be involved in evaluating increasing number of cancers and precancers from this population. Considerations for lesions and screening related to each anatomic site are briefly reviewed.

Lower Anogenital Squamous Terminology (LAST) nomenclature and use of p16 The LAST project adopted nomenclature that unifies histopathologic terminology for precancers across all lower anogenital sites.16 The three grades of intraepithelial neoplasia (-IN) terminology were replaced with low and high grade squamous intraepithelial lesions (LSIL, HSIL). LAST recommends that H&E histologic evaluation is supplemented with detection of p16, a marker reflecting HPV E6/E7-driven cell proliferation, to guide diagnosis when: 1. Differential diagnosis is between HSIL and a mimic of pre-cancer (such as immature squamous metaplasia, reparative epithelial changes, tangential cutting) 2. Histologic diagnosis of –IN grade 2 is considered 3. There is a professional disagreement in diagnosis with differential including HSIL 4. Biopsy referral indicates high grade disease but histology is LSIL or less16

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Unless category 4 applies, use of p16 IHC is not appropriate for negative or LSIL histology. It should not be used when histologic diagnosis is –IN grade3. Strong and diffuse block p16 positivity supports a diagnosis of HSIL, while negative or non-block positive staining strongly favors LSIL or a non HPV-associated pathology. Any identified p16-positive area must meet H&E morphologic criteria for a high-grade lesion to be interpreted as such. These recommendations do not differ in the setting of HIV infection, however p16 IHC may be warranted more frequently in evaluating tissues from HIV-positive patients because of the underlying higher risk for precancer.

Anal Lesions Lesions of the anal canal merit special attention since compared to all other sites, the highest risk of HPV driven disease in HIV patients is of invasive anal cancer. While anal cancer is uncommon, there were 6,577 new cases (4,136 women and 2,441 men) and 900 deaths (529 women and 371 men) in the United States in 2013.17 Patients at high risk of developing anal cancer are men who have sex with men (MSM), HIV positive men and women, organ transplant recipients, and women with history of multicentric lower genital tract neoplasia.13, 18 The pooled anal cancer incidence in HIV positive men was 45·9 per 100,000 men (95% CI 31·2-60·3), in contrast to 5·1 per 100,000 (95% CI 0-11·5) in HIV negative men.18 In HIV-infected MSM, the highest risk group, the incidence of anal cancer is > 30 fold greater than in the general population.19 As in non-HIV patients, anal precursor squamous intraepithelial lesions (ASIL) arise in the transformation zone and anal HSIL has the clear potential to progress to anal cancer. Progression rate of HSIL to anal cancer has been estimated to be about 1 in 377/year in HIV

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positive MSM and 1 in 4,196/year in HIV-negative MSM. The progression of HSIL to anal cancer is lower than rates of progression of cervical HSIL (CIN3) to cancer which is estimated at about 1%/year in HIV-negative women.20 Prevalent anal HSIL was reported to have progressed to anal cancer over an average of 57 months compared to 64 months for incident HSIL.21 Furthermore, the majority of patients with HSIL do not regress while receiving antiretroviral therapy (HAART), and HAART is not associated with a reduced prevalence of ASIL or reduced incidence of invasive anal cancer.22 Among HIV-infected women, 42% have detectable anal and cervical HPV infection compared with 8% of HIV negative (P < 0.001). HIV-infected women are more likely to have the same HPV genotype in the anus and cervix than HIV-uninfected women (18 vs. 3%, P < 0.001).23 The risk of ASIL was found to be 13 times greater for women with HPV detected at 3 or 4 genital sites.24 Genital condylomas tend to be larger, more numerous, and have increased recurrence after treatment in HIV positive persons.25-27 Anal condylomas in HIV positive patients harbor foci of high grade disease more frequently (61% versus 25% for HIV negative patients; p = .005), and also harbor foci of invasive squamous cell carcinoma.28 In addition, condylomas in HIV-positive patients have multiple HPV genotypes more frequently and those with high risk HPV are more likely to harbor high grade disease (see Figure 3). These observations suggest condylomas in HIV-positive patients may require more careful examination than usual. Smaller condylomas should be entirely submitted for histopathologic evaluation and larger condylomas should be well sampled. The slides should be carefully reviewed for foci of high grade squamous intraepithelial lesion (HSIL), including keratinizing SIL that appears as cells with squamous atypia and pleomorphic nuclei but with more eosinophilic cytoplasm than is typically seen in mucosal high grade lesions (Figure 4).16

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Cervical lesions Both HPV infection and HPV-associated cervical abnormalities have long been recognized to be increased in HIV-positive compared with HIV-negative women.10, 14 A systematic review of global data found a wide range of incidence and progression for cervical lesions in HIV-positive women.29 The review reported the median incidence of cervical lesions was 3-fold higher and progression rate twice as high in HIV-positive compared to HIV-negative women. Fewer studies included data on risk by CD4 count, but the majority found lower CD4 counts were associated with increased incidence and progression in HIV-positive women; although statistical significance was not usually achieved.29 Similarly the impact of HAART on incidence and progression was not consistent.

Other lower genital sites Vulvar wart incidence and prevalence has been shown to be increased in HIV-positive women. A 13-year follow-up of a cohort of 2,791 HIV-positive and 953 HIV-negative women found a 2.7-fold higher prevalence (5.3% versus 1.9%) and 3.6-fold higher incidence (33% versus 9%) in HIV-positive than in HIV-negative women.30 This was correlated with CD4 cell counts. However, even in HIV-positive women, regression of warts was common (82%, most within the first year), leading the authors to suggest that persistent warts, even with typical appearance, should be biopsied.30 In this cohort, incident vulvar intraepithelial neoplasia (VIN) of any grade was rare, but 6-fold higher in HIV-positive compared to HIV negative women (0.42% versus 0.07%). HPV-associated disease in HIV-positive women is more likely to be multicentric and involve the vulva, vagina, cervix, and anus compared to HIV-negative

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women.31, 32

Head & neck lesions King et al. reported that the incidence of oral warts in HIV-seropositive patients appears to be increasing in the era of HAART.33 Oral warts were associated with reductions in viral load, suggesting that this may in part be related to immune reconstitution.33 Studies suggest that HIV-infected individuals have a modest (1.5-4 fold) risk for HPV-associated head and neck cancer.34 It is not clear what factors increase the risk of oral HPV incidence, persistence and progression to subsequent head and neck cancer. The relative effects of HIV, reduced immunity, tobacco use, sexual behavior and other factors on the natural history of oral HPV are still not well understood and more long-term studies are needed to explore all these.

Secondary Prevention - Screening Anal cancer screening Routine anal cancer screening with anal cytology in persons with HIV infection, MSM without HIV infection, and the general population is not recommended because more evidence is needed concerning the natural history of anal intraepithelial neoplasia, the best screening methods and target populations, the safety and response to treatments, and other programmatic considerations.35 An annual digital anorectal examination (DARE) may be useful to detect masses on palpation that could be anal cancer in high-risk populations (persons with HIV infection, MSM, and history of receptive anal intercourse). While screening with cytology and/or HPV testing are not recommended, these methods are actively being studied. Anal cytology serves as a screening test for detecting anal squamous intraepithelial

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lesions (ASIL) if treatment is available for patients with HSIL. The entire anal canal from the distal rectal vault to the anal verge distally is the target of screening. This includes the anal transformation zone, and the nonkeratinized and keratinized squamous epithelium of the anal canal. Cytological samples are generally obtained without direct visualization of the anal canal although some clinicians use a small anoscope to introduce the collection device.36, 37 The most commonly used sampling device is a Dacron® or polyester synthetic fiber swab moistened with tap water. The Dacron® swab is better than a cotton swab because it releases its cellular harvest more readily and may be more appropriate for use with liquid-based sampling and the swab is better tolerated by the patient than the cytobrush.38 However, the skill of the operator in collecting an adequate sample is more important than the type of sampling device. “Blind” sampling is reported to be superior to directed sampling of the squamocolumnar junction. Both conventional smears and liquid- based cytological preparations are used. Liquid-based preparations may be better with improved cell yield and less compromising factors such as obscuring fecal material, air-drying, and mechanical artifacts.39 Patient collected samples have also been reported to be sufficient for interpretation.40 A meta-analysis found that overall the sensitivity and specificity of anal cytology for HSIL was comparable to the cervicovaginal Pap test with sensitivity of 69-93% and specificity of 32- 59%.41 However, in HIV-positive men the sensitivity of abnormal cytology to detect high-grade anal neoplasia was higher (87%) compared to HIV-negative MSM (55%).42 Anal cytology interpretations have only moderate to good interobserver agreement with a weighted kappa value of 0.54 for overall agreement for cytology specimens.43 This indicates that continued education in interpretation of anal cytology is needed.44 Anal cytology specimens should be carefully screened for atypical and high grade cells.

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High resolution anoscopy (HRA) is recommended in patients who have HSIL, LSIL, atypical squamous cells cannot exclude HSIL (ASCH), and HPV-positive atypical squamous cells of uncertain significance (ASCUS) cases on anal cytology screening, if resources permit and there is expertise to initiate treatment in patients detected with HSIL. If resources are limited, patients with HSIL and ASCH are prioritized over LSIL and ASCUS/HPV+ cases for HRA.45 Abnormal lesions detected on HRA should be biopsied and screened carefully for foci of HSIL as cytology underestimates the grade of ASIL compared with the corresponding biopsy.46, 47 Atypical parakeratosis is seen in 7% of ASCUS, 41% of LSIL, 53% of ASCH, and 71% of HSIL. Cytology underestimates degree of SIL because of parakeratosis which is seen in 63% negative and 74% of abnormal cases. The prevalence and grade of abnormal anal cytology findings in the HIV positive high-risk population is dramatically different from those seen in cervical screening populations (Table1). The overall abnormal anal cytology rate as well as the rate in each category is 2- to 5-fold higher than the 95th percentile of abnormal cervical cytology rate documented among College of American Pathology (CAP) laboratories. The 7.4% unsatisfactory rate for anal cytology is significantly higher than the 3.4% unsatisfactory rate reported as the 95th percentile for cervical cytology rate documented among CAP laboratories.46, 48 The optimal role of HPV testing for anal cancer screening and triage has yet to be defined.42 Commercially available HPV tests are not FDA-approved for anal samples and HPV testing is not a cost-effective approach for triaging patients because of the high prevalence of HPV, although some have found that reflex HPV testing may be helpful in triaging patients with ASCUS cytology.18, 49, 50 HPV16 genotyping may have a more important role in anal cancer

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screening since most anal squamous cell carcinomas are associated with HPV 16.51 Because combined negative cytology and negative HPV has a high negative- predictive value, a negative HPV test may be useful in high-risk groups.52 A study examining biomarker performance for anal screening found that the best overall performance, as measured by Youden’s index, was observed for HPV E6/E7 mRNA testing, followed by HPV16/18 genotyping, p16/Ki-67 cytology, and HPV DNA testing.53

Cervical cancer screening It is recommended that HIV-positive women younger than 21 years and who are sexually active should be screened for cervical cancer within one year of onset of sexual activity and no later than 21 years of age. In women < 30 years of age, co-testing (Pap test and HPV test) is not recommended. HIV-positive women 21-29 years of age should have a Pap test at the time of the initial diagnosis of HIV infection and annually if the previous Pap test is normal. For ASCUS result, if reflex HPV testing is positive, a referral to colposcopy is recommended. If HPV testing is not available or not done, repeat cytology in 6 to 12 months is recommended. For any result equal to or greater than ASCUS on repeat cytology, referral to colposcopy is recommended. For LSIL or worse [ASCH, atypical glandular cells (AGC) and HSIL] referral to colposcopy is recommended (regardless of reflex HPV result, if done). Recent Centers for Disease Control and Prevention, the National Institutes of Health, and the HIV Medicine Association of the Infectious Diseases Society of America guidelines recommend a Pap test every 3 years after 3 consecutive normal Pap tests in women 21-29 years of age.35 In HIV-positive women 30 years or older a Pap test is recommended every 3 years after 3 consecutive normal Pap tests or Pap test and HPV DNA co-testing every 3 years.

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A negative HPV test has a strong negative predictive value for precancer and cancer similar to HIV-uninfected women.35 Those who are Pap test normal but HPV-positive should have repeat co-testing in one year (unless HPV 16 or 16/8 is positive). If either of the co- tests at one year is abnormal referral to colposcopy is recommended. If the initial Pap test result is ASCUS they should be referred for colposcopy or undergo repeat cytology in 6-12 months and the finding of ASCUS or higher on repeat cytology should result in prompt colposcopy. However, in women < 30 years of age, co-testing (Pap test and HPV test) is not recommended. Screening for cervical cancer in HIV-positive women should continue throughout their lifetime and not end at 65 years as in non- infected women. Women with history of HSIL or invasive cervical cancer should undergo annual testing even after hysterectomy.

Primary Prevention – Vaccination Because HPV infection is a necessary step for cervical cancer, primary cancer prevention is possible through prevention of HPV infection. Currently there are three US FDA-approved and Advisory Committee on Immunization Practices (ACIP) recommended prophylactic HPV vaccines. All three target HPV 16 and 18, the types accounting for approximately 70% of cervical cancers worldwide and the highest proportion of HPV-associated anogenital cancers. All three vaccines are composed of non-infectious L1 protein subunits assembled into virus-like particles (VLPs). The vaccines elicit neutralizing antibodies that bind to type-specific epitopes and prevent binding and cellular entry of HPV. Because the vaccines act to prevent infection, they have no impact once exposure has occurred.54, 55 Gardasil (4vHPV; targeting HPV 6, 11, 16, 18; Merck and Co. Inc., Whitehouse Station, New Jersey) and Gardasil 9 (9vHPV; targeting HPV 6, 11, 16, 18, 31, 33, 45, 52, 58; Merck and

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Co Inc.) are licensed for males and females. Cervarix (2vHPV; targeting HPV 16 and 18; GlaxoSmithKline, Rixensart, Belgium) is licensed for use in females. Currently only 9vHPV is available in the US. The originally approved dosing schedule was for 3-doses, administered at 0, 1-2 months and 6 months, routinely recommended for 11 or 12-year-olds, with extension through age 26 for unvaccinated females or age 21 for unvaccinated males. Recently a two-dose schedule at 0 and 6-12 months was recommended for those initiating the vaccine series at age 9 through 15 years.56 Three doses continue to be recommended for those initiating vaccinations at older ages. Since the HPV types that are causally associated with cancers in the HIV population do not appear to be significantly different from the general population, the same HPV vaccines have the potential to be effective in the HIV population. Because the vaccine formulations are noninfectious, they are safe for use in immunocompromised persons. However, it was not clear if HIV-infected persons could mount an effective immune response. Several studies have shown that HIV-populations, including children, female adolescents and adults, can be safely immunized, although a three-dose series is required and seroconversion rates and titers are lower than in comparable HIV-negative groups.57 Trials are ongoing to demonstrate efficacy for disease endpoints and duration of protection, but prospects for reducing the burden of HPVassociated cancers in those infected with HIV are optimistic.

Conclusion In 1983 Harald zur Hausen’s laboratory published the discovery of HPV 16 and Françoise BarréSinoussi and Luc Montagnier laboratory published the discovery of a novel retrovirus now known as HIV.58, 59 In 2008 the same three investigators shared the Nobel Prize in medicine for

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their discoveries. It seems likely that the scientific and medical progress combating these viruses and their associated illnesses will continue to be related. However, therapies targeting HPV to halt disease progression have proved elusive even while HAART has been successful for HIV. Similarly, prophylactic vaccines for HIV have been unsuccessful despite the progress with HPV vaccines. It is clear that HPV-associated diseases in HIV-positive patients will continue to present challenges for appropriate diagnosis and management until widespread uptake of HPV vaccines reduces HPV prevalence. Unfortunately, the burden of HPV-associated disease remains highest in countries with lowest resources that have yet to implement HPV vaccination, and the same countries have the highest burden of HIV infection. The HIV-HPV interaction remains a public health challenge that requires the attention and resources of the world.

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Table 1 Range of anal cytology abnormalities and corresponding yield of high grade disease found on biopsy. [Data from references 46, 46] Anal Cytology

Anal Cytology Result

Biopsy Diagnosis of HSIL2

Category1

% Range

% Range

ASCUS

20-25

45-53

LSIL

28-30

47-57

ASCH

2.5-6

65-92

HSIL

10-15

81-90

Any Abnormality

60-75

1

cytology Categories from the Bethesda System. ASCUS = Atypical squamous cells of undetermined significance; LSIL = Low grade squamous intraepithelial lesion; ASCH= Atypical squamous cells cannot exclude high grade squamous intraepithelial lesion; HSIL= High grade squamous intraepithelial lesion 2

Diagnosis based on LAST nomenclature for histology. HSIL = High grade squamous intraepithelial lesion

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Figure Legends FIGURE 1 Schematic diagram of genomic organization of open reading frames of HPV. All HPVs are similar in their genetic organization and appearance by electron microscopy. URR, upstream regulatory region.

FIGURE 2 Estimated number of prevalent sexually transmitted infections in the United States, 2008 (All ages). Data was based on age-specific data from the National Health and Nutritional Evaluation Survey 2008 from Satterwhite et al.7

FIGURE 3 3A – Focus of high grade squamous intraepithelial lesion (HSIL; AIN 3) in an anal condyloma (H&E section, original magnification x 100). 3B- p16 immunohistochemical stain highlighting the focus of HSIL in the condyloma (original magnification x 100).

FIGURE 4 4A -Keratinizing dysplasia with atypia in an anal condyloma (H&E section, original magnification x 100). 4B - p16 immunohistochemical stain highlighting the keratinizing dysplasia with atypia as HSIL (AIN 3) in the condyloma (original magnification x 100).

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FIGURE 1

26

FIGURE 2

Estimated Infections in the United States, 2008 (All ages, based on age specific data from NHANES) Trichomoniasis HIV Hepatitis B HPV HSV-2 Syphilis Gonorrhea Chlamydia 0

20,000

40,000

60,000

80,000

100,000

Number Prevalent Infections (thousands)

27

FIGURE 3A

FIGURE 3B

28

FIGURE 4A

FIGURE 4B

29