High Rates of High-risk HPV Anal Infection and Abnormal Cytology in a Cohort of Transgender People Assigned Male at Birth

Abstract

Background

Transgender people assigned male at birth (TG-AMAB) have higher rates of anal human papillomavirus (HPV) infection and anal cancer compared with cisgender populations. In a cohort of TG-AMAB in Washington DC, we determined the prevalence and epidemiological factors associated with anal high-risk HPV (HR-HPV) infection and cytological abnormalities.

Methods

In an urban academic–community clinic, we recruited adults identifying as a gender different than their sex assigned at birth. Participants provided blood samples, anal swabs for HR-HPV and cytology, and completed surveys on sexual behaviors and use of gender-affirming hormones. Logistic regression was used to identify predictors of HR-HPV and abnormal cytology.

Results

Of 97 TG, 80 were AMAB, of whom 66% were people with HIV (PWH); 73% tested positive for any anal HR-HPV, and 48% had abnormal cytology. Only 24% recalled HPV immunization. While TG-AMAB PWH had increased risk of HPV16 infection (37% vs 8%; P = .014), the prevalence rates of any HR-HPV (80% vs 62%; P = .097) and abnormal cytology (56% vs 32%; P = .085) were similarly elevated regardless of HIV status. Among TG-AMAB, estradiol use was not associated with increased odds of any HR-HPV or abnormal cytology (all P > .05); however, higher testosterone levels were associated with any HR-HPV (P = .014). Age ≥35 years was associated with abnormal cytology in TG-AMAB (59% vs 34%; P = .035).

Conclusions

TG-AMAB have a high prevalence of any HR-HPV and abnormal cytology, regardless of age, HIV status, or hormone use. Given high risk for anal cancer and low rates of HPV vaccination, there is a pressing need for primary and secondary anal cancer prevention strategies targeting this population.

Anal cancer, for which high-risk oncogenic human papillomaviruses (HR-HPVs) are considered the main cause, is rare in the general population [1]. However, certain key populations are disproportionately impacted by this cancer, including transgender people (TG), who have an anal cancer prevalence 5.5 times higher than cisgender individuals [2]. Among transgender women (TGW), a high prevalence of anal HR-HPV and HIV infection—which potentiates HR-HPV infection and HPV-related dysplasia—confer an even greater risk of anal cancer [1, 3–6].

ANCHOR, a seminal clinical trial evaluating the impact of treating anal high-grade squamous intraepithelial lesions (HSIL)—the precursor to anal cancer—in people with HIV (PWH), proved that high-resolution anoscopy (HRA)–guided treatment prevents anal cancer [7]. Data from this study also reinforced that TGW are at high risk of developing anal cancer. Of 306 TGW PWH screened for ANCHOR, half had anal HSIL on HRA [7]. ANCHOR led to the development of the first anal cancer screening guidelines by the International Anal Neoplasia Society in 2024 [8], followed by the release of the Department of Health and Human Services anal cancer screening guidelines in people PWH [9]. Regarding TGW, these guidelines recommend screening those PWH ≥35 years old, and those without HIV ≥45 years old, via anal cytology and/or HPV testing. Those with abnormal cytology and/or HPV16 (HPV with the highest oncogenic potential) are referred to HRA [8, 9]. Unfortunately, HRA has limited availability in the United States that cannot meet the high burden of anal dysplasia in populations at risk, including TGW [7, 8, 10]. Further, evidence suggests that efforts for primary prevention of HPV infection via vaccination do not reach TGW. In a study of 49 TGW in 2 US cities, only 10% reported being vaccinated against HPV, and 55% had not heard of the HPV vaccine [11]. These limitations underscore the need for further data to characterize anal cancer risk in TGW, a key population that is underrepresented in anal cancer research [12].

An additional important and unique consideration for HPV in TG is the impact of gender-affirming hormones. In the United States, 80% of TG are estimated to have used or desire to use gender-affirming hormones [13]. However, while anal tissue is known to be a target for sex hormones [14], the impact of these hormones on anal HPV remains unclear [15]. In a cross-sectional study of 272 TGW in Brazil with a 61% prevalence of HR-HPV, self-reported hormone use was not associated with increased prevalence of HR-HPV, although hormone levels and specific hormone use were not reported [16]. The influence of hormones has also been assessed in cisgender populations. In a cohort of 340 cisgender men who have sex with men (MSM), higher testosterone levels were associated with increased anal HPV16 prevalence [17]. Further, the downregulation of estrogen receptor α in 74 human cervical samples was associated with progression of cervical dysplasia to cancer [18].

As such, there remains a limited understanding of factors associated with the natural history of HPV and anal dysplasia in TG, information that is critical for anal cancer prevention in this disproportionately affected community. Therefore, in a cohort of TG in Washington DC, we sought to identify the prevalence of HR-HPV and abnormal anal cytology and understand associated clinical and socio-behavioral factors.

METHODS

Study Design

Through an academic–community–governmental partnership, we have been providing gender-affirming care and treatment and prevention of HIV and sexually transmitted infections (STIs), embedded in a community-based organization providing services to TG. This unique model of health care delivery allowed recruitment of TG across Washington DC, between April 2021 and May 2023, into a longitudinal observational cohort study: “Prospective Analysis of Transgender Individuals At Risk and Living With HIV to Understand the Impact of Integrating HIV Treatment and Prevention Into Transgender Care Utilizing Hormone Therapy (PATCH).” Participants were recruited using convenience sampling and followed every 3 months for 1 year. This analysis includes data from the screening visit. The study protocol was approved by the University of Maryland, Baltimore's Institutional Review Board.

Participants

Eligible participants were 18 years or older and identified as transgender, nonbinary, or a gender identity different than their sex assigned at birth. While acknowledging the diversity of gender identities, all participants are referred to as TG.

Procedures

At screening, from all TG, we collected anal swabs, blood samples, and responses to interviewer-administered surveys related to demographics and sexual behavior. Anal swabs were preferentially collected by clinicians; however, self-collection was allowed per patient preference. The same anal sample was sent for both anal cytology and HR-HPV genotyping using the Roche Cobas linear-array test, which provides individual results for HPV16, HPV18, and pooled results for other HR-HPV variants (31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68) [19]. Participants with inconclusive results were asked to come back for 1 reattempt at the collection of the anal sample. For this analysis, we classified HPV infection based on HPV subset (HPV16, HPV18, or non-16/18 HR-HPV) and included a composite outcome of “any HR-HPV” combining all 3 subsets. We excluded analysis of factors associated with HPV18 alone in the setting of low HPV18 prevalence.

Anal cytology was performed by a Clinical Laboratory Improvement Amendments commercial service. Participants with abnormal anal cytology (defined as high- or low-grade squamous intraepithelial cells or atypical cells of either undetermined significance or without the ability to rule out high-grade changes) were referred to off-site HRA. As our data collection preceded the publication of the current guidelines for anal cancer screening [8], participants testing positive for HPV16 with normal anal cytology were not referred to HRA.

Data and Laboratory Measures

Hormone Use

Estradiol and testosterone levels were measured by electrochemiluminescence immunoassay. In TG-AMAB, current estradiol use was defined based on a serum estradiol level >42.6 pg/mL or, if the lab value was missing, based on self-report. Otherwise, participants were considered not to be on estradiol. Androgen blockade was defined as a testosterone level <264 ng/dL or, if the lab value was missing, based on self-reported use of androgen blockers or a history of orchiectomy (surgical removal of the testicles). Otherwise, participants were considered not to be in androgen blockade. Categorical-level cutoffs were based on the sex-specific assay lab value cutoffs [20, 21] and transgender health guidelines [22].

HIV and STI Data

HIV status was defined based on HIV fourth-generation antigen/antibody testing and medical history. HIV viremia was defined as an HIV viral load ≥200 copies/mL. A diagnosis of active syphilis infection was based on clinician evaluation of rapid plasma reagin testing with reflex treponemal antibody confirmation. Nucleic acid amplification tests were used for diagnosis of chlamydia or gonorrhea infection in the rectum, urine, and pharynx. A positive bacterial STI was defined as active infection with syphilis, chlamydia, and/or gonorrhea.

Behavioral Measures

Transactional sex was defined as sex in exchange for money, housing, or drugs. Prior HPV vaccination, prior anal pap, and substance use were based on self-report [23]. We compared HR-HPV and abnormal cytology prevalence using 35 years as the age cutoff based on the age of screening recommended for TGW PWH [8, 9].

Statistical Analysis

Categorical variables were presented as proportions and continuous variables as means and CIs. The Fisher test and χ² test were used to compare differences based on sex at birth and vaccine status by age. HR-HPV status and cytological abnormalities were compared using logistic regression. Due to the minimal amount of missing data, a complete case analysis was performed. All associations were presented as odds ratios (ORs) and their corresponding 95% CIs. Statistical analyses were conducted in SAS 9.4 (Cary, NC, USA).

RESULTS

Demographics and Sexual Behaviors

Among 97 TG, the median age (interquartile range) was 35 (30–40) years, and 80 (82%) were AMAB. Participants were predominately Black (84%) and unemployed (58%). Among TG-AMAB, in the year before screening, 75% engaged in receptive anal sex and 44% engaged in transactional sex. At screening, 24% tested positive for any bacterial STI (Table 1).

HR-HPV and Abnormal Anal Cytology

Among 75 TG-AMAB with available HPV results, 73% had any anal HR-HPV infection, including 20 (36%) who tested positive for >1 HR-HPV subset. Of participants with any HR-HPV, 52 (69%) tested positive for non-16/18 HR-HPV, 20 (27%) for HPV16, and 9 (12%) for HPV18 (Figure 1).

Of 76 TG-AMAB with available cytology, 36 (48%) had abnormal cytology (Figure 2). Infection with any HR-HPV, HPV16, and non-16/18 HR-HPV were all significantly associated with abnormal anal cytology (P = .002, P = .007, and P = .007, respectively) (Table 2).

HPV Vaccine

In participants AMAB, 24% recalled receiving any HPV vaccination (Figure 3). Reported HPV vaccination was not associated with lower anal HR-HPV, HPV16, or non-16/18 HR-HPV infection or abnormal cytology (all P > .05) (Table 2).

Sex Assigned at Birth

Compared with TG-AMAB, the 17 participants assigned female at birth were significantly less likely to report receptive anal sex in the prior year (75% vs 12%; P < .0001), test positive for any anal HR-HPV (73% vs 8%; P < .0001), or have abnormal anal cytology (48% vs 8%; P = .006) (Table 1). The proportion of participants assigned female at birth who reported HPV vaccination was similar compared with those AMAB (24%; P = 1).

Age and Behavioral Factors

Among participants AMAB, any HR-HPV prevalence was 69% in those age <35 years, and it was 77% in those age ≥35 years (P = .44). In bivariate analysis, people age ≥35 years had greater odds of abnormal anal cytology (59% vs 34%; OR, 2.76; 95% CI, 1.07–7.09; P = .035) (Table 2); however, this association was not significant in the multivariable model (P = .09) (Supplementary Figure 1). Tobacco use, engaging in receptive anal sex, oral sex, insertive penile sex, and transactional sex in the previous year were not associated with HR-HPV or with abnormal anal cytology (all P > .05).

HIV Status

Of 80 TG-AMAB, 66% were PWH, of whom 35% had HIV viremia, and the mean (SD) CD4 count was 739 (423) cells/mm³ (Table 1). Lower mean CD4 count was associated with HPV16 infection (P = .045) and abnormal cytology (P = .035), but not with any HR-HPV or non-16/18 HR-HPV (Figure 4). HIV viremia was not associated with HR-HPV infection or abnormal cytology (all P > .05) (Table 2). In bivariate analysis, TG-AMAB PWH were 6.97 times more likely to test positive for HPV16 compared with those without HIV (95% CI, 1.47–32.99; P = .014). There was no significant difference in prevalence of any HR-HPV infection or abnormal anal cytology by HIV status (all P > .05) (Table 2).

Gender-Affirming Hormones

Of TG-AMAB, 47 (59%) were taking gender-affirming hormones (14% estradiol only, 15% androgen blockade only, 30% both). Six had an orchiectomy. Of the 33 (41%) participants not taking hormones, 14 (42%) reported never taking any hormones. Of 80 TG-AMAB, 82% ever took hormones, and 66% ever took hormones that were prescribed by a provider (Table 1).

In the bivariate analysis, neither estradiol mean serum levels nor current or past estradiol use or androgen blockade was associated with any HR-HPV, HPV16, non-16/18 HR-HPV, or abnormal cytology (all P > .05) (Table 2, Figure 4). However, higher mean testosterone serum levels were associated with any HR-HPV infection (P = .019) and non-16/18 HR-HPV (P = .014) (Figure 4) but not when comparing any HR-HPV status based on categorical testosterone levels (<60 ng/dL vs 60–264 ng/dL; P = .46; and <60 ng/dL vs >264 ng/dL; P = .54) (Table 2).

DISCUSSION

In this anal HPV natural history study focused on TG-AMAB in Washington DC (PATCH), we found a high prevalence of anal HR-HPV and abnormal cytology and low rates of HPV vaccination. These data offer insight into a population that is underrepresented in anal HPV research and that faces unique barriers to health care.

Our data reinforce a high prevalence of HR-HPV in TG-AMAB PWH in PATCH, where 4 out of 5 participants had current anal HR-HPV infection. These findings are similar to prevalence rates found in 2 cohorts of TGW PWH from Latin America (62%–75%) [16, 23] and a meta-analysis of MSM PWH (74%) (Table 3) [24]. These data also reinforce the higher rates of HR-HPV among TG-AMAB and MSM compared with cisgender male PWH who have sex with women, in whom HR-HPV prevalence is 27% (Table 3) [24]. Among TG-AMAB without HIV, we found a striking prevalence of HR-HPV (62%) in PATCH compared with a meta-analysis of MSM without HIV (41%) and of cisgender men without HIV who have sex with women (7%) (Table 3) [24]. Further, rates of abnormal anal cytology were comparable in TG-AMAB from PATCH (49%) and TG-AMAB from Pakistan (48%) [25]. These alarmingly high rates highlight the need to engage TG-AMAB in robust and targeted primary and secondary prevention campaigns. This further highlights the need to better understand anal cancer risk in this population by examining persistent anal HR-HPV infection.

While HIV is a known factor associated with progression of anal dysplasia, in TG-AMAB PWH, HIV viremia was not associated with a higher risk of HPV infection or abnormal cytology in this analysis. However, HPV16 and abnormal cytology were associated with a lower CD4 count. This corroborates previous observations in non-TG that a low CD4 count, rather than HIV suppression, is associated with higher rates of HPV16 infection and anal dysplasia [27]. While current guidelines do not include CD4 count as a factor affecting frequency of anal cancer screening, PWH with a low CD4 count may need more frequent screening given a potential higher risk to develop dysplasia. Further, compared with TG-AMAB PWH, we found no statistically significant difference in rates of HR-HPV infection and abnormal cytology in TG-AMAB without HIV: 2/3 with anal HR-HPV infection and 1/3 with abnormal cytology. While anal cancer risk cannot be estimated based on these cross-sectional data alone, these high rates warrant further clarification about the role of anal cancer screening in TG-AMAB without HIV, particularly as the current guidelines recommend screening this population at age 45 based on data in cisgender MSM [8, 28].

Despite the disproportionate risk of HPV infection, only 1 in 4 participants in our cohort recalled receiving an HPV vaccine. These data suggest that our attempts at primary prevention in this population are woefully inadequate. To date, research has been incapable of estimating HPV vaccination rates in TG-AMAB in the United States [29], although a study in 2 urban US cities found vaccination rates in TGW as low as those from our cohort [11]. The use of gendered language around HPV vaccination, low knowledge regarding HPV prevention, and lack of clinician communication about the HPV vaccine have been examined as potential causes for these low vaccination rates [26, 30, 31]. Of note, in our cohort, participants had low vaccination rates regardless of sex assigned at birth. While some participants may not have been eligible for HPV vaccination in the past, prevention strategies can still adopt catch-up vaccination to protect this population from incident HR-HPV infection [12, 32].

This study offers a unique opportunity to examine associations between sex hormones, anal HPV, and abnormal anal cytology. Among TG-AMAB, we found that higher testosterone levels were significantly associated with a higher risk of any HR-HPV and non-16/18 HR-HPV, similar to a single study of MSM [17]. This association was not seen when comparing HR-HPV status based on categorical testosterone values that reflect true androgen blockade. Whether these levels are associated with overall current or past anal receptive sexual practices remains to be explored. We did not find a correlation between estradiol use or serum estradiol levels and HR-HPV or abnormal cytology, similar to 2 studies from Latin America showing that self-reported hormone use did not increase the risk of anal HPV infection [16, 23]. The use of gender-affirming hormones is dynamic, and longitudinal measurement of serum sex hormone levels will provide important insight to correlations between hormone use and the natural history of anal HPV. Beyond epidemiological associations, our measurement of sex hormone serum levels may best reflect the direct biological impact of gender-affirming hormone therapy on anal dysplasia; however, this biological role should be further explored in pathophysiology studies, particularly as preclinical observations confirm the presence of sex hormone receptors in normal and dysplastic anal tissue [14, 17, 33].

In TG-AMAB age <35 years, the prevalence of any HR-HPV was as high as that of individuals age ≥35 years, regardless of HIV status. These findings parallel what is seen in MSM: Anal HPV prevalence plateaus at a high rate in those age ≥20 years, regardless of HIV status [24]. Additionally, 1/3 TG-AMAB age <35 years had abnormal anal cytology. While conclusions about anal cancer risk are limited based on these data alone, these rates are alarming and highlight the need to implement vaccine strategies—including catch-up vaccination—in this young population, while studying in-depth the risk of developing anal dysplasia in TG-AMAB age <35 years. This is of particular interest as the anal cancer screening guidelines exclude this population from screening recommendations, while data on their longitudinal cancer risk are missing [8].

Our enrollment through a community–academic–governmental collaboration allowed for the engagement of a diverse cohort of TG—a community underrepresented in the HPV literature—into research. Providing community-based HIV treatment and gender-affirming care allowed access to underserved TG and the opportunity to shed light on the HPV burden in this population. As more than half of TG-AMAB reported having a prior prescription of hormones from a provider and 87% of those PWH reported having seen a provider for HIV care in the last year, HIV or gender-affirming care clinics may serve as important locations to engage this population at risk for anal cancer in screening and prevention. Of note, PATCH offers a unique approach to examine anal HPV infection among TG assigned female at birth. This small cohort with low rates of receptive anal sex seemed to have a low risk of anal cancer.

Our study has some limitations. Our sample size is not powered to detect associations with all variables of interest, and the cross-sectional nature of our analysis limits such conclusions. However, our community-based recruitment allowed us to shed light on the burden of HPV in underserved TG. Additionally, a small proportion of our study participants were assigned female at birth, limiting the ability to make conclusions about this population. Further, the measure of HPV vaccination was based on self-report, which may limit accuracy, especially for those vaccinated as children. However, a recent meta-analysis found that self-report of HPV vaccine was adequate in measuring vaccination rates in MSM and TG [29].

In conclusion, in a cohort of TG-AMAB, regardless of HIV status, age, or hormone use, the prevalence of HR-HPV and abnormal cytology was as high as that found in MSM in the literature. History of HPV vaccine receipt was uniformly low. These data emphasize the critical need to scale up primary and secondary strategies for anal cancer prevention in TG-AMAB. Further, HR-HPV prevalence was associated with higher testosterone levels, highlighting the importance of ongoing investigations to better understand the risk of anal cancer based on gender-affirming hormone use in TG-AMAB. Finally, in this cohort, having HIV and being age ≥35 years were not associated with a higher prevalence of HR-HPV or abnormal cytology compared with those without HIV and age <35, emphasizing a need for further research on anal cancer risk and screening in TG-AMAB who are HIV negative and/or age <35 years.

Supplementary Data

Supplementary materials are available at Open Forum Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.

Acknowledgments

The authors thank the study participants for the contribution of their time and commitment to this study. The authors also acknowledge the support of Kathren “Raven” Belter (phlebotomy).

Financial support. This work was supported in part by the Office of AIDS Research (grant HHSN272201300022I). We acknowledge the support of the University of Maryland, Baltimore, Institute for Clinical & Translational Research (ICTR) and the National Center for Advancing Translational Sciences (NCATS) Clinical Translational Science Award (CTSA) (grant UL1TR003098).

Patient consent. Written consent was obtained from all participants. The protocol was approved by the University of Maryland Baltimore's Institutional Review Board committee.

References

Author notes