EVIDENCE FOR THE ASSOCIATION BETWEEN HPV AND ANAL CARCINOMA, BUT NOT FOR COLORRETAL CARCINOMA: A SYSTEMATIC REVIEW

REGISTRO DOI: 10.5281/zenodo.10735449


Larisse Silva Dalla Libera1
Jessica Enocencio Porto Ramos2
Igor Lopes Santos3
Vera Aparecida Saddi4 


ABSTRACT

Introduction: The prevalence of anal cancer has grown considerably and human papillomavirus (HPV) plays a fundamental role in the etiology of these cancers, but in recent years a possible causal role for HPV in colorectal cancer (CRC) has also been investigated. Objective: To describe the prevalence of HPV infection and its genotypes in colorectal and anal cancer in men and women worldwide. Method: Systematic review performed on the databases, PUBMED, LILACS and SCIELO, identified until June 2020. The prevalence of HPV infection in the investigated tumors was estimated by combining data from 42 articles (5834 samples). The x2 test and odds ratio with a 95% confidence interval were used, considering p <0.05. Results: HPV was detected in 88.9% of cases of anal cancer, 10.9% of colorectal cancers, 5.5% in colorectal lesions, polyps and adenomas; 1.2% in colonic tissue adjacent to the tumor and free of neoplasia and 1% in healthy colonic tissues. Genotypes 16 and 18 were the most prevalent in both tumors. HPV was significantly associated with anal squamous cell carcinoma (p <0.0001). Conclusion: HPV may have a causal role in anal cancer, but not in colorectal cancer. 

Key words: Human papillomavirus. Anal cancer. Colorectal cancer.

INTRODUCTION

The incidence of colon cancer (1,148,515 cases), rectum (732,210 cases) and anus (50,865 cases), increased considerably, with 1,931,590 new cases per year and 935,173 deaths (576,858 colon, 339,022 rectum and 19,293 anus) worldwide (1). Genetic susceptibility plays a fundamental role in a subset of CRC cases, but the vast majority of tumors are sporadic and not inherited (2,3). Thus, environmental causes such as infections by pathogens have been investigated as possible causal factors for these cancers (4,5). Anal carcinomas, although rare, are mostly associated with persistent high-risk human papillomavirus (HPV) infections, with HPV 16 and 18 being the most prevalent genotypes (6,7).

HPV is the most common sexually transmitted infection in the population, with sufficient evidence of its carcinogenic effects in different locations (8–10). In colorectal cancer the association of HPV has been investigated in colon and rectal tumors, as well as in precursor lesions and normal colonic tissues, but the results are still controversial (11–14).

Some risk factors can influence susceptibility to viral infection and consequent anal carcinogenesis, such as risky sexual behavior, men who have sex with men, co-infection with the human immunodeficiency virus (HIV), women with intraepithelial cervical lesions, history of anogenital cancer and immunosuppression (15–17). However, the route of viral infection to the colon or rectum is still speculative and includes transmission of HPV through the anal route or contamination during the analysis (18).

Thus, an update of the studies that investigated the association of HPV in colorectal cancer is necessary, since its causal role in these tumors is still questionable (11–14,19,20). In addition, the detection and genotyping of HPV in anal cancers is essential to assess its oncogenic potential and the impact of anti-HPV vaccines on the incidence of these cancers (21). Thus, this review aims to describe the prevalence of HPV infection and its genotypes in colorectal and anal cancers in men and women worldwide.

METHODS

Eligibility Criteria

This study comprises a systematic review of the literature in which complete articles in Portuguese or English were included, assessing the prevalence of HPV and its genotypes in anal carcinomas. The selected articles empoyedPCR and reverse hybridization techniques in paraffinized tissues. The following were excluded: review studies, case reports, studies that investigated only precursor lesions such as anal intraepithelial neoplasms (NIAs); studies that evaluated only a specific group of patients (only men or women, only HIV positive or men who have sex with men, etc.) and studies that used HPV detection by hybrid capture or immunohistochemistry techniques.

The prevalence of HPV in colon and rectum cancers in men and women, was performed for a period of ten years. Studies that evaluated only precursor and benign lesions of the colon and rectum, such as polyps and adenomas, were excluded.

Search and Selection Strategy

The search and collection of data were carried out by means of a search protocol, prepared by the authors, containing a topic of interest, inclusion criteria, search and selection strategies, for obtaining the collected data, analysis and presentation of results and interpretation of the results of the studies.

The publications were consulted on the MEDLINE database through PUBMED; in Latin American and Caribbean Literature in Health Sciences (LILACS) and in the Scientific Electronic Library Online (SCIELO). The consultation period was from January 2020 to June 2020. 

The process of finding and selecting articles was conducted by two researchers independently and blindly to avoid selection bias. When the researchers were unable to establish whether the article would be included or not, a third researcher completed the decision-making process.

To assess the prevalence of HPV in anal and colorectal cancer, bolers and search terms MESH and DECS were used: papillomavirus OR HPV AND anal cancer AND prevalence. And papillomavirus OR HPV AND colorectal cancer AND prevalence.

In order to ensure the inclusion of all articles relevant to the topic, a manual search was carried out in the reference list of review articles, consensus and articles located with this search strategy. No contact was made with clinical researchers to verify possible ongoing research. Duplicate publications have been removed manually.

Data extraction

For each study included, the following were extracted: author; periodical; year of publication; study region; sample size; sample groups, HPV prevalence, sample size tested for HPV DNA; research year; histological type (when stated); origin of the material for DNA extraction from HPV; Primer used to detect HPV DNA; prevalence of HPV genotypes and presence of single or multiple HPV infections. The data extraction was carried out by the researchers independently, in disagreement a third reviewer helped to decide what information would be collected. The figure 1 presents the flowchart of the study search and selection strategy.

Figure 1. Flow diagram outlining the study selection process.

Quality assessment 

This study was carried out according to 27 items included in the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyzes) (22) statement to report recommendations for reporting systematic review and meta-analysis studies. The selected articles were evaluated using the STROBE (Strengthening the Reporting of Observational Studies in Epidemiology) criteria (23).

Statistical Analysis 

Estimates of HPV prevalence were expressed as percentages of all cases tested for HPV DNA and also when there was information by sex, age, study region, sample group and histological type of the tumor. The x2 test and odds ratio with a 95% confidence interval were used, considering p <0.05.

From anal cancer studies that presented estimates of anal intraepithelial lesions and invasive anal cancer, only invasive anal cancer data were collected and the total number of samples was also separated.

Estimates of the prevalence of HPV genotypes were expressed as percentages of all cases tested and the analyzes were limited to those studies that genotyped at least HPV 16 and 18. HPV genotypes were defined according to the included studies, such as high-risk HPV genotypes: 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68 and low-risk genotypes: 6, 11, 32,34, 40, 42, 43, 44, 53, 54, 55, 61, 70, 72, 73, 74, 81, 83, 84 and 89.

RESULTS

A total of 1384 abstracts were identified, but only 42 studies met the inclusion criteria (7,17,24–63) totaling 5,834 anal and colorectal samples tested for HPV detection (Figure 1).

Detection and genotyping of HPV in anal cancer

A total of 1842 cases of anal cancer were tested for HPV, and the prevalence of the virus in these tumors was 89% (95% CI 87.52-90.38), with prevalence variation between 60.6% to 100% in different studies studies (7,24,30–33,40,41,45,46,48,50,52,56–58). Table 1 presents the characteristics of the studies that investigated the prevalence of HPV in anal cancers.

All studies were retrospective cross-sectional studies and the average age of the subjects ranged between 50 and 65.5 years. More than half of the studies evaluated a period equal to or greater than ten years. The largest study was carried out with 496 cases of anal cancer, diagnosed over a period of 25 years, in 24 countries from different continents (7).

All detection methods were based on the polymerase chain reaction (PCR) technique and used paraffinized samples. The primers most used for HPV detection were GP5/6 and MY09/11 (30,32,40,46,57,58), however four studies did not mention the primer set used (31,33,50,52).

Overall, the prevalence of HPV in anal cancers was higher in Europe (96.9%) and lower in Africa (61.9%). Figure 2 shows the geographical distribution of HPV prevalence worldwide.

The prevalence of HPV by sex is shown in Table 2 and the virus was significantly more prevalent among women than among men p <0.0000001, OR: 3.27, 95% CI 2.26-4.74. 

The presence of HPV DNA was associated with squamous cell carcinomas (p 0.0001, OR 9.05, 95% CI 5.37-15.3). Anal adenocarcinomas were investigated in five studies (31,40,45,56,58) (table 3). HPV 16 was the most prevalent genotype in all studies, followed by HPV 18 and 33; the prevalence of HPV 16 ranged from 70% to 93.7%. Multiple infections were detected in 11 of the 16 included studies. Table 4 shows the genotype distribution of HPV in anal cancers.

Detection and genotyping of HPV in colorectal cancer

In total, 3992 colorectal samples were tested for HPV DNA (among tumors, lesions and neoplasia-free tissues) (11,17,26,28,29,34–39,42–44,47,49,51,53–55,59–64). Of these samples, 339 (8.5%. 95% CI 7.67-9.39) were positive for the presence of the virus, 10.9% in tumors, 5.5% in lesions, polyps and adenomas; 1.2% in tissue adjacent to the tumor free of neoplasia and 1% in healthy tissues. Of the 26 publications included, 11 investigated the presence of HPV only in colorectal tumors, without the presence of a control group (11,35,39,42,43,47,53,55,59–61). The study with the highest number of cases included a sample of 555 cases (2.3% HPV +) (39).

Most of the studies were case-control and used samples preserved in paraffin. The most used method for HPV detection was PCR and the most used primers were GP5 / 6 and MY09 / 11 (25,35–38,43,49,51,54,55,59,65). 

HPV was more prevalent in Latin America (20.9%), followed by Asia (9.4%); North America (6.9%) and Europe (5.1%). The most detected genotype was HPV16 followed by HPV 18. Only one study did not find HPV 16 in colorectal carcinomas (37). The Table 5 presents the characteristics of the studies included.

Figure 2. Geographical distribution of the number of cases tested for HPV and the prevalence of the virus by continent worldwide.

Table 2. Prevalence of HPV by sex in cases where the viral genome was detected.

Table 3. Prevalence of HPV by histological type of anal cancer.

DISCUSSION

In the present review, HPV was detected in 88.9% of anal cancer samples, and in 10.9% of colorectal tumors. The high prevalence of HPV in anal tumors is well described in the literature, especially in squamous cell carcinomas (15,46). As demonstrated in the results, anal SCC rarely occurs in the absence of the virus, demonstrating an association between HPV and the development of anal cancer (p <0.0000001 OR: 9.08 95% CI 5.57-14.82).

In fact, anal carcinogenesis is very similar to that of the cervix, mainly because both anatomical sites have a zone with transitional epithelium. In addition, several factors contribute to HPV-induced anal cancer, such as early onset of sexual activity, sexual practices involving anal intercourse, previous exposure to high-risk HPV genotypes, a history of anogenital injury and other HPV associated cancers, especially in risky groups, such as men who have sex with men, transsexual women and individuals with the human immunodeficiency virus (HIV) (15).

In contrast, the role of HPV in colorectal cancer is not yet clear. Several authors have detected the virus in a portion of colon and rectal tumors and a few have described the presence of viral infection in both tumors,precursor lesions and normal colonic tissues (4,11–14,66). In order to establish the causal relationship between HPV and colorectal cancer, HPV DNA should be present in tumors and precursor lesions of CRC, and absent in healthy tissues. In this review, benign and precursor lesions for colorectal cancer such as polyps and adenomas had a prevalence of 5.5% of HPV DNA, while the normal adjacent tissue and healthy colonic tissue, a prevalence of less than 1.5 % was demonstrated, suggesting that there isn’t an association between HPV and colorectal cancer.

Some studies corroborate the idea that HPV is associated with CRC, as shown by the meta-analysis by Damin et al., 2013, carried out with 16 case-control studies that showed a general prevalence for the virus of 31.9% (95 CI %: 19.3-47.9) (12). However, the most complete meta-analysis appears to be that of Baandrup et al., 2014, which involved 37 publications and a total of 2630 colorectal adenocarcinomas, with a combined HPV prevalence of 11.2% (95% CI: 4.9-19.6). This same study showed a prevalence of HPV of 13.4% in normal tissue adjacent to the tumor; 1.6% in distant healthy tissues and 5.1% in adenomas (14).

A meta-analysis published in Brazil in 2016, in Portuguese, involved 1549 samples of colorectal cancers and described 18 included articles. The results demonstrate an HPV prevalence of 51.8% and a significant global effect (p <0.001), which led the authors to suggest an evident association between HPV and CRC (13). However, the criteria for evaluating the quality of the studies were not described in the study, nor even the guidelines employed for carrying out the systematic review and meta-analysis.

In Zhang meta-analysis (2018) only publications involving the Chinese population were included and the prevalence of the virus in tumor tissues and adenomas was higher than that observed in controls OR 10.78% (95% CI 4.22-27.53), high lighting the association of HPV with CCR in the Chinese population.

Both anal and colorectal tumors had a higher prevalence of HPV genotypes 16 and 18. These genotypes are considered high risk for carcinogenesis and are associated with different types of anogenital cancers (67). Currently, a prophylactic vaccine for the most common HPV genotypes (HPV 6, 11, 16 and 18) is available for most countries. The vaccine has the potential to prevent more than two-thirds of anal cancers in women and men, playing an important role in the primary prevention of these tumors in both sexes (7,21). For this reason, studies evaluating the prevalence of HPV genotypes in different regions are of great importance, making it possible to evaluate the effectiveness of the vaccine over the years.

Multiple infections were found in 11 of the 16 anal cancer studies included in this review (7,25,62,26,32,39–41,47,48,54). A study carried out in Australia tested 112 samples of anal cancer, of which 96.4% were positive for HPV, among which 23.2% tested for multiple infections (39). Risky sexual practices, such as multiple sexual partners, increased exposure to different HPV genotypes, in addition to the use of genotyping methodologies with a broad spectrum of HPV genotypes, can detect a greater number of genotypes (7). Through this review, it was not possible to evaluate the prognostic role of multiple infections in patients diagnosed with anal carcinomas, suggesting the need for research involving viral genotyping and investigating the associations between the clinical and pathological characteristics of each individual with the presence of simple or multiple infections.

Studies involving adenocarcinomas and other types of anal cancer did not show a significant association for HPV infection. In contrast to SCC, adenocarcinoma of the anus is not intrinsically related to HPV infection (68). These data suggest that more than half of anal adenocarcinomas are probably not related to HPV, which is also seen in colorectal cancer. However, although the virus does not show tropism for the glandular epithelium, studies are needed to investigate the etiological role of HPV in adenocarcinomas, such as studies of transcriptionally active infection or viral integration.

All detection methods of the studies included in this review are based on the polymerase chain reaction (PCR) technique and mainly use samples fixed in formaldehyde and preserved in paraffin. The most used primers for detection and genotyping were GP5 / 6 and MY09 / 11. By using several sets of specific primers in the same reaction (multiplex PCR) and amplifying fragments of different sizes for HPV genotyping, PCR is one of the most used methodologies for detecting HPV in paraffin material. The main primers used are PGMY09 / 11 (20 bp primers leading to a 450 bp amplicon); GP5 + / GP6 + primers (produce 150 bp amplicon and are more sensitive); and the SPF10 primers that lead to the amplification of a 65 bp viral DNA fragment (very sensitive detection) (69). As paraffinized samples tend to be more susceptible to DNA degradation compared to frozen or fresh samples (70), high performance detection tests are increasingly used, such as the commercial INNO-LiPA HPV Genotyping Extra II test (Fujirebio Europe ®), which is highly sensitive and allows the simultaneous detection of several genotypes in a single sample (71–73).

The prevalence of HPV in anal cancers was relatively higher in women (0.0000001 OR: 3.27 95% CI 2.26-4.74), which can potentially be explained by the persistence of the virus in the anal canal and by pre-existing in the cervix, vagina or vulva (52,54,67,74). A study evaluated 517 cases of anal cancer diagnosed over 13 years in women and showed that a history of grade 3 cervical intraepithelial neoplasia (NIC3), smoking, use of oral contraceptives, nulliparity and tubal ligation, are risk factors for anal cancer in women (75). Another possibility is that the anal mucosa may be sensitive to estrogenic action (76).

The difference in the prevalence of HPV infection has also been described in relation to geographical variation. The European continent had the highest number of cases of anal cancer and the highest prevalence of HPV (96.9%). Unlike another study that reported a higher prevalence of the virus in North America (7). The lowest prevalence of HPV and the lowest number of cases of anal cancer were observed in Africa (61.9%). However, we emphasize that this review did not include studies that assessed the prevalence of HPV in immunocompromised or HIV-infected individuals. Unfortunately, during data collection, most studies carried out in Africa showed specific populations that did not meet our inclusion criteria.

Differences in the prevalence of HPV in the CRC in relation to geographical location were not observed in this review. Among the studies analyzed, HPV investigated in CRCs was more prevalent in Latin America (20.9%). In two previous meta-analyzes (12,14), the prevalence of HPV infection was higher in South America, ranging from 32% to 45%, and the lowest prevalence (3% or less) was observed in North America, Europe and Australia (14). More recent publications (35,65) demonstrate higher prevalence of HPV infection in CRC in the European continent (12.3% to 16.9%) and in North America, respectively (23.8% to 42.2%) (28,64,77). Probably the differences observed in the prevalence of HPV infection in CRCs may be the result of methodological differences, sample preparation and tumor location (78).

HPV infection in anal and colorectal cancer can have variable prevalence and be influenced by the HPV detection method used. Some limitations can be observed for this review, for instance the heterogeneity of the studies, methodological differences such as sensitivity and specificity of the PCR methods used for HPV detection and genotyping, characteristics of the studied populations, variation in the histological diagnosis, anatomical location of the tumors and inadequate material. Even so, these data strongly support the association of HPV with anal cancer and the possible benefit of the anti-HPV vaccine in the population, in both sexes.

CONCLUSION

Anal cancers had a higher prevalence of HPV infection (89%) than colorectal cancers (8.5%), with genotypes 16 and 18 being the most commonly found. HPV was significantly associated with anal squamous cell carcinoma (p <0.0001) and was more prevalent in women with anal cancers (p <0.0001).

Given the similar histopathological characteristics between anal carcinoma and other anogenital carcinomas, in which HPV infection is the causative agent, HPV is likely to play the same etiological role in anal tumors, but not in colorectal cancers.

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1Universidade Federal de Goiás, Programa de Pós-Graduação em Ciências da Saúde, Faculdade de Medicina, Goiânia, Goiás, CEP 74605-020, Brasil. larisse_dalla@hotmail.com. MD, Universidade Federal de Goiás, Anápolis, Goiás, CEP 75130-670, Brasil. Telephone number: +55-62-993298343. Collaboration in the manuscript: All stages, from search and selection of articles, to the discussion of results. She also performed the statistical analysis.
2Pontificia Universidade Católica de Goiás, Goiânia, Programa de Pós-Graduação em Ciências Ambientais e Saúde, Goiás, CEP 74175-120, Brasil. jessicaenocencio14@gmail.com. Collaboration in the manuscript: All stages, from search and selection of articles, to the discussion of results.
3Pontificia Universidade Católica de Goiás, Goiânia, Programa de Pós-Graduação em Ciências Ambientais e Saúde, Goiás, CEP 74175-120, Brasil. igorlsantoss@outlook.com. Collaboration in the manuscript: Discussion of results.
4Universidade Federal de Goiás, Goiânia, Programa de Pós-Graduação em Ciências da Saúde, Faculdade de Medicina, Goiás, CEP 74605-020, Brasil. verasaddi@gmail.com. Collaboration in the manuscript: Orientation, supervision and correction of the manuscript.