206
Review article
Review of the current knowledge on the epidemiology, pathogenesis, and prevention of human papillomavirus infection Asia Asiafa, Shiekh T. Ahmadb, Sheikh O. Mohammadc and Mohammad A. Zargara Human papillomavirus (HPV) infection is a central and necessary, although not sufficient, cause of cervical cancer. Besides HPV, the additional multiple risk factors related with the onset of cervical cancer are early-age sexual activities; high number of sexual partners, which is the most salient risk factor; suppression and alteration of the immune status; long-term use of oral contraceptives; and other hormonal influences. The tumor-suppressor proteins p53 and pRb are degraded and destabilized through ubiquitination by viral oncoproteins E6 and E7. Over 95% of cervical cancer cases worldwide test positive for oncogenic HPV DNA. Although cervical screening procedures have been successful in reducing the disease burden associated with HPV infection because of lack of resources or inadequate infrastructure many countries have failed to reduce cervical cancer mortality. Therefore, prevention may be a valuable strategy for reducing the economic and disease burden of HPV infection. At present, two successful prophylactic HPV vaccines are available, quadrivalent (HPV16/18/6/11) ‘Gardasil’ and bivalent (HPV16/18) ‘Cervarix’ for vaccinating young adolescent girls at or before the onset of puberty. Recent data indicate that vaccination prevents the development of cervical lesions in women who have not already acquired the vaccine-specific HPV types. Moreover, several therapeutic vaccines that are protein/peptide-based, DNA-based,
or cell-based are in clinical trials but are yet to establish their efficacy; these vaccines are likely to provide important future health benefits. The therapeutic vaccination mode of prevention is a promising area of research, as revealed in preclinical trials; however, clinical trials based on large populations are warranted before reaching a valid conclusion. This review summarizes the studies on the epidemiology of HPV infection, the pathogenesis of viral oncoproteins in the oncogenesis of cervical cancer, the economic and health burden of HPV-related diseases, and, finally, focuses on the results of recent clinical vaccination trials. European Journal of Cancer c 2014 Wolters Kluwer Health | Prevention 23:206–224 Lippincott Williams & Wilkins.
Introduction
activities in the field. The main contribution to this was made by the German scientist Harald Zur Hausen, who for the first time showed the robust association between HPV infections and cervical cancer by identifying, cloning, and sequencing the two most important high-risk HPV types, that is, 16 and 18, from cervical tumor specimens (Zur Hausen, 1991, 1996). Subsequent research established that HPV is also associated with a substantial number of oral, esophageal, and other anogenital cancers, namely, cancers of the vulva, vagina, penis, and anus, in addition to the development of cervical cancer (Kashima and Mounts, 1987; Greenspan et al., 1988; Zur Hausen, 1989; Syrjanen, 2002; Dillner et al., 2007). Epidemiological case series have shown that HPV DNA is found in nearly 100% of cervical cancer cases (Walboomers et al., 1999). At present, there are neither effective methods for preventing HPV infection nor promising treatments available for the management of HPV-associated diseases: for
Human papillomaviruses (HPVs) are a group of small, nonenveloped, double-stranded DNA viruses belonging to the family Papovaviridae. This group of viruses infects a variety of organisms across the animal kingdom, ranging from birds to mammals, including humans, and the viruses are specific for their respective hosts. These viruses appear to show similarities with polyomaviruses and consist of about 72 capsomers. The circular HPV genome comprises 7500–8000 bp of DNA that is associated with histones and is compacted into aggregates similar to chromatin (Fuchs et al., 2006). HPV is known to be the main causative agent for cervical carcinomas. In early 1976, it was discovered that novel HPV types are involved in the successful development of cervical tumors, and, since then, various epidemiological, molecular, and biological studies have justified this observation. This led to immense enhancement in research c 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins 0959-8278
European Journal of Cancer Prevention 2014, 23:206–224 Keywords: cervical cancer, dysplasia, E6, E7, human papillomavirus, vaccine a Department of Biochemistry, Faculty of Biological Science, University of Kashmir, Srinagar, bInstitute for Stem Cell Biology & Regenerative Medicine, NCBS (TIFR) Campus, Bangalore and cDepartment of Biochemistry and Molecular Biology, School of life Sciences, Pondicherry University, Puducherry, India
Correspondence to Mohammad A. Zargar, PhD, Department of Biochemistry, Faculty of Biological Science, University of Kashmir, Srinagar 190006, India Tel: + 91 194 242 8723; fax: + 91 194 242 1353; e-mail: [email protected] Received 4 April 2013 Accepted 25 June 2013
DOI: 10.1097/CEJ.0b013e328364f273
Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.
Epidemiology, pathogenesis, and prevention of HPV Asiaf et al. 207
preinvasive lesions such as genital warts and cytological abnormalities, the treatment consists of destruction of tissue through electrocoagulation (use of intense heat generated by electric current), cryotherapy (use of extreme cold), laser ablation, or local surgery, whereas invasive cervical cancers are generally managed with surgery, radiation, both surgery and radiation, and in some cases chemotherapy and observation for recurrence. However, these therapies are effective only when the disease is localized, as in the case of cervical intraepithelial neoplasia (CIN); the possibility of recurrence remains, as treatment does not always eradicate the underlying HPV infection. Prophylactic HPV vaccines (Gardasil and Cervarix), which are constructed using virus-like particles, prevent the acquisition of low-grade and high-grade squamous intraepithelial lesions (LSIL and HSIL) of the cervix by generating strong neutralizing antibodies against capsid antigens L1 in recipients; these antibodies likely mediate the observed protection against HPV infections (McElrath et al., 2008). However, in many HPV-associated neoplasias, detectable levels of capsid antigens (L1 or L2) are not expressed by infected cells. Therefore, current prophylactic vaccines are not likely to be effective in preventing disease progression in preexisting HPV infections or HPV-associated lesions, and there is a burning need to develop therapeutic HPV vaccines. Successful immunotherapy might therefore be a preferred mode of treatment because it can target all HPV-associated lesions irrespective of their location and can induce long-lasting immunity, thus preventing recurrence. Therapeutic cancer vaccines are expected to treat an existing cancer by enhancing the naturally occurring immune response against the cancer.
Low-risk and high-risk human papillomaviruses To date, more than 120 HPV types have been isolated, completely sequenced, and classified broadly into mucosal/genital and cutaneous types based on sequence analyses and clinical manifestations (Bernard et al., 2005). HPVs have been phylogenetically categorized into five genera: alpha, beta, gamma, delta, and mu based on the differences in their nucleotide sequences (De Villiers et al., 2004). The mucosal/genital HPV types belong to the alpha-PV genus and can be divided into nononcogenic (low-risk) or potentially oncogenic (high-risk) types according to their presence in malignant lesions (Bosch et al., 2002; Munoz et al., 2003). Fifteen HPVs have been classified as high-risk types (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, and 82) that cause dysplasia and cancer, 12 have been classified as low-risk types (6, 11, 40, 42, 43, 44, 54, 61, 70, 72, 81, and CP6108) that usually cause low-grade mild dysplasia, genital warts, and respiratory papillomatosis, and three have been classified as probable high-risk types (26, 53, and 66) (Munoz et al., 2003). HPVs 31, 33, 35, 51, and 52 are sometimes
regarded as ‘intermediate-risk’ viruses because they are more common in mild or severe dysplastic lesions than in carcinomas (Fernandes et al., 2009). Cutaneous HPVs constitute more than 75% of the HPVs described to date and are grouped into five different genera: alpha-PV, betaPV, gamma-PV, mu-PV, and nu-PV (De Villiers et al., 2004) that are not generally associated with cancers. Certain beta types have been implicated in the development of nonmelanoma skin cancers in immunosuppressed individuals and epidermodysplasia verruciformis (EV) patients. Their possible role in cancer progression in the general population is currently unresolved (Doorbar et al., 2012). Each genotype is characterized on the basis of a difference of more than 10% in the DNA sequence of the L1 open reading frame (ORF) from the closest known HPV type. If a DNA difference of 2–10% exists, the two viruses are considered subtypes of the same HPV type. Variants are defined if the sequences of their L1 genes are at a maximum 2% dissimilar from one another (De Villiers et al., 2004; Chow et al., 2010). Before the development of full-fledged cervical carcinoma, CIN appears, which is the consequence of persistent cervical infection with a highrisk HPV genotype with specific symptoms until its progression into a tumor (Ho et al., 1995). Among all the HPV types, type 16 is the most carcinogenic HPV genotype (Munoz et al., 2003; De Sanjose et al., 2010) and is responsible for B54.4% of all cervical cancers. The minor differences between the developing and developed regions of the world are shown in Fig. 1. HPV-18 is the next most carcinogenic HPV type and accounts for 16.5% of cervical cancers. Approximately 17 other HPV genotypes cause the remaining 29–30% of cervical cancers (WHO/ICO, 2010). HPV-18 causes glandular cancers, adenocarcinoma, and adenosquamous carcinoma more frequently than squamous cell carcinoma (B32 vs. 8%, respectively) (De Sanjose et al., 2010). Infection with a high-risk HPV type is considered necessary for the development of cervical cancer but by itself is not sufficient to cause cancer because the vast majority of women with HPV infection do not develop cancer.
Human papillomavirus genome organization The HPV genome can be divided, in general, into three major functional regions: early, late, and long control (noncoding) regions. Two major viral promoters induce transcription of polycistronic mRNAs. The major early promoter referred to as either p97 or p105 in the HPV-16/31 or HPV-18 subtypes, respectively, is located just upstream of the E6 ORF (Longworth and Laimins, 2004). The major late promoter, generally referred to as p742, is located further downstream and varies slightly depending on the virus type (Longworth and Laimins, 2004). The early genes of papillomavirus constitute over 50% of the viral genome sequence from its 50 half and encode six early ORFs (E1, E2, E4, E5, E6, and E7) (Danos et al., 1982) that are involved in the proliferation
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208
European Journal of Cancer Prevention 2014, Vol 23 No 3
Fig. 1
Cervical cancer by HPV type in developing countries
Cervical cancer by HPV type 1.2%
3.4% 3.6% 4.4% 4.7%
1.3% 1.3 1.9%
HPV-16
0.8% 3.5%
HPV-58 HPV-33
5.1%
HPV-16
4.3%
HPV-18 HPV-58
5.2%
HPV-45 54.4%
HPV-31 HPV-52
5.1%
1.7% 2% 1.6% 3.2%
HPV-18
16.5%
HPV-33
7.1%
54.7% 16.3%
HPV-35
HPV-45 HPV-52 HPV-31
HPV-39
HPV-35
HPV-59
HPV-59
HPV-51 HPV-56
HPV-39
Others Cervical cancer by HPV type in developed countries 1.6% 0.8% 1.8% 0.8% 1.9% 4% 4.2%
3.3% HPV-16 HPV-18 HPV-33
16.7%
54.1%
HPV-31 HPV-45 HPV-52 HPV-58 HPV-35 HPV-39 HPV-73
Prevalence of human papillomavirus (HPV) genotypes in invasive cancers by different regions of the world. Source: WHO/ICO (2010).
of infected cells, their lateral expansion, and oncogenesis. The late region of papillomaviruses lies downstream of the early region and encodes L1, the major capsid protein, and L2, the minor capsid protein occupying almost 40% of the virus genome. The long control region, a segment of about 850 bp (10% of the HPV genome), has no protein-coding functions but bears the origin of replication as well as enhancer and silencer sequences that are important in the regulation of RNA polymerase IIinitiated transcription from viral early and late promoters and thus regulate DNA replication (Fig. 2). The E1 and E2 genes encode the proteins that activate viral replication by interacting with specific sequences in the HPV origin of replication along with other cellular DNA replication factors. E2 also controls the function of E6 and
E7 genes. The E4 protein product is required for the maturation of virions and their release from differentiated keratinocytes (Brown et al., 1994). The E5 protein when expressed alone has weak transforming properties and is believed to augment the oncogenic function of E6 and E7 and contribute to tumor progression (DiMaio and Mattoon, 2001). The E6 gene codes for proteins that inhibit negative regulators of the cell cycle and further inhibit p53, which is a transcription factor for apoptosis. The E7 gene codes for viral proteins that bind to retinoblastoma tumor-suppressor proteins, thereby permitting the cell to progress through the cell cycle in the absence of normal mitogenic signals (Table 1). Many more gene transcripts such as E6*I, which is the truncated E6 protein, exist. Their precise biological function in cervical carcinogenesis is unclear; however, it has recently been found that E6*I has a novel function in
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Epidemiology, pathogenesis, and prevention of HPV Asiaf et al.
209
Fig. 2
Transforming proteins
Regulatory noncoding region LCR
E6
E7
p97 p105
L1 Major capsid protein
10%
HPV-16 7905 bp
40%
50%
E1 Viral DNA replication
E2 L2 Minor capsid protein
p742
Replication and E4 transcription regulation Assembly and release E5 Membrane signaling protein The viral genome of HPV is a double stranded circular DNA of approximately 8 kb in size with eight ORFs which possess the ability to encode proteins and the long control region which regulates transcription and replication of HPV. High-risk HPV contain two promoters, p97 and p742, encoding early and late genes. HPV, human papillomavirus; ORF, open reading frame.
Table 1
Human papillomavirus genes and their functions
Human papillomavirus gene
Function
E2 and E1
Viral replication, regulation of E6 and E7 expression by E2, autoregulation of E1 and E2 function by E2 E5 Tumor progression and augmentation of the oncogenic function of E6 and E7 E6 P53 degradation, inhibition of apoptosis (inhibition of Baax and Bak), cell cycle progression, cell transformation, and immune dysregulation E7 pRB inhibition, cell cycle progression, cell transformation, immune dysregulation E4 Maturation of virions and facilitation of their release L1 Major viral capsid protein, antigenicity L2 Minor viral capsid protein, antigenicity Long control region Starting point of DNA replication
upregulating the expression of aldo-keto reductases (AKR1C3) in cervical cancer cell lines, which has been implicated in drug resistance (Wanichwatanadecha et al., 2012). The role of E5, E6, and E7 oncogenes in carcinogenesis is discussed in detail in the Pathogenesis of oncogenic human papillomavirus section.
Immune response against human papillomavirus Humoral immunity
The immune response against HPV infection is modest as there is no blood-borne phase of infection. The life cycle
of HPVs is exclusively intraepithelial, and the free virus particles are shed from the surface of squamous epithelia, which limits their access to vascular and lymphatic channels and hence to lymph nodes where immune responses are initiated. The level and effectiveness of this humoral antibody response are dependent on both the viral load and viral persistence (Ho et al., 2004). The established viral infection is not cleared, although the antibodies are present for a long duration (Shah et al., 1997). Detection of serum HPV antibodies is an important tool in epidemiological studies to assess past exposure (Dillner et al., 1996; Dillner et al., 1997; Dillner, 1999). Virus-neutralizing antibodies are generated against both viral capsid proteins; however, they differ in their levels as well in their potency against HPV infection and cross-reactivity. Anti-L1 antibodies are generated against the epitopes at the surface of papillomavirus and are type-specific (Hines et al., 1994; Roden et al., 1996). The L2 protein lies deeper within the capsid and only a small segment is exposed to the outer surface. Anti-L2 antibodies are generated against this exposed segment (Christensen et al., 1991; Kawana et al., 1999; Roden et al., 2000). These anti-L2 antibodies are less effective than anti-L1 antibodies and show some cross-reactivity against heterologous HPV types (Greenstone et al., 1998; White et al., 1999; Roden et al., 2000). In individuals with
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210 European Journal of Cancer Prevention 2014, Vol 23 No 3
persistent HPV infection, that is, detection of HPV DNA of the same type on two occasions at least 6 months apart, seroconversion occurs most frequently between 6 and 18 months after the first detection of HPV DNA (Wideroff et al., 1995; Carter et al., 1996) and rarely in individuals with incident HPV infections, that is, detection of HPV DNA on only one occasion (Carter et al., 2000). Many practical issues make it difficult to assess whether individuals become immune to type-specific reinfections. Contradictory results have been found for protection against reinfections – some studies have found that protection against reinfection is associated with the presence of HPV antibodies (Konya and Dillner, 2001) and others have shown that antibodies elicited by natural infection with a specific HPV type do not confer protection, as seropositivity is not significantly associated with a reduction in reinfection with homologous types (Viscidi et al., 2004). Cell-mediated immunity
Clearance of a naturally acquired HPV infection is triggered by a specific cell-mediated immune response and requires the interaction of viral epitopes with histocompatibility class I molecules (Ostor, 1993). The evidence for the critical importance of the CD4 T-cellregulated cell-mediated immune response in the resolution and control of HPV infections comes from the analysis of HPV infections in HIV-positive women (Palefsky et al., 2006). HIV-infected patients show multiple HPV infections (Levi et al., 2004), multiple recurrences of cervical HPV infections, increased incidence of cervical neoplasia, and persistence of HPV infections (Ahdieh et al., 2001). Antigen-presenting cells such as dendritic cells or Langerhans cells present in the cervical epithelium play a significant role in recognizing HPVinfected cells and stimulating Th1 helper cells, which in turn produce important cytokines for the production of cytotoxic T lymphocytes (Niedergang et al., 2004). These cytotoxic effector cells attack the infected cells, resulting in resolution of the infection (Stern, 2004).
Epidemiology of human papillomavirus infections Genital HPV is acquired through sexual and genital skinto-skin contact. HPV is currently one of the most common sexually transmitted infections in both men and women and a significant source of morbidity and mortality worldwide (Baseman and Koutsky, 2005; Ebrahim et al., 2005; Dunne et al., 2007). It is estimated that B75% of sexually active adults will encounter at least one HPV infection during their lifetime (Koutsky et al., 1988; Baseman and Koutsky, 2005). The WHO/Catalan Institute of Oncology (ICO) Information Centre on HPV and Cervical Cancer reports that at a given point of time about 11.4% of women worldwide with a normal cytology are positive for HPV DNA. HPV prevalence is higher in the less developed regions (14.3%) compared with the more developed regions
of the world (10.3%). African women (21.3%), in particular women in Eastern Africa, have the highest prevalence (33.6%) of HPV infections (WHO/ICO, 2010). According to a meta-analysis that included data from more than one million women across 59 countries with a normal cervical cytology, the global prevalence of genital HPV was found to be 11.7% and the country-specific adjusted HPV prevalences ranged from 1.6 to 41.9%. The most common HPV types were the oncogenic types, namely, types 16, 18, 52, 31, 58, 39, 51, and 56. HPV-31 was more frequently observed in Europe and Latin America but was much less common in Northern America and Asia, where the most common type was HPV-52 (Fig. 3). HPV-45 was the rarest, usually ranking last among the oncogenic types. A higher HPV prevalence was observed in the African and Latin American regions in comparison with the European, North American, and Asian regions (Bruni et al., 2010). In a recent cross-sectional meta-analysis of high-risk HPV type distribution among 115 789 HPV-positive women [including women with a normal cytology, atypical squamous cells of undetermined significance (ASCUS), LSIL and HSIL diagnosed cytologically, CIN grade 1 (CIN1), CIN2, and CIN3 diagnosed histologically, and invasive cervical cancers], from 423 PCR-based studies worldwide, no strong differences in HPV type distribution were found among women with a normal cytology, ASCUS, LSIL, or CIN1. However, the HPV-16 positivity increased steeply from normal/ASCUS/LSIL/CIN1 (20–28%) through CIN2/HSIL (40/47%) to CIN3/ICC (58/63%) (Guan et al., 2012). The relationship between age and HPV prevalence was seen to be inverse in many countries; however, in some of the poorest areas studied, the HPV prevalence was found to be uniformly high across all age groups (Franceschi et al., 2006). In some countries, cross-sectional and cohort studies have shown an interesting trend in the female age-specific distribution of HPV, whereby there is a first peak at a younger age (r 25 years) and a clear second peak among individuals aged 45 years or older (Munoz et al., 2004; Herrero et al., 2005; Franceschi et al., 2006; De Sanjose et al., 2007; Smith et al., 2008; Asiaf et al., 2012). The rationale behind the first peak, which comes soon after the onset of sexual relations, is higher levels of sexual activity with multiple partners and a low viral immunity. The reasons for the smaller second peak during middle age are still to be deciphered; the probable explanations include immunosenescence, hormonal changes before menopause, reactivation of latent infections, or perhaps the higher rates of HPV persistence at older ages compared with new HPV acquisition, partly at the expense of infections with low-risk types (Castle et al., 2005; De Sanjose et al., 2007; Bruni et al., 2010; Gonzalez et al., 2010). HPV not only affects women, but also commonly infects sexually active men. In a study of men, Giuliano et al.
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Epidemiology, pathogenesis, and prevention of HPV Asiaf et al. 211
Fig. 3
HPV types grouped by their specific prevalence
HPV types grouped by their specific prevalence
European continent (N =129 646) 73, 44, 90, 72, 62, 69, 54 42, 81, 68, 83, 59, 61, 35 53, 56, 70, 11 45, 52, 51, 58 6 33, 66 39 18 31 16
Latin America and Caribbean (N =18 248) 89, 71, 73, 40, 72 84, 59, 62, 68, 42, 39, 56, 83, 66, 81 53, 51, 54, 70, 61 52, 11, 35, 45 33 6 58 31, 18 16
0.1 0.2 0.3 0.4 0.5 0.6 0.8 0.9 2.3 4.8 0
1
2
3
4
5
0.2 0.4 0.5 0.7 0.8 0.9 1 1.2 3.3 0
6
African continent (N = 5 872) 54, 68 44 33, 39 81, 42, 56, 53 66 58 45 31, 35, 18 52 16
3.5 2
3
4
2.5 0
73, 43, 74, 62, 83, 11 68, 59, 70, 84, 72 81, 42, 44, 35, 54 6, 33, 66, 45 39, 56, 53, 51 58 31 52 18 16
2 2.1 2.3 5.8 1
0.5
1
1.5
2
2.5
3
World (N =215 568)
0.7 1 1.1 1.2 1.3 1.5
0
4
1.4
North America (N =11 874) 62, 83 45, 31, 56, 84 54 39, 53, 59 66 58, 51 6 52 18 16
3
Review article
Review of the current knowledge on the epidemiology, pathogenesis, and prevention of human papillomavirus infection Asia Asiafa, Shiekh T. Ahmadb, Sheikh O. Mohammadc and Mohammad A. Zargara Human papillomavirus (HPV) infection is a central and necessary, although not sufficient, cause of cervical cancer. Besides HPV, the additional multiple risk factors related with the onset of cervical cancer are early-age sexual activities; high number of sexual partners, which is the most salient risk factor; suppression and alteration of the immune status; long-term use of oral contraceptives; and other hormonal influences. The tumor-suppressor proteins p53 and pRb are degraded and destabilized through ubiquitination by viral oncoproteins E6 and E7. Over 95% of cervical cancer cases worldwide test positive for oncogenic HPV DNA. Although cervical screening procedures have been successful in reducing the disease burden associated with HPV infection because of lack of resources or inadequate infrastructure many countries have failed to reduce cervical cancer mortality. Therefore, prevention may be a valuable strategy for reducing the economic and disease burden of HPV infection. At present, two successful prophylactic HPV vaccines are available, quadrivalent (HPV16/18/6/11) ‘Gardasil’ and bivalent (HPV16/18) ‘Cervarix’ for vaccinating young adolescent girls at or before the onset of puberty. Recent data indicate that vaccination prevents the development of cervical lesions in women who have not already acquired the vaccine-specific HPV types. Moreover, several therapeutic vaccines that are protein/peptide-based, DNA-based,
or cell-based are in clinical trials but are yet to establish their efficacy; these vaccines are likely to provide important future health benefits. The therapeutic vaccination mode of prevention is a promising area of research, as revealed in preclinical trials; however, clinical trials based on large populations are warranted before reaching a valid conclusion. This review summarizes the studies on the epidemiology of HPV infection, the pathogenesis of viral oncoproteins in the oncogenesis of cervical cancer, the economic and health burden of HPV-related diseases, and, finally, focuses on the results of recent clinical vaccination trials. European Journal of Cancer c 2014 Wolters Kluwer Health | Prevention 23:206–224 Lippincott Williams & Wilkins.
Introduction
activities in the field. The main contribution to this was made by the German scientist Harald Zur Hausen, who for the first time showed the robust association between HPV infections and cervical cancer by identifying, cloning, and sequencing the two most important high-risk HPV types, that is, 16 and 18, from cervical tumor specimens (Zur Hausen, 1991, 1996). Subsequent research established that HPV is also associated with a substantial number of oral, esophageal, and other anogenital cancers, namely, cancers of the vulva, vagina, penis, and anus, in addition to the development of cervical cancer (Kashima and Mounts, 1987; Greenspan et al., 1988; Zur Hausen, 1989; Syrjanen, 2002; Dillner et al., 2007). Epidemiological case series have shown that HPV DNA is found in nearly 100% of cervical cancer cases (Walboomers et al., 1999). At present, there are neither effective methods for preventing HPV infection nor promising treatments available for the management of HPV-associated diseases: for
Human papillomaviruses (HPVs) are a group of small, nonenveloped, double-stranded DNA viruses belonging to the family Papovaviridae. This group of viruses infects a variety of organisms across the animal kingdom, ranging from birds to mammals, including humans, and the viruses are specific for their respective hosts. These viruses appear to show similarities with polyomaviruses and consist of about 72 capsomers. The circular HPV genome comprises 7500–8000 bp of DNA that is associated with histones and is compacted into aggregates similar to chromatin (Fuchs et al., 2006). HPV is known to be the main causative agent for cervical carcinomas. In early 1976, it was discovered that novel HPV types are involved in the successful development of cervical tumors, and, since then, various epidemiological, molecular, and biological studies have justified this observation. This led to immense enhancement in research c 2014 Wolters Kluwer Health | Lippincott Williams & Wilkins 0959-8278
European Journal of Cancer Prevention 2014, 23:206–224 Keywords: cervical cancer, dysplasia, E6, E7, human papillomavirus, vaccine a Department of Biochemistry, Faculty of Biological Science, University of Kashmir, Srinagar, bInstitute for Stem Cell Biology & Regenerative Medicine, NCBS (TIFR) Campus, Bangalore and cDepartment of Biochemistry and Molecular Biology, School of life Sciences, Pondicherry University, Puducherry, India
Correspondence to Mohammad A. Zargar, PhD, Department of Biochemistry, Faculty of Biological Science, University of Kashmir, Srinagar 190006, India Tel: + 91 194 242 8723; fax: + 91 194 242 1353; e-mail: [email protected] Received 4 April 2013 Accepted 25 June 2013
DOI: 10.1097/CEJ.0b013e328364f273
Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.
Epidemiology, pathogenesis, and prevention of HPV Asiaf et al. 207
preinvasive lesions such as genital warts and cytological abnormalities, the treatment consists of destruction of tissue through electrocoagulation (use of intense heat generated by electric current), cryotherapy (use of extreme cold), laser ablation, or local surgery, whereas invasive cervical cancers are generally managed with surgery, radiation, both surgery and radiation, and in some cases chemotherapy and observation for recurrence. However, these therapies are effective only when the disease is localized, as in the case of cervical intraepithelial neoplasia (CIN); the possibility of recurrence remains, as treatment does not always eradicate the underlying HPV infection. Prophylactic HPV vaccines (Gardasil and Cervarix), which are constructed using virus-like particles, prevent the acquisition of low-grade and high-grade squamous intraepithelial lesions (LSIL and HSIL) of the cervix by generating strong neutralizing antibodies against capsid antigens L1 in recipients; these antibodies likely mediate the observed protection against HPV infections (McElrath et al., 2008). However, in many HPV-associated neoplasias, detectable levels of capsid antigens (L1 or L2) are not expressed by infected cells. Therefore, current prophylactic vaccines are not likely to be effective in preventing disease progression in preexisting HPV infections or HPV-associated lesions, and there is a burning need to develop therapeutic HPV vaccines. Successful immunotherapy might therefore be a preferred mode of treatment because it can target all HPV-associated lesions irrespective of their location and can induce long-lasting immunity, thus preventing recurrence. Therapeutic cancer vaccines are expected to treat an existing cancer by enhancing the naturally occurring immune response against the cancer.
Low-risk and high-risk human papillomaviruses To date, more than 120 HPV types have been isolated, completely sequenced, and classified broadly into mucosal/genital and cutaneous types based on sequence analyses and clinical manifestations (Bernard et al., 2005). HPVs have been phylogenetically categorized into five genera: alpha, beta, gamma, delta, and mu based on the differences in their nucleotide sequences (De Villiers et al., 2004). The mucosal/genital HPV types belong to the alpha-PV genus and can be divided into nononcogenic (low-risk) or potentially oncogenic (high-risk) types according to their presence in malignant lesions (Bosch et al., 2002; Munoz et al., 2003). Fifteen HPVs have been classified as high-risk types (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, and 82) that cause dysplasia and cancer, 12 have been classified as low-risk types (6, 11, 40, 42, 43, 44, 54, 61, 70, 72, 81, and CP6108) that usually cause low-grade mild dysplasia, genital warts, and respiratory papillomatosis, and three have been classified as probable high-risk types (26, 53, and 66) (Munoz et al., 2003). HPVs 31, 33, 35, 51, and 52 are sometimes
regarded as ‘intermediate-risk’ viruses because they are more common in mild or severe dysplastic lesions than in carcinomas (Fernandes et al., 2009). Cutaneous HPVs constitute more than 75% of the HPVs described to date and are grouped into five different genera: alpha-PV, betaPV, gamma-PV, mu-PV, and nu-PV (De Villiers et al., 2004) that are not generally associated with cancers. Certain beta types have been implicated in the development of nonmelanoma skin cancers in immunosuppressed individuals and epidermodysplasia verruciformis (EV) patients. Their possible role in cancer progression in the general population is currently unresolved (Doorbar et al., 2012). Each genotype is characterized on the basis of a difference of more than 10% in the DNA sequence of the L1 open reading frame (ORF) from the closest known HPV type. If a DNA difference of 2–10% exists, the two viruses are considered subtypes of the same HPV type. Variants are defined if the sequences of their L1 genes are at a maximum 2% dissimilar from one another (De Villiers et al., 2004; Chow et al., 2010). Before the development of full-fledged cervical carcinoma, CIN appears, which is the consequence of persistent cervical infection with a highrisk HPV genotype with specific symptoms until its progression into a tumor (Ho et al., 1995). Among all the HPV types, type 16 is the most carcinogenic HPV genotype (Munoz et al., 2003; De Sanjose et al., 2010) and is responsible for B54.4% of all cervical cancers. The minor differences between the developing and developed regions of the world are shown in Fig. 1. HPV-18 is the next most carcinogenic HPV type and accounts for 16.5% of cervical cancers. Approximately 17 other HPV genotypes cause the remaining 29–30% of cervical cancers (WHO/ICO, 2010). HPV-18 causes glandular cancers, adenocarcinoma, and adenosquamous carcinoma more frequently than squamous cell carcinoma (B32 vs. 8%, respectively) (De Sanjose et al., 2010). Infection with a high-risk HPV type is considered necessary for the development of cervical cancer but by itself is not sufficient to cause cancer because the vast majority of women with HPV infection do not develop cancer.
Human papillomavirus genome organization The HPV genome can be divided, in general, into three major functional regions: early, late, and long control (noncoding) regions. Two major viral promoters induce transcription of polycistronic mRNAs. The major early promoter referred to as either p97 or p105 in the HPV-16/31 or HPV-18 subtypes, respectively, is located just upstream of the E6 ORF (Longworth and Laimins, 2004). The major late promoter, generally referred to as p742, is located further downstream and varies slightly depending on the virus type (Longworth and Laimins, 2004). The early genes of papillomavirus constitute over 50% of the viral genome sequence from its 50 half and encode six early ORFs (E1, E2, E4, E5, E6, and E7) (Danos et al., 1982) that are involved in the proliferation
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European Journal of Cancer Prevention 2014, Vol 23 No 3
Fig. 1
Cervical cancer by HPV type in developing countries
Cervical cancer by HPV type 1.2%
3.4% 3.6% 4.4% 4.7%
1.3% 1.3 1.9%
HPV-16
0.8% 3.5%
HPV-58 HPV-33
5.1%
HPV-16
4.3%
HPV-18 HPV-58
5.2%
HPV-45 54.4%
HPV-31 HPV-52
5.1%
1.7% 2% 1.6% 3.2%
HPV-18
16.5%
HPV-33
7.1%
54.7% 16.3%
HPV-35
HPV-45 HPV-52 HPV-31
HPV-39
HPV-35
HPV-59
HPV-59
HPV-51 HPV-56
HPV-39
Others Cervical cancer by HPV type in developed countries 1.6% 0.8% 1.8% 0.8% 1.9% 4% 4.2%
3.3% HPV-16 HPV-18 HPV-33
16.7%
54.1%
HPV-31 HPV-45 HPV-52 HPV-58 HPV-35 HPV-39 HPV-73
Prevalence of human papillomavirus (HPV) genotypes in invasive cancers by different regions of the world. Source: WHO/ICO (2010).
of infected cells, their lateral expansion, and oncogenesis. The late region of papillomaviruses lies downstream of the early region and encodes L1, the major capsid protein, and L2, the minor capsid protein occupying almost 40% of the virus genome. The long control region, a segment of about 850 bp (10% of the HPV genome), has no protein-coding functions but bears the origin of replication as well as enhancer and silencer sequences that are important in the regulation of RNA polymerase IIinitiated transcription from viral early and late promoters and thus regulate DNA replication (Fig. 2). The E1 and E2 genes encode the proteins that activate viral replication by interacting with specific sequences in the HPV origin of replication along with other cellular DNA replication factors. E2 also controls the function of E6 and
E7 genes. The E4 protein product is required for the maturation of virions and their release from differentiated keratinocytes (Brown et al., 1994). The E5 protein when expressed alone has weak transforming properties and is believed to augment the oncogenic function of E6 and E7 and contribute to tumor progression (DiMaio and Mattoon, 2001). The E6 gene codes for proteins that inhibit negative regulators of the cell cycle and further inhibit p53, which is a transcription factor for apoptosis. The E7 gene codes for viral proteins that bind to retinoblastoma tumor-suppressor proteins, thereby permitting the cell to progress through the cell cycle in the absence of normal mitogenic signals (Table 1). Many more gene transcripts such as E6*I, which is the truncated E6 protein, exist. Their precise biological function in cervical carcinogenesis is unclear; however, it has recently been found that E6*I has a novel function in
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Epidemiology, pathogenesis, and prevention of HPV Asiaf et al.
209
Fig. 2
Transforming proteins
Regulatory noncoding region LCR
E6
E7
p97 p105
L1 Major capsid protein
10%
HPV-16 7905 bp
40%
50%
E1 Viral DNA replication
E2 L2 Minor capsid protein
p742
Replication and E4 transcription regulation Assembly and release E5 Membrane signaling protein The viral genome of HPV is a double stranded circular DNA of approximately 8 kb in size with eight ORFs which possess the ability to encode proteins and the long control region which regulates transcription and replication of HPV. High-risk HPV contain two promoters, p97 and p742, encoding early and late genes. HPV, human papillomavirus; ORF, open reading frame.
Table 1
Human papillomavirus genes and their functions
Human papillomavirus gene
Function
E2 and E1
Viral replication, regulation of E6 and E7 expression by E2, autoregulation of E1 and E2 function by E2 E5 Tumor progression and augmentation of the oncogenic function of E6 and E7 E6 P53 degradation, inhibition of apoptosis (inhibition of Baax and Bak), cell cycle progression, cell transformation, and immune dysregulation E7 pRB inhibition, cell cycle progression, cell transformation, immune dysregulation E4 Maturation of virions and facilitation of their release L1 Major viral capsid protein, antigenicity L2 Minor viral capsid protein, antigenicity Long control region Starting point of DNA replication
upregulating the expression of aldo-keto reductases (AKR1C3) in cervical cancer cell lines, which has been implicated in drug resistance (Wanichwatanadecha et al., 2012). The role of E5, E6, and E7 oncogenes in carcinogenesis is discussed in detail in the Pathogenesis of oncogenic human papillomavirus section.
Immune response against human papillomavirus Humoral immunity
The immune response against HPV infection is modest as there is no blood-borne phase of infection. The life cycle
of HPVs is exclusively intraepithelial, and the free virus particles are shed from the surface of squamous epithelia, which limits their access to vascular and lymphatic channels and hence to lymph nodes where immune responses are initiated. The level and effectiveness of this humoral antibody response are dependent on both the viral load and viral persistence (Ho et al., 2004). The established viral infection is not cleared, although the antibodies are present for a long duration (Shah et al., 1997). Detection of serum HPV antibodies is an important tool in epidemiological studies to assess past exposure (Dillner et al., 1996; Dillner et al., 1997; Dillner, 1999). Virus-neutralizing antibodies are generated against both viral capsid proteins; however, they differ in their levels as well in their potency against HPV infection and cross-reactivity. Anti-L1 antibodies are generated against the epitopes at the surface of papillomavirus and are type-specific (Hines et al., 1994; Roden et al., 1996). The L2 protein lies deeper within the capsid and only a small segment is exposed to the outer surface. Anti-L2 antibodies are generated against this exposed segment (Christensen et al., 1991; Kawana et al., 1999; Roden et al., 2000). These anti-L2 antibodies are less effective than anti-L1 antibodies and show some cross-reactivity against heterologous HPV types (Greenstone et al., 1998; White et al., 1999; Roden et al., 2000). In individuals with
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210 European Journal of Cancer Prevention 2014, Vol 23 No 3
persistent HPV infection, that is, detection of HPV DNA of the same type on two occasions at least 6 months apart, seroconversion occurs most frequently between 6 and 18 months after the first detection of HPV DNA (Wideroff et al., 1995; Carter et al., 1996) and rarely in individuals with incident HPV infections, that is, detection of HPV DNA on only one occasion (Carter et al., 2000). Many practical issues make it difficult to assess whether individuals become immune to type-specific reinfections. Contradictory results have been found for protection against reinfections – some studies have found that protection against reinfection is associated with the presence of HPV antibodies (Konya and Dillner, 2001) and others have shown that antibodies elicited by natural infection with a specific HPV type do not confer protection, as seropositivity is not significantly associated with a reduction in reinfection with homologous types (Viscidi et al., 2004). Cell-mediated immunity
Clearance of a naturally acquired HPV infection is triggered by a specific cell-mediated immune response and requires the interaction of viral epitopes with histocompatibility class I molecules (Ostor, 1993). The evidence for the critical importance of the CD4 T-cellregulated cell-mediated immune response in the resolution and control of HPV infections comes from the analysis of HPV infections in HIV-positive women (Palefsky et al., 2006). HIV-infected patients show multiple HPV infections (Levi et al., 2004), multiple recurrences of cervical HPV infections, increased incidence of cervical neoplasia, and persistence of HPV infections (Ahdieh et al., 2001). Antigen-presenting cells such as dendritic cells or Langerhans cells present in the cervical epithelium play a significant role in recognizing HPVinfected cells and stimulating Th1 helper cells, which in turn produce important cytokines for the production of cytotoxic T lymphocytes (Niedergang et al., 2004). These cytotoxic effector cells attack the infected cells, resulting in resolution of the infection (Stern, 2004).
Epidemiology of human papillomavirus infections Genital HPV is acquired through sexual and genital skinto-skin contact. HPV is currently one of the most common sexually transmitted infections in both men and women and a significant source of morbidity and mortality worldwide (Baseman and Koutsky, 2005; Ebrahim et al., 2005; Dunne et al., 2007). It is estimated that B75% of sexually active adults will encounter at least one HPV infection during their lifetime (Koutsky et al., 1988; Baseman and Koutsky, 2005). The WHO/Catalan Institute of Oncology (ICO) Information Centre on HPV and Cervical Cancer reports that at a given point of time about 11.4% of women worldwide with a normal cytology are positive for HPV DNA. HPV prevalence is higher in the less developed regions (14.3%) compared with the more developed regions
of the world (10.3%). African women (21.3%), in particular women in Eastern Africa, have the highest prevalence (33.6%) of HPV infections (WHO/ICO, 2010). According to a meta-analysis that included data from more than one million women across 59 countries with a normal cervical cytology, the global prevalence of genital HPV was found to be 11.7% and the country-specific adjusted HPV prevalences ranged from 1.6 to 41.9%. The most common HPV types were the oncogenic types, namely, types 16, 18, 52, 31, 58, 39, 51, and 56. HPV-31 was more frequently observed in Europe and Latin America but was much less common in Northern America and Asia, where the most common type was HPV-52 (Fig. 3). HPV-45 was the rarest, usually ranking last among the oncogenic types. A higher HPV prevalence was observed in the African and Latin American regions in comparison with the European, North American, and Asian regions (Bruni et al., 2010). In a recent cross-sectional meta-analysis of high-risk HPV type distribution among 115 789 HPV-positive women [including women with a normal cytology, atypical squamous cells of undetermined significance (ASCUS), LSIL and HSIL diagnosed cytologically, CIN grade 1 (CIN1), CIN2, and CIN3 diagnosed histologically, and invasive cervical cancers], from 423 PCR-based studies worldwide, no strong differences in HPV type distribution were found among women with a normal cytology, ASCUS, LSIL, or CIN1. However, the HPV-16 positivity increased steeply from normal/ASCUS/LSIL/CIN1 (20–28%) through CIN2/HSIL (40/47%) to CIN3/ICC (58/63%) (Guan et al., 2012). The relationship between age and HPV prevalence was seen to be inverse in many countries; however, in some of the poorest areas studied, the HPV prevalence was found to be uniformly high across all age groups (Franceschi et al., 2006). In some countries, cross-sectional and cohort studies have shown an interesting trend in the female age-specific distribution of HPV, whereby there is a first peak at a younger age (r 25 years) and a clear second peak among individuals aged 45 years or older (Munoz et al., 2004; Herrero et al., 2005; Franceschi et al., 2006; De Sanjose et al., 2007; Smith et al., 2008; Asiaf et al., 2012). The rationale behind the first peak, which comes soon after the onset of sexual relations, is higher levels of sexual activity with multiple partners and a low viral immunity. The reasons for the smaller second peak during middle age are still to be deciphered; the probable explanations include immunosenescence, hormonal changes before menopause, reactivation of latent infections, or perhaps the higher rates of HPV persistence at older ages compared with new HPV acquisition, partly at the expense of infections with low-risk types (Castle et al., 2005; De Sanjose et al., 2007; Bruni et al., 2010; Gonzalez et al., 2010). HPV not only affects women, but also commonly infects sexually active men. In a study of men, Giuliano et al.
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Epidemiology, pathogenesis, and prevention of HPV Asiaf et al. 211
Fig. 3
HPV types grouped by their specific prevalence
HPV types grouped by their specific prevalence
European continent (N =129 646) 73, 44, 90, 72, 62, 69, 54 42, 81, 68, 83, 59, 61, 35 53, 56, 70, 11 45, 52, 51, 58 6 33, 66 39 18 31 16
Latin America and Caribbean (N =18 248) 89, 71, 73, 40, 72 84, 59, 62, 68, 42, 39, 56, 83, 66, 81 53, 51, 54, 70, 61 52, 11, 35, 45 33 6 58 31, 18 16
0.1 0.2 0.3 0.4 0.5 0.6 0.8 0.9 2.3 4.8 0
1
2
3
4
5
0.2 0.4 0.5 0.7 0.8 0.9 1 1.2 3.3 0
6
African continent (N = 5 872) 54, 68 44 33, 39 81, 42, 56, 53 66 58 45 31, 35, 18 52 16
3.5 2
3
4
2.5 0
73, 43, 74, 62, 83, 11 68, 59, 70, 84, 72 81, 42, 44, 35, 54 6, 33, 66, 45 39, 56, 53, 51 58 31 52 18 16
2 2.1 2.3 5.8 1
0.5
1
1.5
2
2.5
3
World (N =215 568)
0.7 1 1.1 1.2 1.3 1.5
0
4
1.4
North America (N =11 874) 62, 83 45, 31, 56, 84 54 39, 53, 59 66 58, 51 6 52 18 16
3
