Pathogenesis of Papillomaviruses Infection

C. Pathogenesis

1. Clinical Manifestations

  1. Benign warts - HPVs are responsible for the numerous types of warts such as plantar/common hand warts, juvenile/flat warts, genital warts (condylomata accuminata), macular lesions in immunosuppressed patients and patients with EV, and various premalignant lesions e.g. CIN1, CIN2 etc. Uncommon presentations include conjunctival warts that may be occupational hazards to obstetricians and gynaecologists.
  2. Juvenile laryngeal papillomatosis - laryngeal warts are usually caused by types 6 and 11 which have a tropism for mucosal surfaces. (The same types infect genital areas as well). It is considered to be a life-threatening condition in children because of the danger of airways obstruction. Repeated surgery of laser treatment may be required. There have been documented cases where laryngeal carcinomas developed when laryngeal papillomas were treated by X-radiotherapy.
  3. Epidermodysplasia verruciformis - EV is a rare autosomal recessive condition characterized by the appearance of widespread macular lesions and warts on the body. Some of the HPV types are not normally found in normal people. One-third of EV patients develop squamous cell carcinoma, usually in areas exposed to sunlight.

2. Association with Cervical Cancer

The association between certain strains of HPV and cervical cancer is well known. In the West, cervical cancer is the fourth most common cancer in women. In developing countries though, it is the most common cancer, with 1/2 million new cases per year. Overall, it is the most common virus associated cancer worldwide, followed by hepatocellular carcinoma, NPC and Burkitt's Lymphoma. There is now compelling evidence for an aetiological role of HPV in cervical and other cancers.

a. Epidemiological Evidence ;- it has been known for a long time that the incidence of cervical Ca. is related to sexual activity. Promiscuous women are far more likely to develop cervical Ca. Nuns rarely get cervical Ca. Several sexually transmitted agents have been implicated including Herpes Simplex. It is now fair to say that evidence of a role for HSV is very slim and now very much discounted.

b. Virological Evidence ;-

  1. It has now been proven that certain precancerous conditions for cervical and vulval Ca, namely CIN and VIN, are caused by HPV. Normal human epithelium from the cervix and epithelium purposefully exposed to HPV 16 were transplanted to nude mice. (immunologically immature mice which are not able to reject the graft). Weeks later, lesions that resemble CIN histologically were detected in the epithelium which was infected by HPV.
  2. HPV genomes can be detected in 90 - 95% of tumour cell lines. HPV genomes can be found at a lower frequency in pre cancerous state such as CIN1, CIN2 and CIN3. HPV 16 and 18 are the most commonly found. However, other strains may be present, especially in the less dysplastic lesions. Types 6 and 11 are commonly present in the less dysplastic lesions but not in the more dysplastic ones.
  3. HPV DNA persists in an episomal form in normal infected cells but is integrated in the host cell chromosome in malignant cells. The integration of HPV DNA appears to be random. It is thought that as integration occurs, the breakage always occurs in the E1 region. One of the functions of the E1 region is to negatively control the expression of the E6 and E6 genes. Therefore, it is thought that as a result of the integration, the E1 function is lost and the E6 and E7 genes are derepressed. This leads to an overproduction of the E6 and E7 gene products. E6 is shown to inactivate p53 and E7 the Rb proteins.
  4. HPV DNA is biologically active in vitro. It is able to transform human fibroblasts and keratinocytes. It has effects on epithelial cell differentiation and induces DNA replication in the fibroblast. All the above effects can be nailed down to the E7 reading frame. The constant expression of the E7 and perhaps the E6 reading frames products are essential for the maintenance of transformation.
  5. Antibodies to HPV are more prevalent in patients with CIN or invasive Ca. Gissman et al. looked at the prevalence of antibodies to the E7 and E4 HPV proteins in normal people and those with Ca.Cervix. Although the prevalence of Abs to E4 and E7 were increased in patients with Ca.Cervix compared to the controls. The prevalence of Ab to E7 was much higher ; There was only a 2 fold difference in the prevalence of Ab to E4 between the cancer patients and the normal controls. However there was a 7-fold difference in the prevalence of Ab to E7. This suggest that there may be a role for testing for Ab to the E7 protein in future for use as a marker in monitoring the effectiveness of treatment. However, it is very important to have the correct protein antigen of the particular strain of HPV involved.

It must be borne in mind that other factors are involved in the tumourigenesis of cervical Ca ;-

  1. The actual site in the body
  2. The HPV type
  3. The immune status of the patient; immunocompromised patients have a much higher risk of developing Ca.Cervix.
  4. Smoking
  5. Hormones
  6. Vitamin A deficiency

3. Association with tumours in immunocompromised individuals

10% of the general population have warts at any one time but warts and skin cancers are much more common and of a greater problem in immunocompromised patients, in particular renal transplants and people with AIDs. The actual number of people undergoing renal transplant operations have increased gradually in the last 20 years. As survival times increased, it became apparent that many of them are developing serious problems with warts and skin cancers. The problem seems to be related to the maintenance immunosuppressive therapy, which is prescribed to patients as long-term therapy after their operation. Prednisolone and Azothiaprine are the 2 most common immunosuppressants given.

Many renal transplant patients appeared with horrific intractable warts in all parts of the body. The warts could not be controlled by any methods but may respond to the withdrawal of the immunosuppressive therapy. Some of these patients with warts progressed onwards to malignancy in many parts of the body at once. Several factors are known to contribute to malignancy ;-

  1. Diminished immunosurveillance
  2. UV exposure
  3. Oncogenic effects of the drugs given
  4. Chronic antigenic stimulation

The longer the graft survives, the more likely that patient will develop warts and skin cancers.

  Warts Kerratoses Cancers
< 5 years 35% 15% 2%
> 5 years 83% 36% 13%

The widespread appearance of warts in renal transplant patients resembles those found in the condition Epidermaldysplasia Verruciformis (EV). EV is a rare autosomal recessive condition characterized by the appearance of widespread warts on the surface of the body and 30% of the development of squamous cell carcinomas (SCC). Over 20 types of HPV are associated with EV and the HPV DNA exists mainly in the episomal form in the lesions of these patients. In renal graft survivors the ratio of basal cell carcinoma to SCC is 1:15 compared to a ratio of 1:5 for a normal population. The female renal transplant patients are also at a much higher risk of developing cervical Ca., CIN and VIN than the normal population. The precise role of HPV in the causation of skin cancers in the renal-transplant patients is not known. The skin lesions do resemble those of EV on appearance and the DNA of HPV 5 and 8 ( the HPV strains commonly isolated from patients with EV ) are found in 50% of SCC of the renal transplant patients. HPV 5 and 8 may act as inducing agents for the development of skin tumours. In a recent study, it was claimed that the prevalence of anti-HPV 8 Ab is increased in patients with skin tumours ;-

Controls 20%
Basal Cell Carcinoma 40%
Squamous Cell Carcinoma 80%

If the above can be substantiated, then HPV may have a far more sinister role in human cancers than previously thought. It may have a role in inducing the development of skin and other epithelial cancers. It is not clear whether a withdrawal of immunosuppressive therapy will have a beneficial effect on skin cancers. Although some patients with intractable cancer problems have been offered a withdrawal of immunosuppressive therapy. There have been no cases of metastasis from a SCC arising in an ex-renal transplant patient to date.  

4. Persistence of HPV infection

There is circumstantial evidence that HPVs may persist in the squamous epithelium, without producing recognizable lesions, in a fashion similar to the persistence of polyomaviruses in the kidney. Up to 42% of allograft recipients develops cutaneous lesions within a year after transplant. This suggests that transplanted patients experience either new infections or reactivation of latent virus. HPV DNA sequences have been found in biopsies of normal areas of the larynx of individuals who have had recurrent episodes of laryngeal papillomas.

D. Laboratory Diagnosis

The appearance of warts vary from the scaly flat lesions on the epithelium of individuals with EV, to the aceto-white flat CIN lesion of the cervix. It may be hard to distinguish wart lesions from other lesions such as molluscum. The methods which are in routine use are:

  1. EM of skin scrapings
  2. Detection of the common HPV antigen in the tissue by IF techniques. This is more sensitive than EM even though the antigen is not detectable in 25% of confirmed warts.

DNA-DNA hybridization and PCR techniques have been developed in research laboratories. The Southern blot is the most sensitive and specific hybridization technique but is labor-intensive and time-consuming. Dot-blot is available in commercial kits and is as sensitive as Southern blot but not as specific, being unable to distinguish between cross-reacting closely related viruses. RNA probes are more sensitive than DNA probes because of the stability of the RNA-RNA or RNA-DNA hybrids compared to DNA-DNA hybrids. Commercial high sensitivity hybridization assays such as the Digene hybrid capture will detect HPV types that are associated with cervical cancers from biopsies of genital lesions. However, the usefulness of these tests in a histopathological setting remains controversial and more research is required.


E. Management and Prevention

Although they are a cosmetic nuisance in most cases, warts are notoriously difficult to treat. The treatments which could be used include cryotherapy with liquid nitrogen, podophyllin (an antimitotic agent), lasers (for mucosal lesions such as CIN), electrodiathermy, and surgery. Interferon has been used successfully to treat recurrent laryngeal warts and CIN. However, the cost of the treatment may prove to be prohibitive. Precancerous CIN-2 and CIN-3 lesions should be treated by lasers, electrodiathermy, or surgery. In the case of immunocompromised patients with intractable warts, improvement is often seen. There are mass screening programs for CIN and cervical cancers in a number of countries. Being one of the most common cancers in women, a vaccine against HPV types associated with cervical cancers would be desirable and research is being carried out in this area.