HPV+and+its+Role+in+Cervical+Cancer

=Purpose =

We will understand and analyze the role of Human Papillomavirus (HPV) in causing cervical cancer, as well as the most effective ways of preventing cervical cancer worldwide.

=Introduction =

In 1975, virologist Harold zur Hausen of the German Cancer Research Center in Heidelberg first suggested a connection between cervical cancer and human papillomavirus. A second report in 1999 confirmed zur Hausen’s theory, finding HPV DNA in 99.7% of cervical cancer cases studied. Cervical cancer has always been a worldwide disease: it is the second most common cancer in women with more than 200,000 deaths per year worldwide (Sanchez). Thus, these findings were exciting and promising to researchers everywhere.


 * Figure 1. Leading cancers in developed versus underdeveloped countries.**

Cervical cancer is significantly less prevalent in developed countries than in underdeveloped countries, largely due to the use of Pap smears. This is because wealthier countries, such as the U.S., can afford to provide this procedure for most of its population. Despite the increased diagnoses from Pap smears, the intricacies of cervical cancer were unknown until the discovery of HPV.

=Overview =

What is Human Papillomavirus (HPV)?
HPV is a retrovirus, and is the most common sexually transmitted infection worldwide. It can infect male and female genitals, as well as the throat and mouth. There are over 100 strains of the virus, however only 40 of these infect the genital area. Certain strains cause genital warts, while other strains, such as HPV 16 and 18, cause cervical cancer. It is possible to be infected with more than one strain of HPV at any given time. In 90% of HPV infection cases, the body's immune system kills the infection within a two year period, with no significant symptoms (CDC). The figure below shows a direct correlation between HPV infection and increased risk of cervical cancer.

 The cumulative risk of cervical cancer development is plotted using Kaplan-Meier methods. Plot A shows women under the age of 30: HPV16 infection, cumulative probability = 14.6 (95% CI = 10.0–20.9) and PPV = 11.1 (8.1–14.0); other carcinogenic types, cumulative probability = 7.0 (4.2–11.4), PPV = 3.2 (2.0–4.3); no carcinogenic types, cumulative probability = 1.8 (1.2–2.5) and PPV = 0.7 (0.4–0.9). Plot B shows women over 30: HPV16 infection, cumulative probability = 8.5 (95% CI = 4.1–17.2) and PPV = 13.8 (8.2–19.4); other carcinogenic types, cumulative probability = 3.1 (1.6–6.1) and PPV = 3.7 (2.0–5.4); no carcinogenic types, cumulative probability = 0.7 (0.5–0.9) and PPV = 0.4 (0.3–0.5).
 * Figure 2. Cumulative probability of cervical cancer based on HPV status.**

What is Cervical Cancer?
Not all strains of HPV cause cancer. Only half of the known types are able to infect mucosal tissues, while the other half generally produce common skin warts. Mucosal HPV's are ones that actually cause cervical cancer, the most common of these being HPV 16 and 18--70% of cervical cancers are caused by the 16 and 18 strain worldwide (World Health Organization). These types infect the epithelial cells lying just under the mucosal surface in the cervix, and produce an oncoprotein called E6 (Cohen). E6 binds p53 and retinoblastoma (Rb) proteins, both tumor suppressors, and renders them inactive. Without Rb, cells are able to proliferate unchecked; without p53, cells can replicate even with damaged DNA. Ultimately, the inactivation of tumor suppressors allows the squamous epithelial cells in the cervix to proliferate abnormally and uncontrollably. Cancer is caused, for unknown reasons, when these epithelial cells reach the columnar cells in the transformational zone (Figure 3).  Antibodies triggered by the vaccine shown binding to HPV in transformation zone of the squamous epithelia and columnar cells.
 * Figure 3. Cervical Wall Diagram.**

The E6 Protein:
E6 is one of the E oncoproteins originating from the HPV retrovirus and transcribed from human DNA. When HPV enters the body, it produces an E2 protein that binds to the E2 promoter region of human DNA and induces E6 protein transcription. This unregulated transcription leads to accumulation of excessive amounts of E6 protein (Sanchez). E6 then binds ubiquitin ligase, which under normal circumstances would attach ubiquitin to its target proteins. The E6 protein has a binding site pocket made of three domains: the beginning Zinc C domain, the terminal Zinc N domain, and a linker helix to separate the two. Many human proteins, including p53 and Rb, have binding sites in their conformation consisting of an LxxLL combination, with L being acidic Leucine and x being some hydrophobic amino acid (not always the same). The zinc and linker helix domains on E6 are also very hydrophobic, so these two sites become quickly attracted to one another and bind easily (Zanier). When E6 binds a protein, for example p53, it attaches ubiquitin. This degrades the p53 protein, disabling its tumor suppressor abilities. Degrading proteins like Rb and p53 ultimately causes uncontrolled cell division and growth.

[[image:Screen Shot 2014-06-02 at 4.15.14 PM.png]]
Top: The structure of HPV16 E6 bound to residues 403-414 of ubiquitin ligase. Bottom: The hydrophobic pocket (pink) responsible for LxxLL motif recognition in HPV16 E6. Both structures show helical LxxLL peptides inserted in a deep pocket formed by the two zinc domains.
 * Figure 4. Conformation of E6 protein.**

HPV Vaccine:
<span style="font-family: Arial,Helvetica,sans-serif;">HPV vaccines were FDA approved in 2006 after a long period of clinical trials. There are currently two prophylactic vaccines on the market, made by Merck & Co. and GlaxoSmithKline (GSK) Biologicals. Both companies used HPV 16 and 18 as the backbones of their vaccines (Merck & Co. also included HPV 6 and 11), and coincidentally relied on the same basic technology. The vaccines contain a viral protein called L1. Researchers found that when HPV’s L1 gene was inserted into yeast or another virus, virus-like particles self-assembled. This is because the L1 protein makes up most of HPV’s outer shell. When L1 is inserted into the body, production of antibodies is triggered in the bloodstream. These antibodies then pass through the base membrane of the cervix and into the mucosal surface. If HPV is introduced at a later time, the L1 antibodies bind HPV and prevent it from causing an infection. This was an exciting discovery for two reasons: first, since these empty L1 shells contained none of HPV’s actual DNA, they could safely trigger effective immune responses in people injected with the protein without the risk of actual infection. Second, these virus-like particles could be produced in high quantities, as opposed to HPV itself, which grows poorly in lab cultures (Cohen).

<span style="font-family: Arial,Helvetica,sans-serif;">Clinical trials for both companies’ vaccine were enormously successful. Merck & Co.’s vaccine was tested on 500 women; of those 500, 89% achieved protection against persistent infection and completely prevented cervical cancer. Even more impressively, GSK’s vaccine, tested on 700 women, showed 100% efficacy.

Pap Smear:
The Papanicolaou test, or Pap smear, is a test that swabs the cervix and looks for abnormal cells. This cancer screening method is recommended annually for sexually active women over the age 21. The use of the Pap smear has greatly reduced cervical cancer rates. In 2002, about 5,000 American women died from cervical cancer--an enormous 75% drop in mortality since 1950, before the Pap smear was widely used (Cohen). However, this is not the same case in underdeveloped countries.

<span style="font-family: Arial,Helvetica,sans-serif;">The prevalence of cervical cancer has decreased significantly since the invention of the Pap smear. However, this effective form of screening is still unavailable in many parts of the world, especially developing countries. Thus, cervical cancer cases and deaths have become increasingly concentrated in the poorer areas of the world. This is largely due to the expense of the procedure.
 * <span style="font-family: Arial,Helvetica,sans-serif;">Figure 5. Disproportionate impact. **

=Conclusion=

In developed countries, potential negative impacts of widespread HPV vaccination should be considered. The HPV vaccines used only account for a few types of HPV, so many women could still contract a less common strain. Also, in countries where Pap smears and other cervical cancer screening methods are widely used, the HPV vaccine may be superfluous (Cohen). Women monitor their cervical cancer status so regularly with screening that another form of cancer prevention may not be needed. Widespread HPV vaccination could also have a negative impact on screening; if women believe they are exempt from the risk of cancer, they could potentially feel they do not need to get screening.

However, in developing countries, potential negative impacts of HPV vaccination are irrelevant. Focus should be placed primarily on administering HPV vaccines here because cancer screening is uncommon.

<span style="font-family: Arial,Helvetica,sans-serif;">The World Health Organization (WHO) has proposed an HPV Vaccine Program for girls ages 9-13, which would deliver vaccines to developing countries. They conducted a study in Tanzania testing and analyzing the relative cost of HPV vaccine delivery. The results showed that when countries begin administering a new vaccine, they faced initial high costs as well as incremental costs to deliver the vaccines on an on-going basis. As administration continued, demand for the vaccine increased, which also contributed to net increased costs. The figure below shows a five year cost of introducing a national phased-in HPV vaccine program in Tanzania. <span style="font-family: Arial,Helvetica,sans-serif;">The resource requirements over five years that include shared costs (for example, transport or salaries also used for other vaccines) are approximately US$58 million (Hutubessy).
 * Figure 6. Economic costs of HPV vaccine.**


 * Figure 7. World Health Organization's statistics.**

"The financial delivery costs of nationwide HPV vaccination are higher than those of infant vaccines and can be substantial in resource-poor settings since it requires building up new delivery channels. As a consequence, governments need to plan ahead for these non-vaccine costs so that they will have adequate finances in place for vaccine introduction" (Hutubessy).

Pap smears in the United States cost $50-$200 per person, without insurance. This cost is relatively universal. Additionally, clinicians in underdeveloped countries tend to be less highly trained than those in the US, and the clinics are less established. Because of this, Pap smears in developing countries have a significantly lower specificity and thus a lower accuracy in diagnoses as a whole; this often results in an abnormal amount of false positive diagnoses (Denny). Conversely, WHO estimates that HPV vaccines will cost less than US$5 dollars per person. As we previously discussed, efficacy of the vaccine is extremely high. Thus, upon comparing the two methods of cervical cancer prevention in developing countries, vaccination is clearly the better choice.

When comparing the two methods of cervical cancer prevention and the allocation of these services, one can see a great cost discrepancy. After weighing the expenses of delivery services, supervision, and medical assistants for both treatments, you see that these expenses hardly differ from one another. We can then conclude that the significant difference lies in the cost of administration of treatment per person. The HPV vaccine is much more easily administered and less expensive, making it the better choice in developing countries.

After reviewing WHO's HPV vaccine program, we concluded that it would be beneficial to add an additional phase that focuses on older women already infected with HPV. Methods used for each focused age group of women towards prevention of cervical cancer for all. These treatment options would be focused on in the second phase we are proposing for WHO's program.
 * Figure 8. Overview of programmatic interventions over the life course to prevent HPV infection and cervical cancer.**

The vaccine is not for everyone; there are some qualifications that need to be met in order to receive it. These include not yet being predisposed to HPV (not sexually active) and being between the ages of 9-13. WHO suggests that a pap smear should be given to any women at least once within the ages of 30-49 (World Health Organization pg 9). This is not to the United States standard of preventitive health care. Women are recommended to have a pap smear every other year beginning at the age of 21. By the U.S. standard, we would have a difficult time finding the funding to accomplish this task in underdeveloped countries. The argument we propose to this statement is if each woman had one pap smear, a lot of information could be gained from that alone. Lives could be saved as cancer would be discovered and treated. As for those who have cancer, depending on how aggressive/ far along it is, drugs for cure or comfort could be administered accordingly. This second phase could benefit many women's lives greatly.

By adding other age groups to this cause we can eventually have full coverage of the cervical cancer outcome in underdeveloped countries. Because cervical cancer is the leading killer of women worldwide, we could greatly impact this statistic. Another factor to consider is to vaccinate boys at a young age too, because they can be carriers of the disease. These measures have already been put forth in developed countries and have been successful. Even though this is not a complete cure to cervical cancer, it will greatly change the lives of women in underdeveloped countries.

=<span style="font-family: Arial,Helvetica,sans-serif;">Bibliography =

Cohen, J. "PUBLIC HEALTH: High Hopes and Dilemmas for a Cervical Cancer Vaccine." //Science// 308.5722 (2005): 618-21. Web. 11 May 2014. <http://www.sciencemag.org/content/308/5722/618.full>.

Denny, Lynette, Michael Quinn, and R. Sankaranarayanan. "Chapter 8: Screening for Cervical Cancer in Developing Countries." //Vaccine// 24 (2006): S71-77. //World Health Organization//. 14 Mar. 2006. Web. 04 June 2014. <http://www.who.int/immunization/sage/Dennycervical_cancer.pdf>.

"HPV Vaccine Introduction Clearing House." //WHO//. World Health Organization, n.d. Web. 04 June 2014. <http://www.who.int/immunization/hpv/en/>.

Huibregtse, J. M., M. Scheffner, and P. M. Howley. "A Cellular Protein Mediates Association of P53 with the E6 Oncoprotein of Human Papillomavirus Types 16 or 18." //Embo Journal// 10.13 (1991): 4129-135. Web. 21 May 2014. <http://www.ncbi.nlm.nih.gov/pmc/articles/PMC453163/>.

"Human Papillomavirus (HPV)." //Centers for Disease Control and Prevention//. Centers for Disease Control and Prevention, 05 Feb. 2013. Web. 04 June 2014. <http://www.cdc.gov/hpv/signs-symptoms.html>.

Hutubessy, R., A. Levin, S. Wang, W. Morgan, M. Ally, T. John, and N. Broutet. "A Case Study Using the United Republic of Tanzania: Costing Nationwide HPV Vaccine Delivery Using the WHO Cervical Cancer Prevention and Control Costing Tool." (2012): n. pag. //Pub Med//. Web. 04 June 2014. <http://www.ncbi.nlm.nih.gov/pubmed/23146319>.

Sanchez, I. E., M. Dellarole, K. Gaston, and G. De Prat Gay. "Comprehensive Comparison of the Interaction of the E2 Master Regulator with its Cognate Target DNA Sites in 73 Human Papillomavirus Types by Sequence Statistics." //Nucleic Acids Research// 36.3 (2007): 756-69. //Oxford Journals//. Web. 20 May 2014. <http://nar.oxfordjournals.org/content/36/3/756.abstract>.

Schiffman, M., A. G. Glass, N. Wentzensen, B. B. Rush, P. E. Castle, D. R. Scott, J. Buckland, M. E. Sherman, G. Rydzak, P. Kirk, A. T. Lorincz, S. Wacholder, and R. D. Burk. "A Long-term Prospective Study of Type-Specific Human Papillomavirus Infection and Risk of Cervical Neoplasia Among 20,000 Women in the Portland Kaiser Cohort Study." //Cancer Epidemiology Biomarkers & Prevention// 20.7 (2011): 1398-409. Web. 02 June 2014. <http://cebp.aacrjournals.org/content/20/7/1398.full>.

Scudellari, Megan. "HPV: Sex, Cancer and a Virus." //Nature// 503.7476 (2013): 330-32. Web. 11 May 2014. <http://www.nature.com/news/hpv-sex-cancer-and-a-virus-1.14194>.

World Health Organization. "Comprehensive Cervical Cancer Pr Evention and Contr Ol: A Healthier Future for Girls and Women." //WHO// (2013): 1-16. Web. 4 June 2014. <http://www.who.int/immunization/hpv/learn/comprehensive_cervical_cancer_who_2013.pdf?ua=1>.

Zanier, K., S. Charbonnier, A. O. M. O. Sidi, A. G. Mcewen, M. G. Ferrario, P. Poussin-Courmontagne, V. Cura, N. Brimer, K. O. Babah, T. Ansari, I. Muller, R. H. Stote, J. Cavarelli, S. Vande Pol, and G. Trave. "Structural Basis for Hijacking of Cellular LxxLL Motifs by Papillomavirus E6 Oncoproteins." //Science// 339.6120 (2013): 694-98. Web. 12 May 2014. <http://www.sciencemag.org/content/339/6120/694.full>.

Zelkowitz, R. "CANCER: HPV Casts a Wider Shadow." //Science// 323.5914 (2009): 580-81. Web. 11 May 2014. <http://www.sciencemag.org/content/323/5914/580.full>.