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This article is part of a new series on diagnostics within Women’s Health. We’ll be adding more stories and related content throughout the year. 

In 2020, the US FDA approved the use of p16/Ki-67 dual staining technology [1], which is set to advance cervical cancer triage by enabling more accurate diagnosis and greater risk stratification than current cytology usage or co-testing with HPV16/18 genotyping [2]. This development holds great promise to improve patient outcomes and reduce costs in managing a highly preventable disease.

The value of p16/Ki-67 dual staining biomarkers in triage

In recent years, primary HPV screening has improved in many countries, which now offer expanded programmes with Pap cytology and HPV DNA testing. Since HPV infection does not necessarily lead to cervical cancer, women still need to undergo triage screening after a primary positive HPV test to determine their level of cervical cancer risk. It is unrealistic to direct every positive case to colposcopy, an invasive procedure that is not only costly but  places unnecessary stress on the patient, especially considering that the majority of HPV infections are transient and self-clearing.

Current triage options in most countries include the use of cytology alone or co-testing with HPV genotyping. However, cytology-based methods have been known to be resource-intensive and results are open to interpretation [3]. HPV genotyping aims to alleviate some of the uncertainty that is associated with Pap cytology, but it is unable to provide full assurance to patients and clinicians alike when a positive HPV result is returned.

Staining for the biomarkers p16/Ki-67 in triage has been the subject of multiple studies for years now [4]. Since these two biomarkers are mutually exclusive in a normal cell, an indication of both suggests that a cell is undergoing a transforming HPV infection and the patient is at greater risk.

The landmark Improving Primary Screening and Colposcopy Triage (IMPACT) trial [2], which released its findings in July 2021, involved over 35,000 eligible women (between the ages of 25 to 65) and followed them for a year in the United States. It evaluated the use of dual biomarker staining for p16 and Ki-67 as compared to (a) Pap cytology alone and (b) co-testing with HPV16/18 genotyping. Here are some key findings:

• Dual staining biomarkers are more efficient at detecting cervical disease. At the disease stage of CIN2, Pap cytology alone was able to detect at a rate of 65.9% with co-testing showing a rate of 76.4%. Dual biomarker staining was able to detect at 86.5%. This trend holds true for detecting disease at the CIN3 stage. Of those who were escalated to colposcopy, more dual stained triaged women were found to have the disease than those triaged with Pap cytology.
• Dual staining biomarkers are able to stratify risk. While HPV16/18 genotyping is valuable in detecting the presence of high risk strains 16 and 18, it does not account for the other 12 strains that are not as common but still associated with causing cervical cancer. Within the group of the 12 other HPV-positive strains, dual biomarker staining is able to stratify patients further into groups of significant risk or those at low risk.
• Dual staining biomarkers are able to stratify risk better than Pap cytology. Pap cytology is also used in triage, but dual staining biomarkers show an advantage over cytology-based methods for risk stratification. In the significant risk group, dual staining was able to detect risk at 19.6% vs Pap cytology risk detection at 17.8%. Within the low-risk group, dual stain also outperformed Pap cytology at 3.6% to 7.4%.

Clearly, dual biomarker staining is a screening method that is precise and leaves no room for misinterpretation. It also puts women in better control of their health journey – where once a negative result meant ambiguity, it is now a definite disease-free status. Dual staining provides great value to clinicians too as they need not worry about the loss to follow up, which has been shown to occur in 4 out of 10 women [5].

Given the benefits, countries like France and Germany have incorporated p16/ki-67 dual biomarker staining into clinical guidelines, with even more countries using them in clinical practices.

Reaching the World Health Organisation’s 90-70-90 goals

Dual staining technology is an important complement to HPV vaccination, which is sometimes regarded as the first line of defense against cervical cancer. Of the 14 genotypes associated with cancer-causing HPV strains, most HPV vaccines provide highly-effective protection against the 2 most common high-risk strains, HPV 16 and HPV 18 [3]. Yet infections from other strains can still occur, making screening and triage an enduring necessity.

The value of diagnostics is clearly evident in the World Health Organisation’s Global Strategy to Accelerate the Elimination of Cervical Cancer, which was released in 2020 and includes a 3-pronged approach of vaccination, screening and treatment. Its ‘90-70-90’ target was to ensure that 90% of girls would be fully vaccinated against HPV by 15 years old; 70% of women would be screened with a high-performance test by age 35 and again by 45; and 90% of women identified with cervical disease would receive treatment.

With 604,000 women diagnosed with cervical cancer in 2020, and 342,000 deaths from it, cervical cancer remains the 4th most common type of cancer amongst women [6]. In terms of screening, consistency with high performing tests and early detection will be central to beating a 100% preventable cancer.

With a powerful tool to be added to triage screening, more countries can now be on track to achieving the 90-70-90 goal by 2030 and setting a cervical cancer-free stage within the next century.

References:
[1] Accessdata.fda.gov. 2022. Premarket Approval (PMA). [online] Available at: <https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpma/pma.cfm?id=P190024> [Accessed 4 January 2022].

[2]  Wright, T., Stoler, M., Ranger‐Moore, J., Fang, Q., Volkir, P., Safaeian, M. and Ridder, R., 2021. Clinical validation of p16/Ki‐67 dual‐stained cytology triage of HPV ‐positive women: Results from the IMPACT trial. International Journal of Cancer, 150(3), pp.461-471.

[3] Safaeian, M., Solomon, D. and Castle, P., 2007. Cervical Cancer Prevention—Cervical Screening: Science in Evolution. Obstetrics and Gynecology Clinics of North America, 34(4), pp.739-760.

[4] Wentzensen, N., Schwartz, L., Zuna, R., Smith, K., Mathews, C., Gold, M., Allen, R., Zhang, R., Dunn, S., Walker, J. and Schiffman, M., 2012. Performance of p16/Ki-67 Immunostaining to Detect Cervical Cancer Precursors in a Colposcopy Referral Population. Clinical Cancer Research, 18(15), pp.4154-4162.

[5] Rebolj, M. and Lynge, E., 2010. Incomplete follow-up of positive HPV tests: overview of randomised controlled trials on primary cervical screening. British Journal of Cancer, 103(3), pp.310-314.

[6] World Health Organisation Fact Sheet on Cervical Cancer. Available at: <https://gco.iarc.fr/today/data/factsheets/cancers/23-Cervix-uteri-fact-sheet.pdf> [Accessed 4 January 2022].