UNIVERSITY OF THESSALY

SCHOOL OF HEALTH SCIENCES – FACULTY OF MEDICINE

DEPARTMENT OF O&G UNIVERSITY HOSPITAL LARISA

MANAGEMENT OF HPV POSITIVE CASES

AFTER SCREENING
EBCOG ACCREDITED
2015-2019

Alexandros I. Daponte
M.D. (Thessaloniki), Dr Med (LMU Munich) F.C.O.G. (S.A.)
Head of the Department of Obstetrics and Gynaecology
Associate Professor of Obstetrics and Gynaecology
2008
 AIM: Reduce colposcopies-
unnecessary treatment

Triaging hrHPV positive women, and, therefore, reducing the psychological

burden on women referred for colposcopy or even to unnecessary treatment
is a necessary approach currently investigated worldwide.

To minimise the effect of HPV DNA testing’s low specificity,

several strategies have been investigated for

triage of hrHPV positive women
to colposcopy ? OTHER

according to cervical cancer risk assessment

 COTESTING Cytology + HPV
Co-testing identifies the same number of precancerous lesions
as hrHPV testing,
however,

worse specificity and PPV and requiring more colposcopies to

identify one precancerous lesion.

This reflects the fact that the HPV component is considered the important one
concerning cotesting and that a negative HPV test provides better 3-year
safety for cervical cancer compared to the 5-year one provided by a negative co-test

Gage JC, Schiffman M, Katki HA, Castle PE, Fetterman B, Wentzensen N, Poitras NE, Lorey T,
Cheung LC, Kinney WK(2014) Reassurance against future risk of precancer and cancer
conferred by a negative human papillomavirus test. J Natl Cancer Inst 106 (8).
 While HPV2/NILM cotesting results are associated with low CIN3 risk, HPV

testing had similar screening performance to cotesting and to cytology alone .

 Athena Trial
Οncogenic potential of HPV16 and 18 is greater than the remaining
hrHPVs, since about 70% of cervical cancer cases is attributed to
these two HPV types.

3 years after a HPV test positive the cumulative incidence

rate for CIN3+ is

• 25.2% for HPV 16,

• 10.0% for HPV18,

• 5.4% altogether for other hrHPVs .

Wright TC, Stoler MH, Behrens CM, Sharma A, Zhang G,Wright TL (2015) Primary cervical
cancer screening with human papillomavirus: End of study results from the ATHENA
study using HPV as the first-line screening test. Gynecol Oncol.
 Genotyping
identifies women at highest risk
Risk of developing CIN3+ within 3 years

1 in 4

1 in 9

12 other
hrHPV 1 in 19

Source: Wright et al., Gynecologic Oncology, 2015

HPV 16,18 GENOTYPING IN CHINA
 HPV16/18 positive women to colposcopy and cytology
testing for the remaining hrHPV positive women
The combination of the two methods forming a comprehensive
initial screening strategy seems to be the optimal approach.

According to this, the referral of HPV16/18 positive women to colposcopy and

cytology testing for the remaining hrHPV positive women with subsequent
colposcopy in case of ASCUS+ identified 28.6% more precancerous
lesions. (Huh et al.)
Currently constitutes an interim clinical guidance issued by the American Society
of Colposcopy and Cervical Pathology (ASCCP).
Huh WK et al. (2015) Use of primary high-risk human papillomavirus testing for cervical
cancer screening: interim clinical guidance. Obstet Gynecol
125(2):330–337.
Saslow D, et al, American Cancer S, American Society for C, Cervical P, American Society
for Clinical P (2012) American Cancer Society, American Society for Colposcopy and Cervical
Pathology, and American Society for Clinical Pathology screening guidelines for the
prevention and early detection of cervical cancer. Am J Clin Pathol 137 (4):516–542.
 Similar PPV for cytology and
HPV16/18 genotyping

Larger studies utilizing different HPV assays report

Similar PPV for cytology and HPV16/18 genotyping

concerning triage of HPV positive women to
colposcopy .

Castle PE, Stoler MH, Wright TC Jr, Sharma A, Wright TL, Behrens CM (2011) Performance
of carcinogenic human papillomavirus (HPV) testing and HPV16 or HPV18 genotyping for
cervical cancer screening of women aged 25 years and older: a subanalysis of the
ATHENA study. Lancet Oncol
Agorastos T, Chatzistamatiou K, Katsamagkas T, Koliopoulos G, Daponte A, Constantinidis
T, Constantinidis TC, the Hsg (2015) Primary screening for cervical cancer based on high-
risk humanPapillomavirus (HPV) detection and HPV 16 and HPV 18 Genotyping,
in Comparison to Cytology. PLoS One 10(3):
 Use of primary high-risk human papillomavirus testing
for cervical cancer screening: Interim clinical guidance.
(Huh et al, Gyn Oncology, 07.01.2015) 

Society of Gynecologic Oncology

American Society of Cytopathology
American Cancer Society
American Society for Colposcopy and Cervical Pathology
American College of Obstetricians and Gynecologists
College of American Pathologists
American Society for Clinical Pathology

HPV testing & genotyping with cytology as triage

FDA approved strategy (25+)
Conclude that 12-month surveillance of women who test positive for other high-
risk HPV (not 16/18) and cytology ASC-US/LSIL, appears to provide the best
balance of benefits,harms and cost-effectiveness in the context partial
genotyping for HPV 16/18 within the new Australian primary-HPV based
screening program.
 Τhe spectrum of disease progression

HPV

HPV HPV DNA HPV E6/E7 Cell cycle Cance

infection replication gene expression deregulation r

HPV DNA Test - +

HPV E6/E7 mRNA Test - - +

p16/Ki-67 Test - - - +

15
 Staining with p16/Ki-67
A potential triage system for these cases could be additional staining with p16/Ki-67.

p16 is a cell cycle regulatory protein, which in normal physiological conditions

triggers cell cycle arrest.

Ki-67, on the otherhand, is a proliferation marker.

In normal circumstances, p16 and Ki-67 are rarely seen together.

Overexpression of both p16 and Ki-67 suggests a deregulation of the cell cycle by
HPV and is indicative of a high grade lesion

This concept of dual-stain cytology (CINtec1 PLUS Cytology test) is a morphology-

independent biomarker approach.
Dual staining
Dual staining
 Higher proportion of true vs. false
test results in the dual-stained cytology approach
Dual-stained cytology showed a similar test positivity
rate in HPV-positive women (14 pooled genotypes) as Pap cytology
(28.4% vs. 26.4%, respectively), and in clinical practice this would result
in a similar number of women being referred to colposcopy.

However,
the sensitivity of dual-stained cytology was significantly higher than
the sensitivity of Pap cytology (74.9% vs. 51.9% for CIN3+;p LESS THAN 0.0001).

Also, positive and negative predictive values were significantly higher for dual-stained
cytology compared to Pap cytology, whereas there was no statistically significant
difference in specificity (p=0.3198 at the CIN3+threshold).
 Strategies that utilized dual-stained cytology were
more sensitive or more effective, or both
than strategies that utilized Pap cytology
testing for the triage of primary HPV-positive results

This study, for the first time, analyzed a combination of

HPV16/18 genotyping and p16/Ki-67
dual-stained cytology for the triage
of women in a setting of HPV primary screening.

Highest Sensitivity (86.8%)

NPV (98.2%) for CIN3+

of all the triage strategies that were evaluated

 Use of primary high-risk human papillomavirus testing
for cervical cancer screening: Interim clinical guidance.
(Huh et al, Gyn Oncology, 07.01.2015) 

Society of Gynecologic Oncology

American Society of Cytopathology
American Cancer Society
American Society for Colposcopy and Cervical Pathology
American College of Obstetricians and Gynecologists
College of American Pathologists
American Society for Clinical Pathology

HPV testing & genotyping with cytology as triage

FDA approved strategy (25+)
 Strategies that utilized dual-stained cytology were
more sensitive or more effective, or both than
strategies that utilized Pap cytology
testing for the triage of primary HPV-positive results

Dual-stained cytology as a single triage test for HPV-

positive (14 pooled genotypes) women resulted in the
highest PPV and the lowest number of colposcopies per
CIN3+ detected.
However,
Sensitivity of this approach was 12% lower than a
strategy that refers HPV16/18 positive women to
colposcopy and then triages women positive for 12
“other” HPV genotypes using dual-stained cytology.
We span the spectrum of disease progression

HPV

HPV HPV DNA HPV E6/E7 Cell cycle Cance

infection replication gene expression deregulation r

HPV DNA Test - +

HPV E6/E7 mRNA Test - - +

p16/Ki-67 Test - - - +

23
Triage of AHPV+ by abnormal reflex LBC and the presence of HPV 16/18/45
would result in a significantly lower colposcopy referral rate with similar
CIN2+ and CIN3+ detection rates as the overall HC2+ referral algorithm.
Viral load
 HPV testing’s
worse specificity versus cytology,
The PIPAVIR project (E7 detection)
The PIPAVIR project will contribute to that direction by evaluating the use of a novel E7
oncoprotein detection assay for that purpose.

PIPAVIR database on better triage strategies, using novel assays for hrHPV positive
women.
Based on this algorithm, a detailed analysis has been planned to detect
whether the triage process can be further improved by combining current and novel
triage modalities, namely, the detection of HPV E7 oncoprotein, in different ways.

In more detail, it has been planned to examine the diagnostic accuracy of E7 detection
for the triage to colposcopy of hrHPV 16/18 positive women and of hrHPV (non
16/18) positive women compared to cytology.
Regarding the latter the goal will be to test whether a fully molecular process for both,
cervical cancer screening and triage to colposcopy is more accurate than the
established practices.
The PIPAVIR project (E7 detection)
 Triage of HPV positive alternative
(?self sampling samples)
Few commercial HPV assays have been evaluated, and some of them are specifically
designed to test alternative clinical specimens such as self-collected cervicovaginal
lavage,vaginal and urethral swabs, urine and tissue specimens (in situhybridization-
based HPV tests),

we urgently need an evaluation of the performance of as many commercial HPV tests

as possible on a broader range of alternative clinical specimens;

for example, other types of self-collected anogenital samples (different brushes,

swabsand sponges, tampons), oral swabs, saliva, anal swabs, penile swabs,Guthrie-
type filter paper, and so on,
HPV DNA Self sampling
VAGINA URINE
 HPV 16 DNA screening και self sampling
δείγμα κόλπου University Hospital Larissa 2004-2005
.

Self sampling με τυποποίηση για

HPV 16 σε δείγμα κόλπου
έχει ευαισθησία παρόμοια με
του τραχήλου
HPV 16 DNA screening και self sampling
δείγμα ούρων- FIRST VOID
University Hospital Larissa 2004-2005

Self sampling ούρων με

τυποποίηση για HPV 16 έχει
ικανοποιητική ευαισθησία
στις σοβαρότερες βλάβες
(καρκίνος 88.8%,
βαριά δυσπλασία 76.5% ε
ήπια δυσπλασία 45.5%)
Abstract
Background
HPV testing from clinician-collected cervical and self-collected cervico-vaginal samples is more sensitive for detecting
CIN2+/CIN3+ than cytology-based screening, stimulating interest in HPV testing from urine. The objective was to determine
the performance of the Trovagene HPV test for the detection of CIN2+ from urine and PreservCyt cervical samples.
Methods
Women referred for colposcopy at St Mary’s Hospital London, following abnormal cytology, were recruited to this diagnostic
accuracy study by convenience sampling (September 2011 and April 2013). 501 paired urine and cervical samples were
collected. Primary outcome: sensitivity for CIN2+/CIN3+; specificity for <CIN2. Secondary outcomes: comparisons with other
HPV tests; agreement/kappa values between urine and cervical samples.
Results
Trovagene HPV test sensitivity and specificity from PreservCyt were similar to well-established tests [sensitivity for CIN3+
(n=145) 96·3%(95%CI,89·6-99·2); CIN2+(n=81) 94·5%(95%CI,89·4-97·6); specificity for <CIN2 25·3%(95%CI,20·8-30·1)].
Sensitivity from urine was slightly, but not significantly lower [CIN3+ 91·4%(95%CI,83·0-96·5), P=0·3; CIN2+ 88·3%(95%
CI,81·9-93·0), P=0·06]. Specificity for <CIN2 was similar: 24·7%(95%CI,20·3-29·5), P=0·9. 403 Trovagene HPV tests were
positive and 396 urine tests. Overall agreement between paired samples was 82·6%(95%CI,79·3-86·0).
Conclusion
Trovagene HPV test’s performance on PreservCyt cervical samples was comparable to established HPV tests. Sensitivity in
urine although slightly lower may nevertheless be adequate for selfsampling. This referral population’s higher HPV positivity
rate affects specificity, warranting further studies in a screening population.
Impact
This may prove useful for women not attending for cervical screening.
HPV positive self sampling samples

Piyathilake et al. (Cancer, 2016, 122, 2836-2844) clearly demonstrated this in their
larger study of 502 women, where only 1 in 5 women testing positive for high risk-HPV in their urine
or cervix had CIN2 or worse. Where HPV testing is of particular value is rather the reassurance it
gives women with a negative test; the same group found that the absence of high risk-HPV infection
was truly associated with no disease in over 9 out of 10 cases.
Before urinary HPV testing can be rolled out into routine screening, however, further data is urgently
required concerning the accuracy and acceptability of urinary HPV testing in the general population,
instead of the selected colposcopy referral population that has been the focus of studies so far. The
prevalence of cervical HPV infection is lower in the general population, and this will influence the
sensitivity and negative predictive value of the test. Recruiting ‘hard to reach’ groups is a priority of
future research as it is precisely these women who are most likely to benefit from its use in clinical
practice. If urinary HPV testing overcomes barriers to cervical screening that prohibit its uptake by
these women, it may become an important weapon in the fight against cervical cancer.
 Because additional testing could involve additional costs, a health
economic analysis assessing costs and benefits should be performed
in order to determine the cost-effectiveness and economic impact of
any method.
 UNIVERSITY OF THESSALY
SCHOOL OF HEALTH SCIENCES –
FACULTY OF MEDICINE
MANAGEMENT OF HPV POSITIVE CASES
AFTER SCREENING
THANK YOU FOR YOUR ATTENTION
EBCOG ACCREDITED
2015-2019

Alexandros I. Daponte
M.D. (Thessaloniki), Dr Med (LMU Munich) F.C.O.G. (S.A.)
Head of the Department of Obstetrics and Gynaecology
Associate Professor of Obstetrics and Gynaecology
MANAGEMENT OF HPV POSITIVE CASES
AFTER SCREENING

THANK YOU
Alexandros I. Daponte
M.D. (Thessaloniki), Dr Med (LMU Munich) F.C.O.G. (S.A.)
Head of the Department of Obstetrics and Gynaecology
Associate Professor of Obstetrics and Gynaecology