2004 Summer Meeting - Amsterdam - The Pathological Society of ...

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A Study of Insitu Hybridisation for HPV in Women With
High Grade HPV and Low Grade Smear With Clinical
Progression or Regression
K Morris 2 , B Fakokunde 1 , S Glew 1 , J Pawade 3
1 St Michael's Hospital, Bristol, United Kingdom, 2 University of Bristol,
Bristol, United Kingdom, 3 Bristol Royal Infirmary, Bristol, United
Kingdom
We have investigated for presence of high grade HPV by PCR and insitu
hybridisation using a wide spectrum probe in women presenting with low grade
smear abnormality and koilocytosis and or CIN1 in cervical biopsies and
correlated the findings with clinical regression and progression.
Nine patients (progressors group) on HPV assessment had single HPV type in 3
(2 with HPV 16) and multiple in 6 (5 with HPV 16). The cervical punch
biopsies showed positive insitu signal in all cases with punctate pattern in 6 and
diffuse in 3. There was strong nuclear expression of P16 in 7 biopsies.
Ten patients (regressor group) had single HPV type (HPV 16) and 8 with
multiple HPV ( 4 HPV 16 and 2 with HPV 18). Positive insitu signal was seen
in 9 cases with a diffuse pattern in 7 and punctate in 2. There was positive
staining for p16 in 4 cases.
Integration of HPV in the cells demonstrated by punctate signal by insitu and
strong expression of p16 were features associated with clinical progression.
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Cytokeratin Immunostaining Differentiates Endometrioid
Carcinoma Of Ovary From Metastatic Colorectal Carcinoma
M Jiménez-Liñán 1 , L Happerfield 1 , X Giannakoulopoulos 2 ,
RAF Crawford 2 , MJ Arends 1
1 Department of Pathology, University of Cambridge, Cambridge, United
Kingdom, 2 Addenbrooke's Hospital, Cambridge, United Kingdom
Ovarian metastases from colorectal adenocarcinoma can sometimes
histopathologically mimic primary ovarian carcinomas of endometrioid type.
Distinction of endometrioid from colorectal cancer morphology is widely
recognised as problematic on histological examination. However, it is important
for therapeutic and prognostic reasons to differentiate between primary ovarian
endometrioid carcinoma and metastatic colorectal adenocarcinoma. We
examined a series of 14 cases originally reported as primary endometrioid
carcinomas of ovarian to ascertain whether the site of origin could be
determined by using immunohistochemistry for cytokeratin 7 and cytokeratin
20. Follow-up showed that of the 14 patients, 3 were subsequently shown to
have colorectal carcinomas. Immunohistochemistry showed a cytokeratin
7+/cytokeratin 20- immunophenotype in 82% of the eleven primary ovarian
endometrioid carcinomas; cytokeratin 7+/cytokeratin 20 focally positive pattern
in 18% of the 11 primary ovarian endometrioid carcinomas; and cytokeratin 7-
/cytokeratin 20+ immunophenotype in 100% of the three metastatic colorectal
adenocarcinomas.
We conclude that the combination of cytokeratin 7 and cytokeratin 20
immunohistochemistry is very useful in distinguishing between metastatic
colorectal adenocarcinoma and primary endometrioid carcinoma of ovary.
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COMPARISON OF THE CONVENTIONAL CERVICAL
CYTOLOGY SMEAR, LIQUID-BASED CYTOLOGY AND
HC2 HIGH-RISK HPV DNA TEST USING UCM-
COLLECTED SPECIMENS IN CLINICAL PRACTICE
N Bolger 2 , L Pilkington 2 , N Murphy 1 , W Prendiville 2 , JJ O'
Leary 1
1 Department of Pathology, Trinity College, Dublin, Ireland, 2 The
Coombe Women's Hospital, Dublin, Ireland
Digene’s Universal Collection Medium (UCM) was developed to provide dual
cytologic and direct HC2 DNA testing. The study sought to compare
performance of UCM Pap cytology, conventional Pap smear, combination
testing and HPV stand alone testing.
Each subject consented to a conventional Pap smear and a second sample
collected with Digene’s Cervical Sampler for the UCM Pap test and HPV
test. The Digene brush sample was placed in 1 mL UCM. A 500 µl aliquot
was processed according to the Shandon PapSpin protocol. The remaining
500 µl was processed by the HC2 method for HPV DNA detection. Both
cytology slides were routinely stained and read in a blinded manner.
Overall agreement between conventional and UCM Pap tests was 88%.
Compared to biopsy results, the cytologic sensitivity of conventional Pap smear
and UCM Pap test for HSIL (CIN 2/3) was 32% and 35% respectively. HC2
HPV test alone detected 97% biopsy confirmed HSIL. The combination of
conventional and/or UCM Pap cytology and HPV test resulted in 100%
sensitivity for biopsy confirmed high-grade lesions and cervical cancer.
The preliminary data support the use of UCM-collected cervical samples as a
convenient method of combination LBC and HPV testing for cervical cancer
screening.
Digene Corporation and Thermo Electron Corporation funded this study.
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p16 INK4A , MCM5, CDC6 and High-Risk HPV status: A
Screening Algorithm for Cervical Disease
N Murphy 1 , M Ring 2 , M Griffin 3 , O Sheils 3 , JJ O' Leary 1
1 Department of Pathology, Trinity College, Dublin, Ireland, 2 Department
of Pathology, The Coombe Women's Hospital, Dublin, Ireland, 3
Department of Histopathology, Dublin, Ireland
Increasing knowledge of the underlying pathogenesis of cervical cancer
combined with advances in molecular testing herald the emergence of a
“molecular age” in cervical cancer prevention. This new era promises the use
of HPV testing and molecular biomarkers of cervical dysplasia. The objective
of this study was to develop a screening algorithm which allows risk
stratification of women on the basis of high-risk HPV DNA detection and the
expression of p16 INK4A , MCM5 and CDC6 in squamous and glandular
dysplastic cervical cells.
Generic HPV detection was carried out using a modified SYBR
green assay system. Taqman PCR was employed for the detection of HPV 16,
18, 31 and 33 DNA. Immunohistochemical analysis using mouse monoclonal
antibodies was employed to examine the protein expression pattern of p16 INK4A ,
MCM5 and CDC6 in normal cervical epithelium (n=20), CIN1 (n=38), CIN2
(n=33), CIN3 (n=46), cGIN (n=10), invasive squamous cell carcinomas (n=10)
and adenocarcinomas (n=10) of the cervix. All samples were formalin fixed
and paraffin embedded. Staining intensity was assessed using a 0 to 3 scoring
system. In addition expression status was examined in 10 normal ThinPrep
smears and 15 smears exhibiting mild, moderate or severe dyskaryosis.
p16 INK4A , MCM5 and CDC6 mRNA expression was examined using
quantitative TaqMan RT-PCR.
In this study p16 INK4A identified dysplastic squamous and glandular
cervical lesions with a sensitivity of 99.9% and a specificity of 100%. MCM5
marked squamous and glandular dysplastic lesions with a sensitivity of 95%
and a specificity of 100%. Quantitative TaqMan RT-PCR demonstrated an
increase in MCM5 mRNA expression with increasing severity of dysplasia.
CDC6 identified squamous and glandular dysplastic lesions of the cervix with a
sensitivity of 70% and a specificity of 100% and was preferentially expressed in
high-grade lesions and in invasive disease. TaqMan RT-PCR revealed an
increase in CDC6 mRNA expression in dysplasic cells. The results of this
study strongly indicate that the combined use of HPV DNA testing and MCM5,
CDC6 and p16 INK4A detection methods can be employed to identify cervical
neoplasic cells and can markedly increase the sensitivity and specificity of
cervical cancer screening.
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