Learning Unit: 8

Neoplasia Of The Female Genital Tract

PATHOGENESIS OF CERVICAL CANCER

 Central aetiological factor is persistent infection with high-risk oncogenic

human papillomavirus (HPV) types.
 At least half of sexually active people will have HPV at some point in their
lives, but only a few women will get cervical cancer.
 HPV causes a variety of genital lesions ranging from warts to cervical cancer
 Certain HPV types affect the genitals and are sexually transmitted. At least 15
of these strains have the potential to cause cervical cancer
 Other factors needed: ?smoking ?other STDs ?immune system
HUMAN PAPILLOMAVIRUS

 Cervical cancer is preventable with the HPV vaccine, and it is curable if

detected and treated early.
 Department of Health (SA) started with HPV vaccination programme in 2014,
vaccinating grade 4 girls (Cervarix®)
 HPV is one of the most common STIs worldwide and infection occurs rapidly
following sexual debut, with per-coital transmission rates estimated at 40%.
 Prevalence peaks around age 25, after which it tends to decline - natural
clearance (i.e. transient infection).
 Approximately 200 HPV genotypes, numbered sequentially, have been
identified.
HUMAN PAPILLOMAVIRUS

 High-risk HPV (hrHPV) are strongly associated with anogenital cancers

(particularly cervical) and precursor lesions.
 15 recognized as high-risk (hr)HPVs: genotypes 16, 18, 31, 33, 35, 39, 45, 51,
52, 56, 58, 59, 68, 73 and 82.
 HPV16 is the most prevalent type detected in HPV-associated cancers,
followed by HPV18. These cause about 70% of cervical cancers worldwide.
 The two most common low-risk HPV types are 6 and 11, which together cause
about 90% of genital warts.
 HPV infection is also associated with cancers of the anus, penis, vagina and
vulva, as well as a growing number of head and neck cancers.
HUMAN PAPILLOMAVIRUS

 The genital HPVs are divided into low-risk and high-risk types based on the
frequency of their association with invasive cervical cancer.
 By definition, an HPV is low risk if it has never been isolated from a cervical
carcinoma, and high risk if it ever has been.
 Persistent infection with any one of about 15 high-risk (carcinogenic) types
accounts for virtually all cervical cancers.
HUMAN PAPILLOMAVIRUS

 The virus is believed to enter the body through small, inconspicuous cuts or
abrasions on the skin or mucous membrane.
 The growth is by clonal expansion and, as suggested it pushes aside the
normal epithelium to form benign warts.
 In these low-grade processes virus replication occurs episomally in the
nucleus.
MOLECULAR PATHWAY OF HPV

 HPV initially infect basal (actively dividing) cells.

 Viral DNA is maintained at a low copy number in infected host cells as they
undergo differentiation and move toward the surface of the epithelium.
 In terminally differentiated cells, the virus replicates to a high copy number,
late genes are expressed, and virions are produced.
 Key events of carcinogenesis is the integration of the HPV genome into a host
chromosome.
 A positive HPV test does not necessarily mean the virus is integrated (i.e.
most HPV tests have poor specificity).
MOLECULAR PATHWAY OF HPV

 Small HPV genome consists of about 8000 base pairs of circular double-stranded DNA.
 It codes for only eight genes , which are classified as “early” (E) or “late” (L) depending on the
timing of their expression in the epithelium.
 HPV infection is established in the basal layers of the epithelium, where the HPV genome is
maintained, with expression of the E genes.
 As the epithelium matures toward the surface, gene amplification and viral assembly occur, with
expression of L1 and L2, with eventual viral release.
 L1 is the major viral capsid protein and is the principal component of HPV vaccines.
 The E6 and E7 gene products play the most significant part in cervical oncogenesis.
 They have a number of cellular targets, with a multitude of effects that lead to malignant
transformation.
 The two most important appear to be (1) the binding of E6 to p53, which results in the blocking
of apoptosis, and (2) the binding of E7 to the retinoblastoma tumor suppression protein pRB,
which abolishes cell-cycle arrest and leads to unscheduled cellular proliferation.
MOLECULAR PATHWAY OF HPV

In summary:
 The key genes involved in the pathogenesis of the virus include L1 (viral
capsid), E6, and E7.
 The E6 and E7 genes work by inhibiting apoptosis and enhancing viral
proliferation by inhibiting retinoblastoma protein, p53, and p21, causing an
accumulation of p16 in infected cells. (Both E6 and E7 HPV oncogenes inhibit
the activities of these tumour suppressors)
 Expression of viral E6 and E7 oncogene mRNA is highly associated with
SIL(squamous intraepithelial lesion) development and is necessary and
sufficient for cell immortalization, neoplastic transformation, and
development of invasive cancer.
HPV GENOME AND FUNCTIONS
HUMAN PAPILLOMAVIRUS

 While HPV infections are very common, women with healthy immune systems
usually clear these infections, while women with HIV are far less likely to
clear HPV.
 Therefor HIV+ women are more likely to develop pre-invasive lesions that can
progress to invasive cancer.
 WHO recommends more screening opportunities and adequate treatment to
all HIV+ women
 Cervical cancer diagnosed in HIV is an AIDS-defining illness and HIV+ women
are at 4-5 times greater risk of developing cervical cancer
SQUAMOUS ABNORMALITIES/LESIONS

 Squamous abnormalities encompass the spectrum of non invasive cervical

epithelial abnormalities associated with human papillomavirus (HPV), ranging
from the cellular changes that are associated with transient HPV infection to
those representing high-grade precursors, to invasive squamous cell
carcinoma.
 Cells do not suddenly change into cancer, but gradually develop abnormal
changes: pre-cursor lesion i.e. squamous intraepithelial lesion (SIL).
 Only some women with SIL will develop invasive carcinoma and treating these
SIL lesions will prevent almost all cervical cancers.
 Goal of screening is to find SIL when it is more treatable and curable.
PRE-CURSOR LESIONS

 The Bethesda System recommends a low-grade/high-grade approach to

grading SIL.
 This is based on the evidence that most LSILs are transient infections that
carry little risk for oncogenesis, whereas most HSILs are associated with viral
persistence and a significant potential for progression to invasive cancer.
 LSIL encompasses lesions previously described separately as koilocytosis (flat
condyloma) and mild dysplasia (CIN 1).
 HSIL encompasses lesions previously described as moderate dysplasia (CIN 2)
and severe dysplasia/carcinoma in situ (CIN 3).
PRE-CURSOR LESIONS

 SIL lesions may disappear without treatment, especially LSIL

 Transient infections generally regress over 1–2 year period, and lesions with
HPV persistence are associated with an increased risk of developing HSIL or
invasive ca
 Many lesions may persist without major changes for years while some lesions
progress very quickly
 Invasive cancer only develop in small % of precursor lesions
 Precursor lesions are not able to metastasise, so it is not life threatening
 If a carcinoma penetrate the basement membrane, it is considered invasive
 Invasion less than 3mm is termed micro-invasive carcinoma (histological term)
LSIL: High N/C ratio
neoplastic cells confined
to lower 1/3 of
epithelium. Ample
squamous differentiation
can still occur before cells
are exfoliated at surface.
Atypical cells on surface
has slightly increased
N/C.
Carcinoma: Neoplastic
cells occupy the entire
thickness and cells
HSIL: High N/C ratio invade basement
neoplastic cells occupy membrane
more than lower 1/3 up to
full thickness of the
epithelium. Less
squamous differentiation
can occur before cells are
exfoliated at surface.
Atypical cells at surface
has a high N/C ratio.
Low-Grade Squamous Intraepithelial
Lesion (LSIL)
 LSIL is a low-risk intraepithelial lesion.
 LSIL is caused by a large number of different HPVs, including low-risk and
high-risk types.
 Many LSILs regress spontaneously, but some persist for long periods of time to
progress to HSIL.
LSIL CYTOMORPHOLOGY

 Cells occur singly, in clusters, and in sheets.

 Cytologic changes are usually confined to squamous cells with “mature”
intermediate or superficial squamous cell-type cytoplasm.
 Overall cell size is large, with fairly abundant “mature” well-defined
cytoplasm.
 Nuclear enlargement >3x the area of a normal intermediate nuclei, results in
a low but slightly increased N/C ratio. Nucleus occupies up to 1/3 of the total
area of the cell.
 Nuclei are generally hyperchromatic but may be normochromatic.
LSIL CYTOMORPHOLOGY

 Nuclei show variable size (anisonucleosis).

 Chromatin is uniformly distributed and ranges from coarsely granular to
smudgy or densely opaque.
 Nuclear membranes variable ranging from smooth to very irregular with
notches.
 Binucleation and multinucleation are common.
 Nucleoli are generally inconspicuous.
 Clean or inflammatory background
LSIL CYTOMORPHOLOGY

 Koilocyte: perinuclear cavitation consisting of a broad, sharply defined clear

perinuclear zone and a peripheral rim of densely stained cytoplasm is a
characteristic viral cytopathic feature (but is not required for LSIL).
 Cells may show increased keratinization with dense, eosinophilic cytoplasm
with little or no evidence of koilocytosis.
 Cells with koilocytosis or dense orangeophilia must also show nuclear
abnormalities to be diagnostic of LSIL
 Hyperkeratosis and parakeratosis can also be seen
LSIL CYTOMORPHOLOGY
Classic koilocytes,
as seen in this LSIL

“Nonkoilocytic”
LSIL. Nuclei are significantly
enlarged and show
mild hyperchromasia and
nuclear contour irregularity.
No definite koilocytes are seen
High-Grade Squamous Intraepithelial
Lesion (HSIL)
 Main molecular difference from LSIL and HSIL is the high expression of E6 and
E7 in HSIL.
 Virtually all women (90%) with an HSIL Pap result test positive for high-risk
HPV.
 If left untreated, it carries a significant risk of progression to cervical cancer.
 HSIL is usually a lesion of immature squamous cells.
HSIL CYTOMORPHOLOGY

 Clean or inflammatory background, no tumour diathesis

 Cells are smaller and show less cytoplasmic maturity than LSIL.
 HSIL consists of quite small parabasal-type cells.
 Cells occur singly, in sheets, or in syncytial-like aggregates - cells with no visually
discernable cytoplasmic borders between the cells and loss of nuclear polarity
within the groups.
 Nuclear enlargement is more variable than that seen in LSIL (but nuclei similar
size), and cytoplasmic area is decreased, leading to a marked increase in the N/C
ratio.
 Other cells have very high N/C ratios, but the actual size of the nuclei may be
smaller than that of LSIL
 Nucleus generally occupies ≥½ of the total area of the cell
HSIL CYTOMORPHOLOGY

 Nuclei are generally hyperchromatic but may be normochromatic or even

hypochromatic.
 Chromatin may be fine or coarse and is evenly distributed.
 Nuclear membrane is quite irregular and frequently demonstrates prominent
indentations or grooves.
 Nucleoli are generally inconspicuous.
 Appearance of the cytoplasm is variable; it can appear “immature,” lacy, and
delicate or densely metaplastic; occasionally, the cytoplasm is “mature” and
densely keratinized (keratinizing HSIL).
HSIL CYTOMORPHOLOGY
HSIL: Cells showing variably sized, ovoid nuclei with
prominent
nuclear grooves. In this case, the chromatin is not
particularly hyperchromatic, and cytoplasm has
ill-defi ned borders.

Keratinizing HSIL
ATYPICAL SQUAMOUS CELLS - ASC

 ASC refers to cytologic changes suggestive of SIL, but which are qualitatively
(features) or quantitatively (number of cells) insufficient
 Findings that are most consistent with benign reactive changes should be
classified as negative for intraepithelial malignancy (NILM) when possible.
 ASC requires:
-squamous differentiation
- increased N/C ratio,
-minimal nuclear changes which may include hyperchromasia, chromatin
clumping, irregularity, smudging, and/or multinucleation. Abnormal-
appearing nuclei are a prerequisite for the interpretation of ASC.
ASCUS

 ASC-US: Atypical squamous cells of undetermined significance.

 Changes suggesting LSIL, but are qualitatively or quantitatively not enough for
a definitive diagnosis.
 Unfortunately there is no magic cut-off number between ASC-US and LSIL
(usually <10) and each case is unique.
 Nuclei are approximately 2½ to 3x ICN
 Slightly increased N/C ratio.
 Minimal nuclear hyperchromasia and irregularity in chromatin distribution or
nuclear shape.
 Smooth or slightly irregular nuclear membrane
ASC-H

 Atypical squamous cells of undetermined significance, cannot rule out HSIL.

 Changes suggesting HSIL, but are qualitatively or quantitatively not enough
for a definitive diagnosis.
 Cells are usually sparse and typically have the size and shape of parabasal or
metaplastic cells.
 Cells usually occur singly or in small groups of less than ten cells; cells may
“stream” in strands of mucus.
 Cells are the size of metaplastic cells with nuclei that are about 1.5–2.5 times
larger than ICN (generally smaller than that of HSIL).
 N/C ratio is approximately that of HSIL