Histopathology

The histopathological classification of HPV-associated disease of the anogenital tract prior to 2012 was complex. Terminology changed for different sites and variable terms such as condyloma, dysplasia, intraepithelial neoplasia and carcinoma in-situ were employed. This was partly due to development of a number of different interest groups including pathologists, dermatologists and gynaecologists using their own version of terminology for similar lesions.

To address this problem and to improve communication between the specialties, the Lower Anogenital Squamous Terminology (LAST) Standardization Project for HPV-Associated Lesions was convened, including five major working groups. The findings were first published in June 2012,[1] with two further publications within the following 12 months in order to reach a broad audience.[2][3]

The LAST Standardization Project working groups recognised a number of general principles:[1]

• HPV-related squamous disease reflects a unified epithelial biology.
• Each cytological or histological sample provides only a statistical representation of the patient’s true biology; the more samples or data points available, the more accurate the assessment of the patient’s true biology.
• The true biology represents the risk for cancer at the current time, and the risk for cancer over time.
• Diagnostic variation can be improved by:
  • aligning the number of diagnostic terms with the number of biologically relevant categories
  • the use of biological markers.

The findings of the LAST project have been widely accepted and adopted by members of the Royal College of Pathologists of Australasia (RCPA) and the World Health Organization (WHO)[4], and have been referenced in the first edition (2013) of the RCPA protocol for structured reporting of cervical carcinoma[5].

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A two-tiered nomenclature system has been accepted for non-invasive HPV associated squamous proliferations of cervix and lower anogenital tract. The two groups are LSIL and HSIL. These two groups may be further characterised by the applicable intraepithelial neoplasia or –IN subcategory. The nomenclature addresses pre-invasive mucosal lesions and early invasive mucosal lesions.

LSIL is the morphologic expression of acute HPV infection and is characterised by cells with increased nuclear to cytoplasmic ratios and irregular nuclear membranes. Mitoses are limited to the lowermost third of the epithelium and maturation begins in the middle third. Multi-nucleation and perinuclear halos, characteristic of HPV effect, may be seen. LSIL encompasses changes previously called ‘HPV effect’ and CIN1 (pre-AMBS 2004). The LAST terminology does not support the distinction between these two categories because of poor inter-observer agreement, lack of clinical significance and the common underpinning biology. In cases with LSIL morphology, p16 staining should not be performed.

HSIL is the morphologic expression of persistent HPV infection that has the potential to progress to invasive carcinoma. It is characterised by mitoses seen at any level of the epithelium, little to no cytoplasmic differentiation in the middle third and upper third, increased nuclear size, and irregular nuclear membranes. Where the pathologist is considering a diagnosis of CIN2, p16 staining should be performed.

The result for p16 is reported as positive if there is strong and diffuse block staining for p16. In squamous epithelia, this is defined as continuous strong nuclear, or nuclear plus cytoplasmic, staining of the basal cell layer with extension upward involving at least one-third of the epithelial thickness.[2] The LAST Standardization Project group notes that this height restriction is somewhat arbitrary but adds specificity, and that full-thickness staining or extension into the upper third or upper half is specifically not required to call a specimen positive.[2]

When the p16 stain is negative the lesion is either LSIL or a mimic of HSIL and, accordingly, should not be diagnosed as HSIL.

HSIL encompasses lesions previously called ‘CIN2’ and ‘CIN3’. The following subcategories should continue to be used:

• HSIL (CIN2) – used when p16 positive
• HSIL (CIN3).

This practice will enable continued measurement of the prevalence of CIN3 in Australian women during and after transition to the renewed NCSP.

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The term ‘microinvasive carcinoma’ is no longer recommended, and the term ‘superficially invasive squamous cell carcinoma’ (SISCCA) should be used instead. It is recognised that SISCCA has a favourable prognosis.

A report including the finding of SISCCA must include a comment on the presence or otherwise of lymphatic invasion and a comment on the number and size of multifocal carcinomas, once the presence of a single carcinoma is excluded. The International Federation of Gynecology and Obstetrics (FIGO) staging should be based on the highest FIGO stage of an individual focus, rather than adding multiple foci together.[6]

Squamous cell carcinoma is an invasive epithelial tumour showing variably differentiated squamous cells.[7] The majority are of large cell keratinizing type, demonstrating sheet-like growth, with surrounding desmoplastic stromal response. Virtually all SCCs are thought to arise from a pre-cancerous intraepithelial lesion (HSIL).[7]

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The LAST group assessed data on p16, Ki-67 (Mib1), ProEx C, L1, HPV 16/18 mRNA, telomerase/TERC, and HPV genotyping. It concluded that p16 was the only biomarker for which there was sufficient evidence to recommend its use for distinguishing between types of mucosal pre-cancerous lesions. Staining for p16 is particularly useful for lesions with inflammation or atrophy, lesions affected by diathermy, and thin lesions.

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Adenocarcinoma in-situ (AIS)

‘Adenocarcinoma in situ’ (AIS) is the only currently recommended term in Australasia for glandular mucosal pre-invasive lesions.[8] The term ‘glandular dysplasia’ is not currently used in Australia, but has been used historically and is in use in the United Kingdom (where the synonym is ‘low grade cervical glandular intraepithelial neoplasia’).[9]

AIS is an intraepithelial lesion containing malignant-appearing glandular epithelium. Nuclei are hyperchromatic and stratified, mitoses are frequently seen towards the apex of the cell, and apoptotic debris may be seen towards the base of the cell. The use of p16 can be very effective in distinguishing AIS from mimics including tubular metaplasia following previous surgical treatment.[9]

Invasive adenocarcinoma of cervix

Adenocarcinoma of cervix is an invasive epithelial tumour showing glandular differentiation.[7] There is significant morphological overlap with adenocarcinoma in situ. However, clues to the invasive nature of the lesion include a desmoplastic stromal response, small angulated claw-like glands branching from the areas of AIS and complex architectural proliferations such as solid, papillary cribriform and labyrinthine growth patterns.[10] Several subtypes are currently recognised.[7][11]

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