Classifying Anal Intraepithelial Neoplasia 2 Based on LAST Recommendations

Abstract

Objectives: The Lower Anogenital Squamous Terminology (LAST) recommendations classify human papillomavirus-associated squamous lesions into low- and high-grade squamous intraepithelial lesions (LSILs/HSILs). Our study aimed to assess interobserver agreement among 6 experienced pathologists in assigning 40 anal lesions previously diagnosed as anal intraepithelial neoplasia 2 (AIN 2) to either HSIL or non-HSIL categories.

Methods: Agreement based on photomicrographs of H&E alone or H&E plus p16 immunohistochemistry was calculated using κ coefficients.

Results: Agreement was fair based on H&E alone (κ = 0.42; 95% confidence interval [CI], 0.34-0.52). Adding p16 improved agreement to moderate (κ = 0.55; 95% CI, 0.54-0.62). On final diagnosis, 21 cases (53%) had unanimous diagnoses, and 19 (47%) were divided. When designating p16 results as positive or negative, agreement was excellent (κ = 0.92; 95% CI, 0.83-0.95). Among variables (staining location, extent, and intensity), staining of the basal/parabasal layers was a consistent feature in cases with consensus for positive results (20/20). Of the 67 H&E diagnoses with conflicting p16 results, participants modified 32 (48%), downgrading 23 HSILs and upgrading 9 non-HSILs.

Conclusions: Although p16 increased interobserver agreement, disagreement remained considerable regarding intermediate lesions. p16 expression, particularly if negative, can reduce unwarranted HSIL diagnoses and unnecessary treatment.

Keywords: Anal intraepithelial neoplasia 2; Human papillomavirus; Interobserver agreement; p16 Immunohistochemistry.

Image 1

Interpretation of p16 immunohistochemistry (IHC) results. A, Case 15 with diffuse, continuous staining of basal and parabasal layers with upward extension. All participants interpreted as positive. Positive to negative: 6:0. B, Case 7 with discontinuous staining throughout the epithelium but not basal and parabasal layers. All participants interpreted as negative. Positive to negative: 0:6. C, Case 9 with focal staining in the basal and parabasal layers. Two participants interpreted as positive, and 4 interpreted as negative. Positive to negative: 2:4. D, Case 13 with patchy staining of the epithelium, including limited basal and parabasal staining. Five participants interpreted as positive, and 1 interpreted as negative. Positive to negative: 5:1. (p16 IHC, original magnification ×100.)

Image 2

Impact of p16 immunohistochemistry (IHC) results on final diagnosis. A, Case 28 was diagnosed as high-grade squamous intraepithelial lesion (HSIL) by all 6 participants based on H&E morphology. B, Given the negative p16 result, 4 participants downgraded it to non-HSIL, and 2 maintained the HSIL diagnosis. C, Case 11 was diagnosed as non-HSIL by all 6 participants based on H&E morphology. D, Given the positive p16 result, 3 participants upgraded it to HSIL, whereas 3 maintained the non-HSIL diagnosis. (A and C, H&E, ×200; B and D, p16 IHC, ×200.)

Image 3

Lesions originating from anal transitional zone. A, Case 19 was diagnosed as high-grade squamous intraepithelial lesion (HSIL) by 3 participants and non-HSIL by 3 based on H&E morphology. B, Given the positive p16 result, 5 participants diagnosed it as HSIL, and 1 maintained a non-HSIL diagnosis. C, Case 3 was diagnosed as HSIL by 4 and non-HSIL by 2 based on H&E morphology. D, Given the negative p16 result, 5 participants diagnosed it as non-HSIL, whereas 1 maintained HSIL. (A and C, H&E, ×200; B and D, p16 immunohistochemistry, ×100.)