[1312] Partial p16 Immunoreactivity in Oropharyngeal Squamous Cell Carcinoma – Extent and Pattern of Staining Correlate with the Presence of Transcriptionally-Active Human Papillomavirus

James S Lewis, Rebecca D Chernock, Xiao-Jun Ma, John J Flanagan, Yuling Luo, Xiaowei Wang, Tian Zhang, Wade L Thorstad, Samir K El-Mofty. Washington University, St. Louis, MO; Advanced Cell Diagnostics, Inc., Hayward, CA

Background: Human papillomavirus (HPV)-related oropharyngeal squamous cell carcinoma (SCC) is a tumor with unique biology and better outcomes. p16 immunostaining is extensively used as a surrogate marker for transcriptionally-active HPV. While diffuse staining is generally accepted as being positive, the significance of partial staining has not been established, nor has the cutoff for extent of p16 staining that should be used to identify a tumor as HPV-related or unrelated.
Design: From 3 other large studies (>400 cases) utilizing p16 immunohistochemistry, we identified all cases with partial positive staining. The p16 stained slides were reviewed by 3 study pathologists for staining (both nuclear and cytoplasmic) extent (in quartiles) and also for % of staining that was confluent (i.e. back to back positive cell staining). Tumors were histologically typed (keratinizing, nonkeratinizing, or nonkeratinizing with maturation) and tested for high risk HPV by RNA in situ hybridization (ISH) and reverse transcriptase PCR.
Results: For the 16 cases, there were 2 4+ (13%), 5 3+ (31%), 6 2+ (38%), and 3 1+ (19%) p16 staining tumors. Extent of staining ranged from 5 to 90% of cells positive with 25% or more confluent staining in 4/16 (25%). Table 1 shows histology, HPV testing, and p16 results by quartile.

Table 1 - HPV Testing by p16 Staining Extent

p16	HPV RNA ISH	HPV RTPCR	Nonkeratinizing	p16 Staining (avg %)	p16 Confluent Staining (25% cutoff)
1+ (0-25%)	0/3	1/3	0/3	12	0/3
2+ (26-50%)	0/6	0/5	0/6	35	0/6
3+ (51-75%)	3/5	2/4	2/5	61	3/5
4+ (76-100%)	2/2	2/2	2/2	85	1/2

Five of 16 (31%) cases were HPV-related based on RNA ISH and RT-PCR. All of these cases had >50% staining, 4/5 (80%) had more than 25% confluent staining, and 4/7 (57%) had nonkeratinizing histologic features. Only 1 of the p16 1+/2+ tumors was HPV RNA positive (by RTPCR only and low level). All 1+/2+ cases were keratinizing type or undifferentiated.
Conclusions: By sensitive detection methods, most partial p16 positive SCC cases with >50% staining harbor transcriptionally-active HPV, and most HPV+ tumors have significant amounts of confluent staining. Cases with <50% p16 staining and lacking significant confluent staining rarely harbor HPV. Thus, our study suggests that >75% p16 staining or, alternatively, >50% staining combined with >25% confluent areas, are suitable criteria for use for positivity for p16 in OSCC.
Category: Head & Neck

Wednesday, March 21, 2012 9:30 AM

Poster Session V # 182, Wednesday Morning