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[966] Combined In Situ Hybridization and Immunohistochemistry for the Detection of Human Papillomavirus (HPV) Infection in Penile Carcinomas

George J Netto, Antonio L Cubilla, Rajni Sharma, Jessica Hicks, Kristen L Lecksell, Alcides Chaux. Johns Hopkins University, Baltimore, MD; Instituto de Patologia e Investigacion, Asuncion, Paraguay

Background: Approximately one half of penile carcinomas are HPV positive. Polymerase chain reaction (PCR) assay is the gold standard for HPV detection. Alternative methods of detection that are less technically demanding and less costly are desirable especially in developping countries. Herein, we evaluated a combination of in situ hybridization and immunohistochemistry for defining HPV status in penile cancers.
Design: Forty-eight cases of penile squamous cell carcinoma (SCC), with previously determined HPV status by SPF-10 PCR assay, were included in a set of 4 tissue microarrays (TMA). Presence of high-risk HPV (HR-HPV) was assessed by in situ hybridization (ISH). Additionally, immunoexpression of 5 cell cycle control markers (p53, p16^INK4a, MDM2, cyclin-D1, and Ki-67) was evaluated. TMA spots were scanned using the APERIO system and uploaded to the TMAJ platform (http://tmaj.pathology.jhmi.edu). Discrimination of markers for defining HPV status was estimated using receiver operating characteristic (ROC) curves, and performances were compared by changes in the area under the ROC curve (AUC).
Results: Positive HR-HPV by ISH and increased p16^INK4a and Ki-67 expression were significantly associated with HPV status (P < .008). ROC analyses are shown in the table, with 95% confidence intervals in parenthesis.

% Sensitivity % Specificity AUC
p16INK4a alone 65 (38, 86) 90 (74, 98) 0.77 (0.65, 0.90)
HR-HPV ISH alone 47 (23, 72) 100 (89, 100) 0.73 (0.61, 0.86)
Ki-67 alone 65 (38, 86) 71 (52, 86) 0.68 (0.54, 0.82)
p16INK4a and HR-HPV ISH 35 (14, 62) 100 (89, 100) 0.68 (0.56, 0.79)
p16INK4a or HR-HPV ISH 76 (50, 93) 90 (74, 98) 0.83 (0.72, 0.95)
p16INK4a and Ki-67 59 (33, 82) 94 (79, 99) 0.76 (0.63, 0.89)
p16INK4a or Ki-67 71 (44, 90) 68 (49, 83) 0.69 (0.55, 0.83)
Ki-67 and HR-HPV ISH 35 (14, 62) 100 (88, 100) 0.68 (0.56, 0.79)
Ki-67 or HR-HPV ISH 76 (50, 93) 71 (52, 86) 0.74 (0.60, 0.87)

Conclusions: As a stand-alone marker, p16^INK4a performed better than HR-HPV ISH or Ki-67 to define presence of HR-HPV. Specificity was high for p16^INK4a and HR-HPV ISH positivity (90% and 100% individually; 100% when both positive). Positivity for either p16^INK4a or HR-HPV ISH (rather than requiring both to be positive) had the best AUC performance. The latter combination could serve as an acceptable surrogate to the gold standard PCR assay.
Category: Genitourinary (including renal tumors)

Monday, March 19, 2012 11:30 AM

Platform Session: Section A, Monday Morning

	% Sensitivity	% Specificity	AUC
p16INK4a alone	65 (38, 86)	90 (74, 98)	0.77 (0.65, 0.90)
HR-HPV ISH alone	47 (23, 72)	100 (89, 100)	0.73 (0.61, 0.86)
Ki-67 alone	65 (38, 86)	71 (52, 86)	0.68 (0.54, 0.82)
p16INK4a and HR-HPV ISH	35 (14, 62)	100 (89, 100)	0.68 (0.56, 0.79)
p16INK4a or HR-HPV ISH	76 (50, 93)	90 (74, 98)	0.83 (0.72, 0.95)
p16INK4a and Ki-67	59 (33, 82)	94 (79, 99)	0.76 (0.63, 0.89)
p16INK4a or Ki-67	71 (44, 90)	68 (49, 83)	0.69 (0.55, 0.83)
Ki-67 and HR-HPV ISH	35 (14, 62)	100 (88, 100)	0.68 (0.56, 0.79)
Ki-67 or HR-HPV ISH	76 (50, 93)	71 (52, 86)	0.74 (0.60, 0.87)

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