[215] Delayed Formalin Fixation Effect on Breast Biomarkers

T Khoury, S Sait, G Wilding, R Chandrasekhar, N Daoud, S Kulkarni. Roswell Park Cancer Institution, Buffalo

Background: Delayed formalin fixation (DFF) may invalidate hormone-receptor and HER2 analyses. Invalid results of tumor markers could significantly impact the type of adjuvant therapy a patient receives and potentially impact outcome. The purpose of this study was to determine the effects of progressive DFF on breast biomarkers.
Design: Ten palpable invasive breast cancers were resected and underwent immediate gross evaluation. For each case, the procured tumor was divided into 8 parts and consecutively fixed after 0, 10, 30 minutes, 1, 2, 4, and 8 hours; one section was kept in saline and stored overnight at 4C (ON). Two tissue microarray blocks were constructed. ER, PR, HER2 immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) were perfomred. Q-score was used to evaluate ER and PR. For HER2 FISH and IHC, the ASCO-CAP (2007) recommendations were followed. HER2 FISH was considered invalid whe at least one of the following was present, vague cellular outline, non-uniform signal, and/or poor nuclear resolution in more than 75% of the tumor cells. Statistical analyses including non-parametric sign test and exact McNemar's test were used.
Results: All ten cases were invasive ductal carcinomas. Q-score 6 was identified in 5 cases for ER and 4 for PR. Mean Q-score started to decline at 2 hr mark for ER and 1 hr mark for PR. Lowest score was at 8 hr mark for ER and ON for PR. HER2 FISH started to be invalid for interpretation at 2 hr mark (p < 0.008).
Conclusions: Delayed formalin fixation more than 1 hr has significant effect on HER2 FISH. Although changes due to DFF were not statistically significant for ER and PR, it is recommended not to delay fixation for more than 1 hr as well and not to store specimens ON.

Results of biomarkers changes with time.

	ER*	PR*	HER2 FISH
	Mean Q score (range)	Mean Q score (range)	# valid cases	p-value
0 min	7 (7-7)	6.75(6-7)	10	NS
10 min	7 (7-7)	6.75(6-7)	10	NS
30 min	7 (7-7)	6.75(6-7)	10	NS
1 hr	7 (7-7)	6 (4-7)	10	<0.008**
2 hr	6.8 (6-7)	6 (4-7)	2	<0.008
4 hr	6.4 (6-7)	6 (4-7)	2	<0.008
8 hr	5.6 (3-7)	5.75 (4-7)	2	<0.008
ON	5.8 (5-7)	5.25 (3-7)	1	<0.004

*p-value for ER and PR was calculated using sign test (not significant). ** p-value for HER2 FISH was calculated using exact McNemar test. NS, not significant.


Category: Breast

Wednesday, March 11, 2009 1:00 PM

Poster Session VI # 32, Wednesday Afternoon