Comparison of quantitative immunofluorescence with conventional methods for HER2/neu testing with respect to response to trastuzumab therapy in metastatic breast cancer.
BACKGROUND
Selection for trastuzumab therapy depends on a companion diagnostic assessment of HER2 by either immunohistochemistry (IHC) for protein overexpression or fluorescence in situ hybridization (FISH) to detect gene amplification. Although many studies have compared IHC to FISH, few have compared the tests to the true gold standard, tumor response.


OBJECTIVE
To compare HER2 testing by FISH and IHC along with a third immunofluorescence-based assay (automated quantitative analysis-tissue microarray [AQUA-TMA]) and to assess the value of each test for prediction of response to trastuzumab.


METHODS
Immunohistochemistry and FISH assays were done on both whole slides (IHC-WS and FISH-WS) and on TMAs (IHC-TMA and FISH-TMA). AQUA was only done on TMAs (AQUA-TMA). Response was assessed according to modified Response Evaluation Criteria in Solid Tumors.


RESULTS
AQUA-TMA scores showed a significant linear relationship to both the FISH signal ratio and IHC scores on whole sections and TMAs. Assay assessment by outcome showed no association between response and FISH-WS ratio (P = .96), FISH-TMA (P = .55), IHC-WS (P = .75), or IHC-TMA (P = .06), but a significant relationship between AQUA score and categoric response was observed (P = .01). Assessed as a function of outcome using models of logistic regression, both AQUA-TMA and IHC-TMA were equally significant (P = .01). FISH-WS was the most sensitive assay, with a significantly higher true-positive fraction than all other tests except AQUA-TMA, although it was the least specific. IHC-TMA was the most specific assay. The lowest misclassification rate was achieved using AQUA-TMA (0.30).


CONCLUSIONS
Both AQUA-TMA and IHC-TMA were substantially more predictive than the FISH or IHC-WS tests. Although these results are derived from a small retrospective series, they suggest that accurate measurement of protein expression and unbiased selection of tissue for measurement may be key factors in prediction of response.
Departments of Pathology, Yale University School of Medicine, New Haven, Connecticut 06520-8023 , USA.
Jennifer M
JM
Giltnane
Annette
A
Molinaro
Huan
H
Cheng
Andrew
A
Robinson
Dmitry
D
Turbin
Karen
K
Gelmon
David
D
Huntsman
David L
DL
Rimm
D000818:Animals; D000911:Antibodies, Monoclonal / Q000627:therapeutic use*; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000378:metabolism* / Q000556:secondary; D002460:Cell Line; D006224:Cricetinae; D005260:Female; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods*; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D016015:Logistic Models; D008875:Middle Aged; D011237:Predictive Value of Tests; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism*; D012189:Retrospective Studies; D012680:Sensitivity and Specificity; D046888:Tissue Array Analysis; D000068878:Trastuzumab; D016896:Treatment Outcome
D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D018719:Erb-b2 Receptor Tyrosine Kinases; D000068878:Trastuzumab