Identification and Validation of an Anthracycline/Cyclophosphamide–Based Chemotherapy Response Assay in Breast Cancer

Mulligan, Jude M.
Hill, Laura A.
Deharo, Steve
Irwin, Gareth
Boyle, David
Keating, Katherine E.
Raji, Olaide Y.
McDyer, Fionnuala A.
O’Brien, Eamonn
Bylesjo, Max
Quinn, Jennifer E.
Lindor, Noralane M.
Mullan, Paul B.
James, Colin R.
Walker, Steven M.
Kerr, Peter
James, Jacqueline
Davison, Timothy S.
Proutski, Vitali
Salto-Tellez, Manuel
Johnston, Patrick G.
Couch, Fergus J.
Paul Harkin, D.
Kennedy, Richard D.

Affiliations of authors: Almac Diagnostics, Craigavon, UK (JMM, LAH, SD, KEK, OYR, FAM, EO, MB, SMW, PK, TSD, VP, PGJ, DPH, RDK); Centre for Cancer Research and Cell Biology, Queen’ s University Belfast, Belfast, UK (GI, DB, JEQ, PBM, CRJ, JJ, TSD, MS-T, PGJ, DPH, RDK); Department of Health Science Research, Mayo Clinic, Scottsdale, AZ (NML); Department of Medical Genetics, Mayo Clinic, Rochester, MN (FJC).

Correspondence to: Richard D. Kennedy, MB, PhD, Centre for Cancer Research & Cell Biology, Queen’s University Belfast, 97 Lisburn Rd, Belfast, BT9 7BL, Northern Ireland, UK (e-mail: [email protected]).

Received March 12, 2012

Accepted October 10, 2013

Journal of the National Cancer Institute 106(1), January 2, 2014. | DOI: 10.1093/jnci/djt335

Background

There is no method routinely used to predict response to anthracycline and cyclophosphamide–based chemotherapy in the clinic; therefore patients often receive treatment for breast cancer with no benefit. Loss of the Fanconi anemia/BRCA (FA/BRCA) DNA damage response (DDR) pathway occurs in approximately 25% of breast cancer patients through several mechanisms and results in sensitization to DNA-damaging agents. The aim of this study was to develop an assay to detect DDR-deficient tumors associated with loss of the FA/BRCA pathway, for the purpose of treatment selection.

Methods

DNA microarray data from 21 FA patients and 11 control subjects were analyzed to identify genetic processes associated with a deficiency in DDR. Unsupervised hierarchical clustering was then performed using 60 BRCA1/2 mutant and 47 sporadic tumor samples, and a molecular subgroup was identified that was defined by the molecular processes represented within FA patients. A 44-gene microarray-based assay (the DDR deficiency assay) was developed to prospectively identify this subgroup from formalin-fixed, paraffin-embedded samples. All statistical tests were two-sided.

Results

In a publicly available independent cohort of 203 patients, the assay predicted complete pathologic response vs residual disease after neoadjuvant DNA-damaging chemotherapy (5-fluorouracil, anthracycline, and cyclophosphamide) with an odds ratio of 3.96 (95% confidence interval [Cl] =1.67 to 9.41; P = .002). In a new independent cohort of 191 breast cancer patients treated with adjuvant 5-fluorouracil, epirubicin, and cyclophosphamide, a positive assay result predicted 5-year relapse-free survival with a hazard ratio of 0.37 (95% Cl = 0.15 to 0.88; P = .03) compared with the assay negative population.

Conclusions

A formalin-fixed, paraffin-embedded tissue-based assay has been developed and independently validated as a predictor of response and prognosis after anthracycline/cyclophosphamide–based chemotherapy in the neoadjuvant and adjuvant settings. These findings warrant further validation in a prospective clinical study.