Garcia-Casado et al. BMC Cancer 2010, 10:36 http://www.biomedcentral.com/1471-2407/10/36 The major biological characteristic of BC is that two thirds of cases express estrogen (ER) and/or progester- one receptors (PgR) [2,6,7], endocrine manipulation being an effective treatment for these patients. Tamoxi- fen, a selective ER modulator, has for decades been the gold standard therapy in ER/PgR [+] BC patients [8]. However, in postmenopausal patients, aromatase inhibi- tors like letrozole have consistently demonstrated to be more effective than tamoxifen [9]. Letrozole is highly specific for the aromatase enzyme and inhibits whole- body aromatization of almost 99%, providing a highly and essentially complete withdrawal of estrogen in post- menopausal women [10]. Neoadjuvant hormonal therapy is a safety option for postmenopausal women with large operable or locally advanced hormone sensitive BC, increasing the possibili- ties of breast-conserving surgery (BCS) [11]. In this con- text, letrozole has shown to be more effective than tamoxifen, obtaining higher response and BCS rates fol- lowing 4 months of preoperative therapy [12,13]. How- ever, less than 60% of patients will respond to this optimal endocrine therapy [14]. To date, only the pre- sence and intensity of hormonal receptors (ER/PgR) are useful tools as predictive markers in clinical practice. Identification of more accurate markers for more effi- cient patient selection to exclude non-responsive cases remains crucial. Genetic polymorphisms in the CYP19A1 gene have been associated with altered sex hormone levels in serum and urine [15-17], providing an explanation for an elevated risk for BC in relation to estrogen expo- sure. For example, the tetranucleotide repeat poly- morphism in intron 4 (TTTA)n has been associated with BC risk in initial studies [18-22]. The polymorph- ism rs10046 in the 3’ untranslated region (3’-UTR) also showed inconsistent associations with BC risk [23-26] and has also been related with tumor stage [25], circulating sex hormone levels [15], HER2 status and disease free and overall survival [27,28]. rs4646 has been reported to be associated with circulating steroid hormone levels [17] and, as in rs10046, with the HER2 status of the tumor [27]. Hence, it is biologi- cally plausible that the polymorphisms in the CYP19A1 gene may be associated with the response to aromatase inhibitors. Indeed, recently the rs4646 CYP19A1 poly- morphism has been associated with letrozole efficacy in advance disease [29]; however, no evidence between CYP19A1 polymorphisms and therapeutic efficacy of aromatase inhibitors in early stage BC has yet been established. In the present study, we perform a genetic analysis of three CYP19A1 polymorphisms in a series of postmeno- pausal endocrine-sensitive BC patients treated with neoadjuvant letrozole and describe their association Page 2 of 11 both with radiological response at 4 months (RR4M) of treatment and with progression free survival (PFS). Methods Patient selection and DNA samples We identified a total of 153 postmenopausal patients treated in our institution with letrozole as primary sys- temic therapy for stage II - III BC between April 1999 and March 2008. Only those cases which met the fol- lowing criteria were eligible for the study: 1) histological diagnosis of invasive BC by tru-cut or biopsy; 2) positive hormonal receptors status measured by immunohisto- chemistry (> 10% ER and/or PgR); 3) preserved tumor block; 4) bidimensionally measurable disease by mam- mogram and/or breast ultrasound; 5) adequate radiolo- gical follow-up at 4 months (± 14 days); 5) treatment with letrozole (Femara: Novartis Pharma AG, Basel, Switzerland) as neoadjuvant therapy by 2.5 mg/day for a minimum of 4 months in the absence of progression or side effects; and 6) patients able to provide a blood sample. Finally, 95 patients fulfilled all criteria, were contacted by their physician (AGZ, ALC, ARS, JG, MAC, VG) and agreed to participate on the study. All patients gave their written consent for the use of their DNA for this specific research proposal. In addition, this study was reviewed and approved by the FIVO Science and Ethics Committees. Peripheral blood samples (7 ml) were obtained from these patients and processed for DNA extraction at the Laboratory of Molecular Biology. Response to treatment at four months was done using the WHO criteria: complete response (CR) required the complete disappearance of all disease, partial response (PR) was defined as a reduction of ≥ 50% in tumor volume and stable disease (SD) was any reduction < 50% in tumor volume. The response evaluation was done by an independent radiologist (JCD). Pre-treatment study included patient characteristics, menopausal status, disease history, histological grade and diagnosis, evaluation of hormone receptor status, HER2 and Ki67 status by immunohistochemical analy- sis on core-cut biopsies, ECOG performance status and tumor assessment. The clinical data were prospec- tively reviewed and stored within a specific database. Histological and immunohistochemical analysis Core-cut biopsies were taken with a 14-guage needle before treatment and fixed in neutral buffered formalin for histological studies. The breast tumors were graded according to the modified Bloom and Richardson score on H&E-stained slides [30]. Four-micrometer sections from embedded blocks were cut on poly-L-lysine-coated slides, and dewaxed,