EGFR And AKT1 Gene Copy Numbers By Quantitative Multi-Gene Fluorescence In Situ Hybridization Impact On Prognosis In Breast Cancer

Objective: 1. The primary goals of this study were to evaluate the rates of EGFR and AKT1 copy number alterations in breast cancers by the quantitative multi-gene fluorescence in situ hybridization(QM-FISH). 2. To analyze the relationship between EGFR and AKT1 gene copy number and the clinical pathological characteristics in breast cancer patients. 3. To investigate the rates of EGFR and AKT1 gene copy number alterations in the prognosis prediction and thus to select patients for anti-EGFR/AKT1-targeted therapy. 4. To evaluate the combining effects of EGFR inhibitor erlotinib and AKT1 inhibitor LY294002 in vitro in EGFR/AKT1 MCF-7 breast cancer cellsMethods: 1. This research selected 205 female breast cancer patients with definite pathological diagnosis from August 2007 to August 2008 by Tianjin Medical University Cancer Institute & Hospital. We selected excised tumor paraffin pathologic specimens and pathologic testing results.QM-FISH was used to evalute EGFR and AKT1 gene copy number. 2. In vitro studies, the inhibitory effect of EGFR inhibitor erlotinib or AKT1 inhibitor LY294002 and combination treatments of the both in EGFR/AKT1 MCF-7 breast cancer cells were quantified by MTT assays.Results: 1. High EGFR gene copy number was observed in 71/205(34.6%) of cases and from the total of 205 tumors, 57(27.8%) presented high AKT1 copy number by QM-FISH. Overall, 24(11.7%) of the cases was identified with EGFR/AKT1-cohighed gene copy number, 101(49.3%) of cases were negative for both. 2. AKT1 and EGFR gene copy number has no relation with the patient age, tumor stage, histologic grade, ER, PR, HER2 status, molecular subtype, Ki67 and p53 expression.3. Although changes in EGFR and AKT1 gene copy numbers had no correlation with patient’s age, tumor stage, histological grade and the expression status of other molecular makers, high EGFR(P=0.0002) but not AKT1(P=0.1177) gene copy numbers correlated with poor 5-year overall survival(OS). The patients with co-heightened EGFR/AKT1 gene copy numbers displayed a poorer prognosis than tumors with only high EGFR gene copy numbers(P=0.0383). Both Univariate(U) and COX multivariate(C) analyses revealed that high EGFR and AKT1 gene copy numbers [(P=0.000(U), P=0.0001(C)], similar to histological grade [P=0.001(U), P=0.012(C)] and lymph node metastasis [P=0.046(U), P=0.158(C)], were independent prognostic indicators of 5-year OS. 4. The preliminary results showed that combined administrations of EGFR and AKT1 inhibitors exhibited an additive and dose-dependent suppression of MCF-7/EGFR+AKT1 cell growth compared to individual treatments.Conclusion: 1. These results indicate that high EGFR and AKT1 gene copy numbers were relatively frequent in breast cancer. 2. The changes in EGFR and AKT1 gene copy numbers had no correlation with patient’s age, tumor stage, histological grade and the expression status of other molecular makers. 3. Co-heightened EGFR/AKT1 gene copy numbers had a worse outcome than those with only high EGFR gene copy numbers. High EGFR and AKT1 gene copy numbers was independent prognostic indicator of 5-year OS. 4. Combined administrations of EGFR and AKT1 inhibitors exhibited an additive and dose-dependent suppression of MCF-7 cell growth compared to individual treatments. This piece of preliminary data partially sustains our speculation that a combination of anti-EGFR and anti-AKT1 treatment of these breast cancer patients may meliorate the 5-year OS...