The Efficacy Of Anthracycline-based Neoadjuvant Chemotherapy In Breast Cancer Patients

ObjectiveBreast carcinoma has been thought to be a systemic disease in recent years. Neoadjuvant chemotherapy(NAC)is commonly used as a systemic treatment of locally advanced breast cancer.NAC is usually given to downstage tumors and promotes higher conservative breast surgery rates and could be used to indicate suitable further therapy by evaluating NAC efficacy.Breast cancer is hormone-dependent tumor,under the control of estrogen and progesterone.The majority of breast cancer tissue expresses the estrogen and progesterone receptor(ER,PR),and a considerable portion of breast cancer exist the expression of HER-2 oncogene.Research in the expression of tumor biological markers in NAC treatment has been a hot topic in recent studies of cancer,but there is no agreement on the relationship between ER,PR,HER-2,p53 and NAC efficacy,Taxanes have emerged as fundamental drugs in the treatment of breast cancer and the targeted therapy drugs like Herceptin be used in the clinical work,but in our country,the status for anthracycline as the first-line drug used in the treatment for breast cancer can not be shaken.We carried out this retrospective study in order to further discuss the relationship between the status of ER,PR,HER-2,p53 protein expression and the NAC efficacy, and provide guidance for clinical treatment.Materials and Methods1,Materials(1)Patients Forty cases with locally advanced breast carcinoma(invasive carcinoma)were enrolled in the study.All of them were from the First affiliated Hospital of China Medical University from Jan 2006 to Jun 2008 and histologically diagnosed as breast carcinoma by core needle biopsy of the breast.Staging of tumor was defined referring to the International Tumor Node Metastasis classification.The median age was 50.4 (25-70).25 cases were≤50,15cases were＞50;27 cases were pre-menopause,13 cases were post-menopause.7 cases were stageⅡ,33 cases were stageⅢ.39 cases were infiltrating ductal carcinoma,1 case was infiltrating lobular carcinoma.None was found to have systemic metastasis.(2)NAC regimens3 cases received CE regimen,37 cases received CE regimen with the dosage of epirubicin 60～80mg/m2,,and the dosage was dependent on the patients' physical status.Chemotherapy was prescribed for the patients every 3 weeks.In fact,30 cases received 2 cycles,4 cases received 3cycles and 6cases received 4 or more cycles.(3)Assessment of clinical tumor response to NACBefore and after NAC,all cases were subject to the same measurement to evaluate the efficacy.33 cases take breast ultrasound examination,7cases take mammography examination.It was classified as a clinical complete remission(CR),partial remission(PR),stable disease(SD)or progressive disease(PD)according to standard International Union Against Cancer criteria.CR was defined as the disappearance of all clinical evidence of the tumor including the axillary site;partial remission was defined as a reduction of 50%or more in the diameters of the products of measured lesions, without the appearance of new lesions.SD was defined as a decrease of less than 50% in the diameters of the products of measured lesions or an increase of less than 25%, without the appearance of new lesions.Any measured increase greater than 25%or appearance of new lesions was defined as PD.CR plus PR was defined as objective response rate(OR).SD plus PD was defined as no response(NR). 2,Methods(1)mmunohistochemistry staining for ER,PR,p53 and c-erbB2 Serial sections 4-μm thick were taken from paraffin-embedded tissue for ER,PR, p53 and c-erbB2 immunostaining.Immunohistochemical study was performed following the manufacturer's instructions.Briefly,this procedure included the deparaffinization and rehydration steps,followed by an epitope retrieval step.The slide was then subjected to a series of alternating washes in tris(hydroxymethyl) aminomethane hydrochloride buffer and incubation steps with,first,a peroxidase-blocking reagent and then with monoclonal antibody for ER monoclonal antibody for PR,monoclonal antibody for p53,and monoclonal antibody for c-erbB2, followed by a visualization reagent for 30 minutes each,and finally incubated with a 3,3'-diaminobenzidine chromogen solution.After a final wash,the slide was counterstained with hematoxylin.For ER,PR and p53 nuclear staining of invasive tumor cells was scored as positive.The threshold for ER,PR and p53 was 10%. Scoring of c-erbB2 results was done as suggested for the HercepTest(DAKO)using the following categories:(-),negative result or membrane staining in＜30%tumor cells; (+),weak and incomplete membrane staining in＞30%tumor cells;(++),weak or moderate,complete membrane staining in＞30%tumor cells;(+++),strong,complete membrane staining in＞30%tumor cells.In this study,the patients with c-erbB2(-)and (+)were thought to be HER-2 gene non-amplified;the patient c-erbB2(++)and(+++) tissues were chosen for HER-2 gene amplification testing by Fluorescence In Situ Hybridization(FISH).(2)FISH detect HER-2 gene amplificationFISH was performed on formalin-fixed,paraffin-embedded tissue specimens from patients using the PathVysion kit(Vysis,Downers Grove,IL,USA).This kit includes probes to the HER-2 gene locus at 17q11.2-12(labeled with Spectrum Orange)and to the centromeric region of chromosome 17(CEP17;labeled with Spectrum Green). Briefly,unstained 4μm thick paraffin sections were cut from blocks and placed on positively charged slides.The slides were placed in an oven at 94℃for approximately 5 hours,deparaffinized in xylene,and dehydrated in a series of ethanol washes.After pretreatment in 0.2 N hydrochloric acid and sodium thiocyanate solutions,digestion in a protease solution for 16 minutes,and fixation in 10%neutral buffered formalin,the slides were subjected to denaturation and hybridization with 10μL of the PathVysion probe/buffer mixture.Fluorescence microscopy integrated with a cooled CCD camera system and Smart Capture software(CytoVision Chromophour System;Applied Imaging Ltd.,Carlsbad,CA,USA)for chromosome arrangement was used to investigate and analyze the FISH results.FISH analysis was performed according to the manufacturer's instructions.Count 30 cells,If Ratio value＜1.8,the case was thought to be HER-2 gene non-amplified;If Ratio value＞2.2,the case was thought to be HER-2 gene amplified;If Ratio value was 1.8~2.2,count 100 cells or redo FISH experiment.ResultsAll patients' clinical tumor response was assessed after NAC according to the International Union Against Cancer criteria.CR was observed in 4 patients(10%),1 case was pCR,PR was observed in 26 cases(65%),SD 9 cases(22.5%),PD 1 case(2.5%).out of 40 patients,whereas NR was observed in 10 patients(30.6%).No significant deviation was observed between these two groups' clinical tumor response to NAC.HER-2 gene non-amplication status and p53-negative status were correlated with a higher OR(P=0.029,P=0.041).The CR rates of patients received≥4 cycles NAC was 33.33%,but the CR rates of patients received＜4 cycles NAC was 5.88%, The CR rates of patients received≥4 cycles NAC was significantly higher than that of＜4 cycles(P＜0.05).In this group,the patients who obtained CR were received three cycles or more NAC.ConclusionsThe patients with HER-2 gene non-amplication and p53 overexpression may sensitive to anthracycline-based chemotherapy,and suitable increased NAC cycles may increase CR rates.