Stiffness at shear-wave elastography and patient presentation predicts upgrade at surgery following an ultrasound-guided core biopsy diagnosis of ductal carcinoma in situ.

AIM: The aim of this study is to establish predictors of invasion in lesions yielding an ultrasound-guided biopsy diagnosis of ductal carcinoma in situ (DCIS). MATERIALS AND METHODS: Patients subjected to ultrasound-guided core biopsy yielding DCIS were studied. At shear-wave elastography (SWE) a threshold of 50 kPa was used for mean elasticity (Emean) to dichotomise the elasticity data between invasive and non-invasive masses. Data recorded included the mammographic and ultrasound features, the referral source, and grade of DCIS in the biopsy. The chi-square test was used to detect statistical significance. RESULTS: Of 57 lesions, 24 (42%) had invasion at excision. Symptomatic patients and patients with stiff lesions were more likely to have invasion than patients presenting through screening and with soft lesions (58% [14 of 24] versus 30% [10 of 33], p=0.03) and (51% [20 of 39] versus 22% [4 of 18], p=0.04). No other factors showed a relationship with invasion. Combining the two predictors of invasion improved risk stratification with symptomatic and stiff lesions having a risk of invasion of 67% (12 of 18) and soft lesions presenting at screening having only a 17% (2 of 12) risk of invasion (p=0.02). CONCLUSION: Stiffness on SWE and the referral source of the patient are predictors of occult invasion in women with an ultrasound-guided core biopsy diagnosis of DCIS.

Viewpoint: Availability of oestrogen receptor and HER2 status for the breast multidisciplinary meeting discussion; time to get it right.

The efficacy and pivotal role of the multidisciplinary meeting (MDM) in informed decision making is well established. It aims to provide a forum in which clinical evidence combines with individual patient data to create a personalized treatment plan. It does not fulfil this role adequately when undertaken without the full results of the patient's investigations being available. Neither doctor nor patient can make an informed decision about treatment options without knowledge of the tumour receptor status. Both targeted therapies and the aim to treat a majority of patients within clinical trials must now drive MDM decision making to be based on accuracy and best available treatment choices. A fully informed decision on treatment delayed by 1-2 weeks is clearly preferable to rushed time target-driven decisions made without the patient being offered a fully informed choice as ratified by a multidisciplinary team. Whilst the early anxiety of waiting for all relevant information to be available may be stressful for patients, not being sure that they have been offered fully informed treatment choices is also stressful and could cause longer lasting anxiety both during and after treatment. MDMs need to develop (along with targeted therapies) to retain their role as a forum whereby patients receive a correct, but specifically a full diagnosis and allow a fully informed discussion of all treatment options, including pre-operative clinical trials.

Magnetic resonance imaging texture analysis classification of primary breast cancer.

OBJECTIVES: Patient-tailored treatments for breast cancer are based on histological and immunohistochemical (IHC) subtypes. Magnetic Resonance Imaging (MRI) texture analysis (TA) may be useful in non-invasive lesion subtype classification. METHODS: Women with newly diagnosed primary breast cancer underwent pre-treatment dynamic contrast-enhanced breast MRI. TA was performed using co-occurrence matrix (COM) features, by creating a model on retrospective training data, then prospectively applying to a test set. Analyses were blinded to breast pathology. Subtype classifications were performed using a cross-validated k-nearest-neighbour (k = 3) technique, with accuracy relative to pathology assessed and receiver operator curve (AUROC) calculated. Mann-Whitney U and Kruskal-Wallis tests were used to assess raw entropy feature values. RESULTS: Histological subtype classifications were similar across training (n = 148 cancers) and test sets (n = 73 lesions) using all COM features (training: 75%, AUROC = 0.816; test: 72.5%, AUROC = 0.823). Entropy features were significantly different between lobular and ductal cancers (p < 0.001; Mann-Whitney U). IHC classifications using COM features were also similar for training and test data (training: 57.2%, AUROC = 0.754; test: 57.0%, AUROC = 0.750). Hormone receptor positive and negative cancers demonstrated significantly different entropy features. Entropy features alone were unable to create a robust classification model. CONCLUSION: Textural differences on contrast-enhanced MR images may reflect underlying lesion subtypes, which merits testing against treatment response. KEY POINTS: • MR-derived entropy features, representing heterogeneity, provide important information on tissue composition. • Entropy features can differentiate between histological and immunohistochemical subtypes of breast cancer. • Differing entropy features between breast cancer subtypes implies differences in lesion heterogeneity. • Texture analysis of breast cancer potentially provides added information for decision making.

What are the characteristics of breast cancers misclassified as benign by quantitative ultrasound shear wave elastography?

OBJECTIVES: Shear wave elastography (SWE) is a promising adjunct to greyscale ultrasound in differentiating benign from malignant breast masses. The purpose of this study was to characterise breast cancers which are not stiff on quantitative SWE, to elucidate potential sources of error in clinical application of SWE to evaluation of breast masses. METHODS: Three hundred and two consecutive patients examined by SWE who underwent immediate surgery for breast cancer were included. Characteristics of 280 lesions with suspicious SWE values (mean stiffness >50 kPa) were compared with 22 lesions with benign SWE values (<50 kPa). Statistical significance of the differences was assessed using non-parametric goodness-of-fit tests. RESULTS: Pure ductal carcinoma in situ (DCIS) masses were more often soft on SWE than masses representing invasive breast cancer. Invasive cancers that were soft were more frequently: histological grade 1, tubular subtype, ≤10 mm invasive size and detected at screening mammography. No significant differences were found with respect to the presence of invasive lobular cancer, vascular invasion, hormone and HER-2 receptor status. Lymph node positivity was less common in soft cancers. CONCLUSION: Malignant breast masses classified as benign by quantitative SWE tend to have better prognostic features than those correctly classified as malignant. KEY POINTS: • Over 90 % of cancers assessable with ultrasound have a mean stiffness >50 kPa. • 'Soft' invasive cancers are frequently small (≤10 mm), low grade and screen-detected. • Pure DCIS masses are more often soft than invasive cancers (>40 %). • Large symptomatic masses are better evaluated with SWE than small clinically occult lesions. • When assessing small lesions, 'softness' should not raise the threshold for biopsy.

CYP2D6 genotype and adjuvant tamoxifen: meta-analysis of heterogeneous study populations.

The International Tamoxifen Pharmacogenomics Consortium was established to address the controversy regarding cytochrome P450 2D6 (CYP2D6) status and clinical outcomes in tamoxifen therapy. We performed a meta-analysis on data from 4,973 tamoxifen-treated patients (12 globally distributed sites). Using strict eligibility requirements (postmenopausal women with estrogen receptor-positive breast cancer, receiving 20 mg/day tamoxifen for 5 years, criterion 1); CYP2D6 poor metabolizer status was associated with poorer invasive disease-free survival (IDFS: hazard ratio = 1.25; 95% confidence interval = 1.06, 1.47; P = 0.009). However, CYP2D6 status was not statistically significant when tamoxifen duration, menopausal status, and annual follow-up were not specified (criterion 2, n = 2,443; P = 0.25) or when no exclusions were applied (criterion 3, n = 4,935; P = 0.38). Although CYP2D6 is a strong predictor of IDFS using strict inclusion criteria, because the results are not robust to inclusion criteria (these were not defined a priori), prospective studies are necessary to fully establish the value of CYP2D6 genotyping in tamoxifen therapy.

Progesterone receptor expression is an independent prognostic variable in early breast cancer: a population-based study.

BACKGROUND: Progesterone receptor (PR) expression assessment in early invasive breast cancer remains controversial. This study sought to re-evaluate PR expression as a potential therapeutic guide in early breast cancer; particularly in oestrogen receptor (ER)-positive, lymph node (LN)-negative disease. METHODS: A population cohort of 1074 patients presenting to a single Cancer Centre over 4 years (2000-2004) underwent surgery for primary invasive breast cancer with curative intent. Prospective data collection included patient demographics, pathology, ER and PR expression, HER2 status, adjuvant chemotherapy and endocrine therapy. Progesterone receptor expression was compared with (all causes) overall survival (OS), breast cancer-specific survival (BCSS) and disease-free survival (DFS). RESULTS: Overall survival was 71.0% and BCSS was 83.0% at median follow-up of 8.34 years. Absent PR expression was significantly associated with poorer prognosis for OS, BCSS and DFS (P<0.0001, log-rank), even within the ER-positive, LN-negative group (hazard ratio for BCSS 3.17, 95% CI 1.43-7.01) and was not influenced by endocrine therapy. Cox's regression analysis demonstrated that PR expression was an independent prognostic variable. CONCLUSION: Absence of PR expression is a powerful, independent prognostic variable in operable, primary breast cancer even in ER-positive, LN-negative patients receiving endocrine therapy. Absence of PR expression should be re-evaluated as a biomarker for poor prognosis in ER-positive breast cancer and such patients considered for additional systemic therapy.

HER2 testing for breast carcinoma: recommendations for rapid diagnostic pathways in clinical practice.

Human epidermal growth factor receptor 2 (HER2) testing is required for newly diagnosed breast cancer and advised for recurrent and metastatic breast cancer, to determine treatment planning using HER2-directed therapy in the neoadjuvant, adjuvant and advanced disease settings. Wide variation, nationally, in the turnaround time for HER2 testing may hinder equity of access for patients to both clinical trials and the timely implementation of HER2-directed therapy particularly in the neo-adjuvant setting. Process mapping from three recognised laboratories in the UK was applied to the logistics of HER2 testing in different geographic hub and spoke models. Consequently, recommendations for HER2 testing likely to facilitate access to clinical trials and timely patient care are presented.

Clinical value of internal mammary lymph node metastases found incidentally during free flap recipient vessel exposure.

INTRODUCTION: The clinical importance of internal mammary (IM) lymph node (LN) metastases in breast cancer remains unclear. The aim of this study was to determine the clinical value of opportunistic IMLN sampling at the time of free flap breast reconstruction using the IM recipient vessels and whether it affected the adjuvant treatment given. METHODS: A prospective study was conducted of IMLN exploration performed by a single surgeon as part of free flap breast reconstruction using IM recipient vessels. Patients where IMLNs were positive for metastatic disease were reviewed with the breast cancer multidisciplinary team for changes in therapy. RESULTS: 122 patients met the inclusion criteria, 111 of whom had immediate reconstructions. 63 patients had IMLNs excised, and of these 13 were positive for metastatic disease, all in immediate breast reconstructions. The adjuvant treatment given was changed in 1 patient with no axillary LN disease as a result of finding a positive IMLN. Positive IMLNs were significantly associated with larger tumours and axillary metastases, but not tumour location. All patients with positive IMLNs were alive at last follow-up with no local or distant recurrences, with mean disease-free survival of 20.5 months (5-56 months). CONCLUSION: Incidental IMLNs positive for metastatic disease were found in 13 of 122 free flap breast reconstructions and resulted in a change in adjuvant treatment in one patient. IMLNs found incidentally during recipient IM vessel exposure for free flap breast reconstruction therefore may upstage a patient's disease and influence the adjuvant treatment for their breast cancer and should therefore be submitted for pathological assessment.

Can additional histopathological examination of ultrasound-guided axillary lymph node core biopsies improve preoperative diagnosis of primary breast cancer nodal metastasis?

AIM: To assess whether an additional histopathological examination of ultrasound-guided core biopsy (USCB)/fine-needle aspiration (FNA) of abnormal axillary lymph nodes (ALN) can improve the preoperative diagnosis of axillary nodal metastasis. MATERIALS AND METHODS: Women with suspected invasive breast cancer and abnormal axillary ultrasound (AUS), but negative USCB on standard histopathological assessment were included. From the core biopsies six additional levels were sectioned for haematoxylin and eosin examination, and two levels were sectioned for immunohistochemistry with AE1/3. The presence of metastatic disease was noted. RESULTS: The USCB of 102 patients were submitted for additional histopathological examination, of whom 58 had screen-detected lesions and 44 had symptomatic lesions. Eighty underwent axillary surgery for invasive carcinoma (n = 74) or for ductal carcinoma in situ (DCIS) requiring mastectomy (n = 6). Twelve patients were found to have nodal disease with a mean of two nodes involved. The additional histopathological assessment of the nodal USCBs revealed tumour not seen at the standard examination in only three cases, which consisted of isolated tumour cells (n = 2) and micrometastasis (n = 1). All three patients underwent subsequent axillary node clearance; however, no upgrade of axillary disease was found at final histopathology. CONCLUSION: Additional histopathological examination of USCBs of radiologically abnormal ALN does not improve the preoperative diagnosis of axillary nodal metastasis in primary breast cancer and may lead to unnecessary axillary clearance.

Multiparameter DNA content analysis identifies distinct groups in primary breast cancer.

BACKGROUND: Multiparameter flow cytometry is a robust and reliable method for determining tumour DNA content applicable to formalin-fixed paraffin-embedded (FFPE) tissue. This study examined the clinical and pathological associations of DNA content in primary breast cancer using an improved multiparametric technique. METHODS: The FFPE tissue from 201 primary breast cancers was examined and the cancers categorised according to their DNA content using multiparametric flow cytometry incorporating differential labelling of stromal and tumour cell populations. Mathematical modelling software (ModFit 3.2.1) was used to calculate the DNA index (DI) and percentage S-phase fraction (SPF%) for each tumour. Independent associations with clinical and pathological parameters were sought using backward stepwise Binary Logistic Regression (BLR) and Cox's Regression (CR) analysis. RESULTS: Tumours were grouped into four categories based on the DI of the tumour cell population. Low DI tumours (DI=0.76-1.14) associated with progesterone receptor-positive status (P=0.012, BLR), intermediate DI (DI=1.18-1.79) associated with p53 mutant tumours (P=0.001, BLR), high DI (DI1.80) tumours with human epidermal growth factor receptor 2 (HER2)-positive status (P=0.004, BLR) and 'multiploid tumours' (two or more tumour DNA peaks) did not show any significant associations. Tumours with high SPF% (10%) independently associated with poor overall survival (P=0.027, CR). CONCLUSION: Multiparametric flow analysis of FFPE tissue can accurately assess tumour DNA content. Tumour sub-populations associated with biomarkers of prognosis or likely response to therapy. The alterations in DNA content present the potential for greater understanding of the mechanisms underlying clinically significant biomarker changes in primary breast cancer.

Sentinel node biopsy in breast cancer has a greater node positivity rate than axillary node sample: results from a retrospective analysis.

BACKGROUND: Sentinel node biopsy as a surgical method of axillary staging for early breast cancer has been widely accepted as an alternative to traditional four-node axillary node sampling, and is the recommended technique by the Association of Breast Surgery in the United Kingdom. In selected units axillary sampling has been compared with either radioisotope sentinel node or blue dye only techniques with comparable node positivity rates. There are no studies directly comparing combined method sentinel node biopsy (SNB) with conventional axillary (four) node sampling (ANS). METHODS: Data for all patients undergoing axillary staging by axillary node sample or sentinel node biopsy were collected, including those proceeding to axillary clearance as a second procedure, but excluding those undergoing axillary clearance as a first procedure. RESULTS: From January 2005 to January 2011, 641 axillary staging procedures were performed (SNB n=231 (36.0%), ANS n=410 (64.0%)). Baseline tumour characteristics were similar for the two groups except for a higher frequency of breast conservation in the SNB group (95.6 vs. 75.6%; p<0.0001). The proportion of cases with positive nodes was higher in the SNB group (20.8 vs. 14.4%; p=0.042). In patients who had presented with symptomatic disease, there was a significantly higher node positivity rate with SNB (30.9%) than with ANS (15.5%; p=0.002), despite similar baseline characteristics in both groups. CONCLUSION: Combined method sentinel node biopsy is more sensitive at detecting low volume axillary disease than traditional four-node sample.

Involved anterior margins after breast conserving surgery: is re-excision required?

BACKGROUND: Complete tumour excision in breast conserving surgery (BCS) is critical for successful outcome; involved circumferential resection margins are associated with increased disease recurrence. However, the importance of an involved anterior margin (IAM) is less clear. The purpose of this study was to review an aggressive approach to IAM and hence assess whether anterior margin re-excision (RE) yields clinical benefit. METHODS: A review of prospectively collected clinical and pathology data was performed for all patients who underwent BCS between 2006 and 2010 through a single cancer centre. An involved margin was defined as < 1 mm clearance of invasive or in-situ breast cancer. RESULTS: 1667 patients underwent BCS for invasive and/or in-situ disease, of whom 114 underwent RE. A total of 170 involved margins were identified: most commonly the anterior (52 margins) followed by the posterior (39 margins) and inferior (31 margins) margin. Patients with IAM were more likely to have grade 3 invasive disease (p = 0.0323) but less likely to have residual disease found at re-excision (2/49 vs. 32/101 margins, p = 0.0033); there were no differences when in-situ characteristics were compared. CONCLUSIONS: RE of IAM after BCS rarely yields further disease; multi-disciplinary teams should consider whether further therapy for an IAM is required on a patient by patient basis.

Axillary lymph node core biopsy for breast cancer metastases -- how many needle passes are enough?

AIM: To determine the diagnostic yield of each of three core passes when sampling abnormal lymph nodes in patients presenting with breast cancer. MATERIALS AND METHODS: All patients suspected of having breast cancer had axillary ultrasound as part of initial assessment. Radiologically abnormal nodes (cortical thickness >2.3mm or round shape) were biopsied with three passes of a 22 mm throw 14 G core biopsy needle and sent for histopathology in separate numbered pots. Data were collected prospectively, and analysis performed on the data of 55 consecutive patients who had positive nodes on at least one core biopsy needle pass. RESULTS: Of 55 patients with a positive node on core biopsy, tumour was noted in all three cores taken in 39 (70.9%). Lymph node metastasis was detected in 45 (81.8%) first core biopsies. With the first two cores taken, positive results were detected in 53 of 55 cases (96.4%). In both cases where tumour was only found on a third core biopsy pass, no lymph node tissue was present in the first two biopsy passes. CONCLUSION: Two well-directed 14 G core biopsy samples from an abnormal axillary node are adequate for diagnosis of breast cancer metastasis.

High frequency of complex TP53 mutations in CNS metastases from breast cancer.

BACKGROUND: Brain metastasis from breast cancer is usually associated with a poor prognosis and early death. Alteration of p53 may contribute to malignant progression by abrogation of apoptosis induced by oncogene activation and by acquisition of gain-of-function properties, which promote tumour aggression. Mutation in TP53 occurs at high frequency in carcinomas of the lung and gastro-intestinal tract, but is much less frequent, at 25%, in primary breast cancer. The frequency of TP53 alteration in the central nervous system (CNS) metastatic breast cancer is not known. METHODS: In all, 23 cases of histologically confirmed CNS metastatic breast cancer were identified and the coding sequence of TP53 determined. TP53 was also sequenced in two control series of primary breast carcinomas from independent clinical centres. RESULTS: We demonstrate a strikingly high frequency of TP53 mutation in the CNS metastatic lesions with an over-representation of complex mutations (non-sense/deletions/insertions). Complex mutations occur in metastatic lesions in both triple-negative breast cancer and hormone receptor/HER2-positive cases. Analysis of paired primary carcinomas and brain metastatic lesions revealed evidence for both clonal selection and generation of new mutations (missense and complex) in progression from a primary breast carcinoma to brain metastasis. CONCLUSION: Mutation in TP53 is the most common genetic alteration reported during metastasis to the brain in breast cancer.

The RNA helicase p68 modulates expression and function of the Δ133 isoform(s) of p53, and is inversely associated with Δ133p53 expression in breast cancer.

The RNA helicase p68 is a potent co-activator of p53-dependent transcription in response to DNA damage. Previous independent studies have indicated that p68 and the Δ133p53 isoforms, which modulate the function of full-length p53, are aberrantly expressed in breast cancers. Here we identify a striking inverse association of p68 and Δ133p53 expression in primary breast cancers. Consistent with these findings, small interfering RNA depletion of p68 in cell lines results in a p53-dependant increase of Δ133p53 in response to DNA damage, suggesting that increased Δ133p53 expression could result from downregulation of p68 and provide a potential mechanistic explanation for our observations in breast cancer. Δ133p53α, which has been shown to negatively regulate the function of full-length p53, reciprocally inhibits the ability of p68 to stimulate p53-dependent transcription from the p21 promoter, suggesting that Δ133p53α may be competing with p68 to regulate p53 function. This hypothesis is underscored by our observations that p68 interacts with the C-terminal domain of p53, co-immunoprecipitates 133p53α from cell extracts and interacts only with p53 molecules that are able to form tetramers. These data suggest that p68, p53 and 133p53α may form part of a complex feedback mechanism to regulate the expression of Δ133p53, with consequent modification of p53-mediated transcription, and may modulate the function of p53 in breast and other cancers that harbour wild-type p53.

Increased mortality in HER2 positive, oestrogen receptor positive invasive breast cancer: a population-based study.

BACKGROUND: This study assessed the impact of human epidermal growth factor receptor 2 (HER2) status on the outcomes in an unselected population of breast cancer patients who did not receive HER2-targeted therapy. METHODS: HER2 status by immunohistochemistry and fluorescence in situ hybridisation was compared with clinicopathological data, overall survival (OS) and disease-free survival (DFS) for all patients presenting with breast cancer over 3 years. RESULTS: In 865 patients (median follow up 6.02 years), HER2 positivity was identified in 13.3% of all cancers and was associated with higher tumour grade (P<10(-8)), lymphovascular invasion (P<0.001) and axillary nodal metastasis (P=0.003). There was a negative association with oestrogen-receptor (ER) and progesterone-receptor expression (P<10(-8)), but the majority (57%) of HER2+tumours were ER+HER2 positivity was associated with poorer OS (P=0.0046) and DFS (P=0.0001) confined to the lymph node-positive (LN+) and ER+ subgroups. CONCLUSION: HER2-positive cancers were less common in this population-based cohort than most selected series. The association of HER2 positivity with poor prognosis was confined to the ER+ and LN+ subgroups. The survival deficit for the 7.5% of patients with ER+/HER2+ cancer compared with ER+/HER2- patients points to a significant subgroup of women who may not (currently) be considered for HER2-directed therapy.

p53 mutation, deprivation and poor prognosis in primary breast cancer.

BACKGROUND: The deprivation gap for breast cancer survival remains unexplained by stage at presentation, treatment, or co-morbidities. We hypothesised that p53 mutation might contribute to the impaired outcome observed in patients from deprived communities. METHODS: p53 mutation status was determined using the Roche Amplichip research test in 246 women with primary breast cancer attending a single cancer centre and related to deprivation, pathology, overall, and disease-free survival. RESULTS: p53 mutation, identified in 64/246 (26%) of cancers, was most common in 10 out of 17 (58.8%) of the lowest (10th) deprivation decile. Those patients with p53 mutation in the 10th decile had a significantly worse disease-free survival of only 20% at 5 years (Kaplan-Meier logrank chi(2)=6.050, P=0.014) and worse overall survival of 24% at 5 years (Kaplan-Meier logrank chi(2)=6.791, P=0.009) than women of deciles 1-9 with p53 mutation (c.f. 56% and 72%, respectively) or patients in the 10th decile with wild-type p53 (no disease relapse or deaths). CONCLUSION: p53 mutation in breast cancer is associated with socio-economic deprivation and may provide a molecular basis, with therapeutic implications, for the poorer outcome in women from deprived communities.

Meta-analysis of sentinel node biopsy in ductal carcinoma in situ of the breast.

BACKGROUND: The need for sentinel lymph node (SLN) biopsy in patients with a preoperative diagnosis of ductal carcinoma in situ (DCIS) is debated. Advocates recommend such biopsy based on a high incidence of SLN involvement in some series. Opponents discourage SLN biopsy based on a perceived low incidence of nodal involvement in this setting. These contradictory arguments are generally based on small studies. The present study is a meta-analysis of the reported data on the incidence of SLN metastasis in patients with DCIS. METHODS: A search of electronic databases identified studies reporting the frequency of SLN metastases in DCIS. The random-effects method was used to combine data. RESULTS: Twenty-two published series were included in the meta-analysis. The estimate for the incidence of SLN metastases in patients with a preoperative diagnosis of DCIS was 7.4 (95 per cent confidence interval (c.i.) 6.2 to 8.9) per cent compared with 3.7 (95 per cent c.i. 2.8 to 4.8) per cent in patients with a definitive (postoperative) diagnosis of DCIS alone. This was a significant difference with an odds ratio of 2.11 (95 per cent c.i. 1.15 to 2.93). CONCLUSION: Patients with a preoperative diagnosis of DCIS should be considered for SLN biopsy.

Differential p53 protein expression in breast cancer fine needle aspirates: the potential for in vivo monitoring.

Fine needle aspiration (FNA) biopsy is the least invasive method of sampling breast cancer in vivo and provides material for breast cancer diagnosis. FNA has also been used to examine cellular markers to predict and monitor the effects of therapy. The aim of this study was to assess the accuracy of using FNA material compared with resected cancer for Western blotting studies of the p53 pathway, a key to tumour response to radiotherapy and chemotherapy. Paired samples of breast cancer FNAs collected pre-operatively and post-operatively were compared with tissue samples obtained at the time of surgical resection. Western blots were probed for p53 using the antibodies DO12 and DO1, and for levels of downstream proteins p21/WAF1 and p27. The protein extracted by FNA was sufficient for up to 5 Western blot studies. p53 expression and phosphorylation did not differ significantly pre- and post-operatively, indicating that intra-operative manipulation does not affect p53 expression or downstream activation in breast cancer. However, expression of p53, p21 and p27 varied between individual patients suggesting a range of p53 pathway activation in breast cancer. Immunohistochemistry confirmed that the cancer cells accounted for the protein expression detected on Western blots. FNA yields adequate protein for Western blotting studies and could be used as a method to monitor p53 activity in vivo before and during anti-cancer treatment possibly providing early evidence of tumour response to therapy.