There is a quest for novel noninvasive diagnostic markers for the detection of breast cancer. The goal of this study is to identify circulating microRNA (miRNA) signatures using a cohort of Asian ...Chinese patients with breast cancer, and to compare miRNA profiles between tumor and serum samples.
miRNA from paired breast cancer tumors, normal tissue, and serum samples derived from 32 patients were comprehensively profiled using microarrays or locked nucleic acid real-time PCR panels. Serum samples from healthy individuals (n = 22) were also used as normal controls. Significant serum miRNAs, identified by logistic regression, were validated in an independent set of serum samples from patients (n = 132) and healthy controls (n = 101).
The 20 most significant miRNAs differentially expressed in breast cancer tumors included miRNA (miR)-21, miR-10b, and miR-145, previously shown to be dysregulated in breast cancer. Only 7 miRNAs were overexpressed in both tumors and serum, suggesting that miRNAs may be released into the serum selectively. Interestingly, 16 of the 20 most significant miRNAs differentially expressed in serum samples were novel. MiR-1, miR-92a, miR-133a, and miR-133b were identified as the most important diagnostic markers, and were successfully validated; receiver operating characteristic curves derived from combinations of these miRNAs exhibited areas under the curves of 0.90 to 0.91.
The clinical use of miRNA signatures as a noninvasive diagnostic strategy is promising, but should be further validated for different subtypes of breast cancers.
Genome-wide association studies (GWAS) have proven highly successful in identifying single nucleotide polymorphisms (SNPs) associated with breast cancer (BC) risk. The majority of these studies are ...on European populations, with limited SNP association data in other populations. We genotyped 51 GWAS-identified SNPs in two independent cohorts of Singaporean Chinese. Cohort 1 comprised 1294 BC cases and 885 controls and was used to determine odds ratios (ORs); Cohort 2 had 301 BC cases and 243 controls for deriving polygenic risk scores (PRS). After age-adjustment, 11 SNPs were found to be significantly associated with BC risk. Five SNPs were present in <1% of Cohort 1 and were excluded from further PRS analysis. To assess the cumulative effect of the remaining 46 SNPs on BC risk, we generated three PRS models: Model-1 included 46 SNPs; Model-2 included 11 statistically significant SNPs; and Model-3 included the SNPs in Model-2 but excluded SNPs that were in strong linkage disequilibrium with the others. Across Models-1, -2 and -3, women in the highest PRS quartile had the greatest ORs of 1.894 (95% CI = 1.157-3.100), 2.013 (95% CI = 1.227-3.302) and 1.751 (95% CI = 1.073-2.856) respectively, suggesting a direct correlation between PRS and BC risk. Given the potential of PRS in BC risk stratification, our findings suggest the need to tailor the selection of SNPs to be included in an ethnic-specific PRS model.
Aims
Ductal carcinoma in situ (DCIS) of the breast is a heterogeneous disease that has risen to prominence and more recently controversy, with the advent of screening mammography. Debate concerning ...the true biological potential of low nuclear grade DCIS continues to challenge therapeutic considerations. In this study, we carried out a comprehensive literature review of the behaviour, outcomes and current management trials of low‐grade DCIS, as well as a retrospective study of a large single institutional series of low‐grade DCIS diagnosed at our hospital.
Methods and results
The study cohort comprised 195 cases of low‐grade DCIS diagnosed at the Singapore General Hospital from 1994 to 2010. Clinicopathological parameters and follow‐up data were retrieved and compared between screen‐detected and symptomatic low‐grade DCIS. Immunohistochemistry was performed for ER, PR and HER2. Among 195 cases, 123 (63.1%) were screen‐detected, while 72 (36.9%) were symptomatic. Screen‐detected cases had frequent calcifications (P < 0.001) and were smaller (P = 0.018) than symptomatic cases. All cases were ER‐positive and rate of PR expression was high. No HER2 overexpression was observed. Mean and median follow‐up periods were 107.8 and 109.6 months, respectively. Six patients recurred ipsilaterally, and one patient developed direct distant metastasis. One breast cancer‐related death was recorded. Positive surgical margins (P = 0.023) were significantly associated with a higher risk of ipsilateral recurrences, as well as poorer disease‐free survivals (P = 0.010).
Conclusion
Our data indicate that low‐grade DCIS may be followed by invasive recurrences and even metastatic disease, requiring more study before being regarded as innocuous and indolent.
The National Comprehensive Cancer Network (NCCN) has proposed guidelines for the genetic testing of the BRCA1 and BRCA2 genes, based on studies in western populations. This current study assessed ...potential predictive factors for BRCA mutation probability, in an Asian population.
A total of 359 breast cancer patients, who presented with either a family history (FH) of breast and/or ovarian cancer or early onset breast cancer, were accrued at the National Cancer Center Singapore (NCCS). The relationships between clinico-pathological features and mutational status were calculated using the Chi-squared test and binary logistic regression analysis.
Of 359 patients, 45 (12.5%) had deleterious or damaging missense mutations in BRCA1 and/or BRCA2. BRCA1 mutations were more likely to be found in ER-negative than ER-positive breast cancer patients (P=0.01). Moreover, ER-negative patients with BRCA mutations were diagnosed at an earlier age (40 vs. 48 years, P=0.008). Similarly, triple-negative breast cancer (TNBC) patients were more likely to have BRCA1 mutations (P=0.001) and that these patients were diagnosed at a relatively younger age than non-TNBC patients (38 vs. 46 years, P=0.028). Our analysis has confirmed that ER-negative status, TNBC status and a FH of hereditary breast and ovarian cancer (HBOC) are strong factors predicting the likelihood of having BRCA mutations.
Our study provides evidence that TNBC or ER-negative patients may benefit from BRCA genetic testing, particularly younger patients (<40 years) or those with a strong FH of HBOC, in Asian patients.
Background
Recent experience with the ipsilateral TRAM flap has shown that it has the advantage of a longer functional pedicle length, which allows tension-free inset of well-vascularized tissue into ...the breast pocket. This leads to better positioning and shaping of the reconstructed breast with minimal disruption of the inframammary fold. The purpose of this article was to provide an illustrated approach to the ipsilateral TRAM flap and to clarify the technique when applied in the context of immediate breast reconstruction following cancer extirpation.
Methods
A prospective evaluation of 89 patients who underwent immediate breast reconstruction following skin-sparing mastectomy for breast cancer was performed. All patients underwent ipsilateral TRAM reconstruction. The innate insetting advantage of the ipsilateral TRAM flap is illustrated in the article. The key steps of the technique were as follows: (1) The ipsilateral corner of the flap was used as the axillary tail, leaving the more bulky part to form the main body of the breast; (2) To avoid undesirable twists, a right TRAM was rotated clockwise so that its apex points superiorly; (3) This flap was subsequently tunneled into the breast pocket while preserving the inframammary fold. The opposite maneuvers were done for the left side; (4) If the flap was congested, venous augmentation was performed where the tributary of the axillary vein or the thoracodorsal vein was anastomosed with the inferior epigastric vein from the flap with an interposed vein graft (17% of cases).
Results
All flaps survived and flap-related complications included partial necrosis of tissue across the midline (2.2%), palpable fat necrosis (22%), and hematoma requiring drainage (2.2%). All flaps were raised concurrent with the resection, and the combined operative time ranged from 3.5 to 6 h, with a mean hospital stay of 7 days.
Conclusion
The ipsilateral TRAM flap was a reliable flap with low complication rates and short surgery time. It was our preferred choice for pedicled breast reconstruction in all cases, except for the ptotic breast or if abdominal scarring excludes its use.
The current need for high-throughput genotyping platforms for targeted validation of disease-associated single nucleotide polymorphisms (SNPs) motivated us to evaluate a novel nanofluidics platform ...for genotyping DNA extracted from peripheral blood and buccal wash samples. SNP genotyping was performed using a Fluidigm 48.48 Dynamic Array biochip on the BioMark polymerase chain reaction platform and results were compared against standard TaqMan assays and DNA sequencing. Pilot runs using these dynamic arrays on 90 samples against 20 SNP assays had an average call rate of 99.7%, with 100% call rates for 16 of the assays. Manual TaqMan genotyping of these samples against three SNPs demonstrated 100% correlation between the two platforms. To understand the influence of DNA template variability, three sources of blood samples (CH-1, n = 20; CH-2, n = 47; KK, n = 47) and buccal washes ( n = 37) were genotyped for 24 SNPs. Although both CH-1 and CH-2 batches showed good base calling (≥98.8%), the KK batch and buccal wash samples exhibited lower call rates (82.1% and 94.0%). Importantly, repurification of the KK and buccal wash samples resulted in significant improvements in their call rates (to ≥97.9%). Scale-up for genotyping 1698 cases and controls for 24 SNPs had overall call rates of 97.6% for KK and 99.2% for CH samples. The Dynamic Array approach demonstrated accuracy similar to that of TaqMan genotyping, while offering significant savings in DNA, effort, time, and costs.
Current histopathologic systems for classifying breast tumors require evaluation of multiple variables and are often associated with significant interobserver variability. Recent studies suggest that ...gene expression profiles may represent a promising alternative for clinical cancer classification. Here, we investigated the use of a customized microarray as a potential tool for clinical practice.
We fabricated custom 188-gene microarrays containing expression signatures for three breast cancer molecular subtypes luminal/estrogen receptor (ER) positive, human epidermal growth factor receptor 2 (HER2), and "basaloid", the Nottingham prognostic index (NPI-ES), and low histologic grade (TuM1). The reliability of these multiple-signature arrays (MSA) was tested in a prospective cohort of 165 patients with primary breast cancer.
The MSA-ER signature exhibited a high concordance of 90% with ER immunohistochemistry reported on diagnosis (P < 0.001). This remained unchanged at 89% (P < 0.001) when the immunohistochemistry was repeated using current laboratory standards. Expression of the HER2 signature showed a good correlation of 76% with HER2 fluorescence in situ hybridization (FISH; ratio > or =2.2; P < 0.001), which further improved to 89% when the ratio cutoff was raised to > or =5. A proportion of low-level FISH-amplified samples (ratio, 2.2-5) behaved comparably to FISH-negative samples by HER2 signature expression, HER2 quantitative reverse transcription-PCR, and HER2 immunohistochemistry. Luminal/ER+ tumors with high NPI-ES expression were associated with high NPI scores (P = 0.001), and luminal/ER+ TuM1-expressing tumors were significantly correlated with low histologic grade (P = 0.002) and improved survival outcome in an interim analysis (hazard ratio, 0.2; P = 0.019).
The consistency of the MSA platform in an independent patient population suggests that custom microarrays could potentially function as an adjunct to standard immunohistochemistry and FISH in clinical practice.
In a clinical setting, next-generation sequencing (NGS) approaches for the enrichment and resequencing of DNA targets may have limitations in throughput, cost, or accuracy. We evaluated an NGS ...workflow for targeted DNA sequencing for mutation detection. Targeted sequence data of the BRCA1 and BRCA2 genes, generated using a PCR-based, multiplexed NGS approach using the SOLiD 4 ( n = 24) and Ion Torrent PGM ( n = 20) next-generation sequencers, were evaluated against sequence data obtained by Sanger sequencing. The overall sensitivity for SOLiD and PGM were 97.8% (95% CI = 94.7 to 100.0) and 98.9% (95% CI = 96.8 to 100.0) respectively. The specificity for the SOLiD platform was high, at 100.0% (95% CI = 99.3 to 100.0). PGM correctly identified all 3 indels, but 68 false-positive indels were also called. Equimolar normalization of amplicons was not necessary for successful NGS. Both platforms are highly amenable to scale-up, potentially reducing the reagent cost for BRCA testing to <US$200. Only 325 ng of DNA per patient is required, with similar coverage and accuracy obtained using DNA derived from peripheral blood or buccal wash samples. The strategy described is accurate and easy to incorporate into conventional workflow, and shows potential for mutation screening of clinically important gene targets in genetic disorders.
To define a predictive model for clinical behaviour of breast phyllodes tumours (PT) using histological parameters and surgical margin status.
Cases of breast PT diagnosed in the Department of ...Pathology Singapore General Hospital between January 1992 and December 2010 were stratified into benign, borderline and malignant grades based on a combination of histological parameters (stromal atypia, hypercellularity, mitoses, overgrowth and nature of tumour borders). Surgical margin status was assessed. Clinical follow-up and biostatistical modelling were accomplished.
Of 605 PT, 440 (72.7%) were benign, 111 (18.4%) borderline and 54 (8.9%) malignant. Recurrences, which were predominantly local, were documented in 80 (13.2%) women. Deaths from PT occurred in 12 (2%) women. Multivariate analysis revealed stromal atypia, overgrowth and surgical margins to be independently predictive of clinical behaviour, with mitoses achieving near significance. Stromal hypercellularity and tumour borders were not independently useful. A nomogram developed based on atypia, mitoses, overgrowth and surgical margins (AMOS criteria) could predict recurrence-free survival at 1, 3, 5 and 10 years. This nomogram was superior to a total histological score derived from adding values assigned to each of five histological parameters.
A predictive nomogram based on three histological criteria and surgical margin status can be used to calculate recurrence-free survival of an individual woman diagnosed with PT. This can be applied for patient counselling and clinical management.
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