Resting-state functional magnetic resonance imaging (rs-fMRI) has been shown to elucidate reliable patterns of brain networks in both children and adults. Studies in adults have shown that rs-fMRI acquisition times of ∼5 to 6 min provide adequate sampling to produce stable spatial maps of a number of different brain networks. However, it is unclear whether the acquisition time directly translates to studies of children. While there are many similarities between the brains of children and adults, many differences are also evident. Children have increased metabolism, differences in brain morphology and connectivity strengths, greater brain plasticity, and increased brain noise. Furthermore, there are differences in physiologic parameters, such as heart and respiratory rates, and compliance of the blood vessels. These developmental differences could translate into different acquisition times for rs-fMRI studies in pediatric populations. Longer scan times, however, increase the subject burden and the risk for greater movement, especially in children. Thus, the goal of this study was to assess the optimum acquisition time of rs-fMRI to extract stable brain networks in school-age children. We utilized fuzzy set theory in 84 six-to-eight year-old children and found that eight networks, including the default mode, salience, frontal, left frontoparietal, right frontoparietal, sensorimotor, auditory, and visual networks, all stabilized after ∼5½ min. The sensorimotor network showed the least stability, whereas the salience and auditory networks showed the greatest stability. A secondary analysis using dual regression confirmed these results. In conclusion, in young children with little head motion, rs-fMRI acquisition times of ∼5½ min can extract the full complement of brain networks.
acquisition time; children; independent component analyses; network stability; resting-state fMRI
The efficacy of chemotherapy depends on the level of risk of the individual patient. Because of this, careful estimation of the risk level is mandatory. In addition to well-established clinicopathological factors, validated gene expression signatures might be useful in selected patients if all other criteria are inconclusive for therapeutic decision-making. If indicated, chemotherapy can be used either after surgery (adjuvant) or before surgery (neoadjuvant). Both approaches lead to comparable long-term survival. The neoadjuvant setting offers the additional opportunity for elaborate translational studies to develop and validate predictive biomarkers and to discover mechanisms of resistance to therapy. If possible, chemotherapy regimens should include both anthracyclines and taxanes. Docetaxel should be used every 3 weeks; better tolerability with equivalent efficacy favors the concurrent over the sequential approach. Paclitaxel, on the other hand, should be administered sequentially, either weekly or every 2 weeks. Especially, intense dose-dense sequential chemotherapy with granulocyte colony-stimulating factor support is very effective in high-risk breast cancer patients. In order to decrease toxicities, anthracycline-free regimens or a shortening of the duration of adjuvant chemotherapy are potential options that should be further explored.
Breast cancer; Prognosis; Chemotherapy
Humanized tumor mice (HTM) were generated by the co-transplantation of human hematopoietic stem cells and human breast cancer cells overexpressing HER2 into neonatal NOD-scid IL2Rγnull (NSG) mice. These mice are characterized by the development of a human immune system in combination with human breast cancer growth. Due to concurrent transplantation into newborn mice, transfer of MHC-mismatched tumor cells resulted in solid coexistence and immune cell activation (CD4+ T cells, natural killer cells, and myeloid cells), but without evidence for rejection. Histological staining of the spleen of HTM revealed co-localization of human antigen-presenting cells together with human T and B cells allowing MHC-dependent interaction, and thereby the generation of T cell-dependent antibody production. Here, we investigated the capability of these mice to generate human tumor-specific antibodies and correlated immunoglobulin titers with tumor outgrowth. We found detectable IgM and also IgG amounts in the serum of HTM, which apparently controlled tumor development when IgG serum concentrations were above 10 µg/ml. Western blot analyses revealed that the tumor-specific antibodies generated in HTM did not recognize HER2/neu antigens, but different, possibly relevant antigens for breast cancer therapy. In conclusion, HTM offer a novel approach to generate complete human monoclonal antibodies that do not require further genetic manipulation (e. g., humanization) for a potential application in humans. In addition, efficacy and safety of the generated antibodies can be tested in the same mouse model under human-like conditions. This might be of particular interest for cancer subtypes with no currently available antibody therapy.
humanized tumor mice (HTM); tumor-specific antibodies; breast cancer; IgG; IgM
Working memory (WkM) is a fundamental cognitive process that serves as a building block for higher order cognitive functions. While studies have shown that children and adolescents utilize similar brain regions during verbal WkM, there have been few studies that evaluate the developmental differences in brain connectivity. Our goal was to study the development of brain connectivity related to verbal WkM in typically developing children and adolescents.
Thirty-five healthy children and adolescents, divided into three groups: 9–12 (children), 13–16 (young adolescents) and 17–19 (older adolescents) years, were included in this fMRI study. The verbal WkM task involved a modified Sternberg item recognition paradigm using three different loads. Brain connectivity analysis was performed using independent component analyses and regressing the components with the design matrix to determine task-related networks.
Connectivity analyses resulted in four components associated solely with encoding, four solely with recognition and two with both. Two networks demonstrated age-related differences with respect to load, 1) the left motor area and right cerebellum, and 2) the left prefrontal cortex, left parietal lobe, and right cerebellum. Post hoc analyses revealed that the first network showed significant effects of age between children and the two older groups. There was increasing connectivity with increasing load for adolescents. The second network demonstrated age-related differences between children and older adolescents. Children have higher task-related connectivity at lower loads, but they tend to equalize with the adolescents with higher loads. Finally, a non-load related network involving the orbital frontal and anterior cingulate cortices showed less connectivity in children.
adolescents; brain connectivity; children; development; fMRI; verbal working memory
It is well known that smoking during pregnancy can affect offspring health. Prenatal tobacco exposure has been associated with negative behavioral and cognitive outcomes in childhood, adolescence, and young adulthood. These associations between prenatal tobacco exposure and psychopathology in offspring could possibly be explained by the influence of prenatal tobacco exposure on brain development. In this prospective study, we investigated the association between prenatal tobacco exposure, behavioral and emotional functioning and brain morphology in young children. On the basis of age and gender, we matched 113 children prenatally exposed to tobacco with 113 unexposed controls. These children were part of a population-based study in the Netherlands, the Generation R Study, and were followed from pregnancy onward. Behavioral and emotional functioning was assessed at age 6 with the Child Behavior Checklist. We assessed brain morphology using magnetic resonance imaging techniques in children aged 6–8 years. Children exposed to tobacco throughout pregnancy have smaller total brain volumes and smaller cortical gray matter volumes. Continued prenatal tobacco exposure was associated with cortical thinning, primarily in the superior frontal, superior parietal, and precentral cortices. These children also demonstrated increased scores of affective problems. In addition, thickness of the precentral and superior frontal cortices was associated with affective problems. Importantly, brain development in offspring of mothers who quit smoking during pregnancy resembled that of nonexposed controls (no smaller brain volumes and no thinning of the cortex). Our findings suggest an association between continued prenatal tobacco exposure and brain structure and function in school-aged children.
addiction & substance abuse; brain morphology; development; developmental disorders; epidemiology; neuroanatomy; pediatric brain; prenatal tobacco exposure; prenatal tobacco exposure; pediatric brain development; brain morphology
Cyclooxygenases (COX) play a key role in prostaglandin metabolism and are important for tumor development and progression. The aim of this study was to analyze the prognostic impact of COX-2 expression in a cohort of lymph node-negative breast cancer patients not treated in the adjuvant setting.
COX-2 expression was determined by immunohistochemistry (IHC) in tumor tissue of 193 node-negative breast cancer patients. Additionally, mRNA expression was determined in corresponding tumor samples using microarray based gene-expression data. Univariate and multivariate Cox regression analyses adjusted for age at diagnosis, tumor size, histological grade, human epithelial growth factor receptor 2 (HER2), estrogen receptor (ER) and progesterone receptor (PR) were performed to evaluate the association of both COX-2 protein and mRNA expression with survival. Survival rates were determined by the Kaplan-Meier method. Correlations between COX-2 expression and established prognostic factors were analyzed using the Chi-square test. A potential correlation between COX-2 protein expression and COX-2 mRNA expression was assessed utilizing the Kruscal-Wallis-H-test.
COX-2 protein expression was positive in 24.9% of the breast cancer samples. Univariate analysis showed that COX-2 protein expression was associated with shorter disease-free survival (DFS) (P = 0.0001), metastasis-free survival (MFS) (P = 0.002) as well as breast cancer specific overall survival (OS) (P = 0.043). In multivariate analysis COX-2 expression retained its significance independent of established prognostic factors for shorter DFS (P < 0.001, HR = 2.767, 95% CI = 1.563-4.901) and for inferior MFS (P = 0.002, HR = 2.7, 95% CI = 1.469-5.263) but not for OS (P = 0.096, HR = 1.929, 95% CI = 0.889-4.187). In contrast, COX-2 mRNA expression was not related to survival and failed to show a correlation with protein expression (P = 0.410).
The present findings support the hypothesis that COX-2 protein but not mRNA expression is associated with an unfavorable outcome in node-negative breast cancer.
Electronic supplementary material
The online version of this article (doi:10.1186/1471-2407-14-952) contains supplementary material, which is available to authorized users.
COX-2; Breast cancer; Node-negative; Prognosis
Protein levels of urokinase plasminogen activator (uPA) and its inhibitor (PAI-1) determined by enzyme-linked immunosorbent assay from fresh-frozen tumor tissue have been evaluated as prognostic factors in prospectively randomized trials in breast cancer. However, the role of uPA and PAI-1 in the context of breast cancer subtypes and for mRNA expression of these factors is less clear.
We evaluated uPA and PAI-1 mRNA expression using the Affymetrix HG-U 133A array within molecular subgroups of breast cancer in cohorts of patients with systemic treatment (cohort A, n=362) and without systemic treatment (cohort B, n=200). We validated mRNA expression in a cohort of HER2-positive breast cancer patients (cohort C, n=290). Luminal, triple-negative, and HER2-positive subcohorts were defined by ESR1 and ERBB2 mRNA expression using predefined cutoffs.
In the entire cohort A, elevated PAI-1 but not uPA mRNA expression was associated with shorter disease-free survival (P=0.007 for PAI and 0.069 for uPA). Regarding different molecular subgroups, 67% (n=244) of tumors were luminal, 14% (n=49) were HER2-positive, and 19% (n=69) were triple-negative. Elevated PAI-1 mRNA expression was associated with shorter disease-free survival only in the HER2-positive subgroup (P=0.031). The same disease-free survival results were found for uPA in HER2-positive patients (P=0.011). In contrast, no association between either marker and survival was observed in the luminal or triple-negative subgroups. In the HER2-positive validation cohort C, elevated uPA and PAI-1 mRNA expression also showed strong associations with shorter disease-free survival (P=0.014 for PAI-1, P<0.001 for uPA).
In this study, the prognostic impact of uPA and PAI-1 expression was mainly observed in patients with HER2-positive tumors.
urokinase plasminogen activator; urokinase plasminogen activator inhibitor-1; HER2; breast cancer; prognosis
Similar to adults, children and adolescents with schizophrenia present with significant working memory (WkM) deficits. However, unlike adults, findings of abnormal activity in the prefrontal cortex in early-onset schizophrenia (EOS) are not consistently reported. Since WkM continues to develop through adolescence and into early adulthood, patterns of activation in adolescents may be different than those found in adults. The goal of this study was to evaluate the functional neurobiology of WkM in patients with EOS.
Participants included 22 patients with EOS (mean age 15 ±2.8 years) and 24 controls (mean age 15.0 ±3.0 years). Diagnoses were confirmed using the KIDDIE-SADS-PL. All subjects underwent a functional MRI paradigm involving a visuospatial working memory task with three separate loads.
The behavioral results demonstrated deficits in EOS patients at all three WkM loads. On functional imaging, EOS patients demonstrated increased activation in the anterior cingulate cortex (ACC), medial temporal lobe structures, the insula, and bilateral lateral temporal lobes.
Patients with EOS demonstrate increased activity in limbic structures and regions involved in processing primary and secondary sensory information. In addition, EOS patients had load dependent decreased activity in the parietal lobe. Unlike studies in adults, we did not find that EOS patients had activation differences in frontal cortical regions. One possibility is that abnormalities in PFC function are related to secondary downstream or developmental processes which are ‘unmasked’ during development. Finally, our findings support growing evidence that EOS patients have aberrations in limbic and temporal lobe regions.
fMRI; Working Memory; Early-Onset Schizophrenia; Temporal Lobe; Limbic
The importance of inflammation is increasingly noticed in cancer. The aim of this study was to analyze the prognostic influence of pre-operative serum C-reactive protein (CRP) in a cohort of 148 lymph node-negative breast cancer patients. The prognostic significance of CRP level for disease-free survival (DFS), metastasis-free survival (MFS) and overall survival (OS) was evaluated using univariate and multivariate Cox regression, also including information on age at diagnosis, tumor size, tumor grade, estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) status, proliferation index (Ki67) and molecular subtype, as well as an assessment of the presence of necrosis and inflammation in the tumor tissue. Univariate analysis showed that CRP, as a continuous variable, was significantly associated with DFS (P = 0.002, hazard ratio [HR] = 1.04, 95% confidence interval [CI] = 1.02–1.07) and OS (P = 0.036, HR = 1.03, 95% CI = 1.00–1.06), whereas a trend was observed for MFS (P = 0.111). In the multivariate analysis, CRP retained its significance for DFS (P = 0.033, HR = 1.01, 95% CI = 1.00–1.07) as well as OS (P = 0.023, HR = 1.03, 95% CI = 1.00–1.06), independent of established prognostic factors. Furthermore, large-scale gene expression analysis by Affymetrix HG-U133A arrays was performed for 72 (48.6%) patients. The correlations between serum CRP and gene expression levels in the corresponding carcinoma of the breast were assessed using Spearman's rank correlation, controlled for false-discovery rate. No significant correlation was observed between CRP level and gene expression indicative of an ongoing local inflammatory process. In summary, pre-operatively elevated CRP levels at the time of diagnosis were associated with shorter DFS and OS independent of established prognostic factors in node-negative breast cancer, supporting a possible link between inflammation and prognosis in breast cancer.
It has long been known that specific visual frequencies result in greater blood flow to the striate cortex. These peaks are thought to reflect synchrony of local neuronal firing that is reflective of local cortical networks. Since disrupted neural connectivity is a possible etiology for schizophrenia, our goal was to investigate whether localized connectivity, as measured by aberrant synchrony, is abnormal in children and adolescents with schizophrenia. Subjects included 25 children and adolescents with schizophrenia and 39 controls matched for age and gender. Subjects were scanned on a Siemens 3 Tesla Trio scanner while observing flashing checkerboard presented at either 1, 4, 8, or 12 Hz. Image processing included both a standard GLM model and a Fourier transform analysis. Patients had significantly smaller volume of activation in the occipital lobe compared to controls. There were no differences in the integral or percent signal change of the hemodynamic response function for each of the four frequencies. Occipital activation was stable during development between childhood and late adolescence. Finally, both patients and controls demonstrated an increased response between 4 and 8 Hz consistent with synchrony or entrainment in the neuronal response. Children and adolescents with schizophrenia had a significantly lower volume of activation in the occipital lobe in response to the flashing checkerboard task. However, features of intact local connectivity in patients, such as the hemodynamic response function and maximal response at 8 Hz, were normal. These results are consistent with abnormalities in regional connectivity with preserved local connectivity in early-onset schizophrenia.
Early-Onset Schizophrenia; Functional Magnetic Resonance Imaging; fMRI; Flashing Checkerboard; Hemodynamic Response Function
Strongly increased uptake of glucose is a hallmark of solid malignant tumors. This phenotype can be triggered by hypoxia-induced gene expression changes or can occur independently of hypoxia as a consequence of malignant transformation itself, and is often referred to as the Warburg effect. The glycolytic phenotype has been associated with malignant progression and resistance to radio- and chemotherapy.
We have chosen squamous cell carcinomas of the vulva (SCC-V) as a representative solid tumor entity to study the central players of this pathway, namely glucose transporter (GLUT)-1, carbonic anhydrase (CA) IX, hexokinase (HK)-2 and pyruvate kinase (PK)-M2, and have investigated their relationships to tumor microvessels (CD34, αSMA) and proliferation (Ki67). Expression of these proteins was analyzed in 38 SCC-Vs, 5 vulvar dysplasias and 10 non-neoplastic squamous epithelia of the vulva using multiparametric immunohistochemistry in registered serial sections (MIRSS).
Expression of GLUT-1 in invasive carcinomas was predominantly located in the outer layers of the tumor cell aggregates close to the vascularized tumor stroma, and only to a lesser extent colocalized with CA IX, which was repeatedly found at larger diffusion distances away from microvessels. CA IX expression was lower in invasive carcinomas compared to dysplasias and non-neoplastic tissue and higher in recurrent vs. primary tumors. Ki67-positive proliferating cells were partially colocalized with GLUT-1. However, HK-2 and PK-2 - proteins centrally involved in the Warburg phenotype - did not show such a correlation.
Consistent with prior studies, the pattern of GLUT-1 clearly indicated that a large part of its expression is presumably unrelated to hypoxia. However, there was also no association with HK-2 and PK-M2, suggesting that the functional background of this expression is also independent of aerobic glycolysis. CA IX may be worth consideration as a marker of biological hypoxia, as should its pathophysiological consequences in SCC-V.
Electronic supplementary material
The online version of this article (doi:10.1186/1471-2407-14-760) contains supplementary material, which is available to authorized users.
Vulvar carcinoma; Hypoxia; Glucose transporter; Carbonic anhydrase IX; Hexokinase 2; Pyruvate kinase M2; Warburg effect
The growth inhibitory effect of tamoxifen, which is used for the treatment of hormone receptor–positive breast cancer, is mediated by its metabolites, 4-hydroxytamoxifen and endoxifen. The formation of active metabolites is catalyzed by the polymorphic cytochrome P450 2D6 (CYP2D6) enzyme.
To determine whether CYP2D6 variation is associated with clinical outcomes in women receiving adjuvant tamoxifen.
Design, Setting, and Patients
Retrospective analysis of German and US cohorts of patients treated with adjuvant tamoxifen for early stage breast cancer. The 1325 patients had diagnoses between 1986 and 2005 of stage I through III breast cancer and were mainly postmenopausal (95.4%). Last follow-up was in December 2008; inclusion criteria were hormone receptor positivity, no metastatic disease at diagnosis, adjuvant tamoxifen therapy, and no chemotherapy. DNA from tumor tissue or blood was genotyped for CYP2D6 variants associated with reduced (*10, *41) or absent (*3, *4, *5) enzyme activity. Women were classified as having an extensive (n = 609), heterozygous extensive/intermediate (n = 637), or poor (n = 79) CYP2D6 metabolism.
Main Outcome Measures
Time to recurrence, event-free survival, disease-free survival, and overall survival.
Median follow-up was 6.3 years. At 9 years of follow-up, the recurrence rates were 14.9% for extensive metabolizers, 20.9% for heterozygous extensive/intermediate metabolizers, and29.0%for poor metabolizers, and all-cause mortality rates were 16.7%, 18.0%, and 22.8%, respectively. Compared with extensive metabolizers, there was a significantly increased risk of recurrence for heterozygous extensive/intermediate metabolizers (time to recurrence adjusted hazard ratio [HR], 1.40; 95% confidence interval [CI], 1.04–1.90) and for poor metabolizers (time to recurrence HR, 1.90; 95% CI, 1.10–3.28). Compared with extensive metabolizers, those with decreased CYP2D6 activity (heterozygous extensive/intermediate and poor metabolism) had worse event-free survival (HR, 1.33; 95% CI, 1.06–1.68) and disease-free survival (HR, 1.29; 95% CI, 1.03–1.61), but there was no significant difference in overall survival (HR, 1.15; 95% CI, 0.88–1.51).
Among women with breast cancer treated with tamoxifen, there was an association between CYP2D6 variation and clinical outcomes, such that the presence of 2 functional CYP2D6 alleles was associated with better clinical outcomes and the presence of nonfunctional or reduced-function alleles with worse outcomes.
The placenta-specific 1 (PLAC1) gene encodes a membrane-associated protein which is selectively expressed in the placental syncytiotrophoblast and in murine fetal tissues during embryonic development. In contrast to its transcriptional repression in all other adult normal tissues, PLAC1 is frequently activated and highly expressed in a variety of human cancers, in particular breast cancer, where it associates with estrogen receptor α (ERα) positivity. In a previous study, we showed that ERα-signaling in breast cancer cells transactivates PLAC1 expression in a non-classical pathway. As the members of the p160/nuclear receptor co-activator (NCOA) family, NCOA1, NCOA2 and NCOA3 are known to be overexpressed in breast cancer and essentially involved in estrogen-mediated cancer cell proliferation we asked if these proteins are involved in the ERα-mediated transactivation of PLAC1 in breast cancer cells.
Applying quantitative real-time RT-PCR (qRT-PCR), Western Blot analysis and chromatin immunoprecipitation, we analyzed the involvement of NCOA1, NCOA2, NCOA3 in the ERα-mediated transactivation of PLAC1 in the breast cancer cell lines MCF-7 and SK-BR-3. RNAi-mediated silencing of NCOA3, qRT-PCR, Western blot analysis and ERα activation assays were used to examine the role of NCOA3 in the ERα-mediated regulation of PLAC1 in further detail. Transcript expression of NCOA3 and PLAC1 in 48 human breast cancer samples was examined by qRT-PCR and statistical analysis was performed using Student’s t-test.
We detected selective recruitment of NCOA3 but not NCOA1 or NCOA2 to the PLAC1 promoter only in ERα-positive MCF-7 cells but not in ERα-negative SK-BR-3 breast cancer cells. In addition, we demonstrate that silencing of NCOA3 results in a remarkable decrease of PLAC1 expression levels in MCF-7 cells which cannot be restored by treatment with estradiol (E2). Moreover, significant higher transcript levels of PLAC1 were found only in ERα-positive human breast cancer samples which also show a NCOA3 overexpression.
In this study, we identified NCOA3 as a selective co-activator of ERα-mediated transactivation of PLAC1 in MCF-7 breast cancer cells. Our data introduce PLAC1 as novel target gene of NCOA3 in breast cancer, supporting the important role of both factors in breast cancer biology.
NCOA3; PLAC1; Estrogen-signaling; Breast cancer; Estrogen receptor α; Tumor antigen
We examined if a combination of proliferation markers and estrogen receptor (ER) activity could predict early versus late relapses in ER-positive breast cancer and inform the choice and length of adjuvant endocrine therapy.
Baseline affymetrix gene-expression profiles from ER-positive patients who received no systemic therapy (n = 559), adjuvant tamoxifen for 5 years (cohort-1: n = 683, cohort-2: n = 282) and from 58 patients treated with neoadjuvant letrozole for 3 months (gene-expression available at baseline, 14 and 90 days) were analyzed. A proliferation score based on the expression of mitotic kinases (MKS) and an ER-related score (ERS) adopted from Oncotype DX® were calculated. The same analysis was performed using the Genomic Grade Index as proliferation marker and the luminal gene score from the PAM50 classifier as measure of estrogen-related genes. Median values were used to define low and high marker groups and four combinations were created. Relapses were grouped into time cohorts of 0–2.5, 0–5, 5-10 years.
In the overall 10 years period, the proportional hazards assumption was violated for several biomarker groups indicating time-dependent effects. In tamoxifen-treated patients Low-MKS/Low-ERS cancers had continuously increasing risk of relapse that was higher after 5 years than Low-MKS/High-ERS cancers [0 to 10 year, HR 3.36; p = 0.013]. High-MKS/High-ERS cancers had low risk of early relapse [0–2.5 years HR 0.13; p = 0.0006], but high risk of late relapse which was higher than in the High-MKS/Low-ERS group [after 5 years HR 3.86; p = 0.007]. The High-MKS/Low-ERS subset had most of the early relapses [0 to 2.5 years, HR 6.53; p < 0.0001] especially in node negative tumors and showed minimal response to neoadjuvant letrozole. These findings were qualitatively confirmed in a smaller independent cohort of tamoxifen-treated patients. Using different biomarkers provided similar results.
Early relapses are highest in highly proliferative/low-ERS cancers, in particular in node negative tumors. Relapses occurring after 5 years of adjuvant tamoxifen are highest among the highly-proliferative/high-ERS tumors although their risk of recurrence is modest in the first 5 years on tamoxifen. These tumors could be the best candidates for extended endocrine therapy.
The validated EndoPredict assay is a novel tool to predict the risk of metastases of patients with estrogen receptor positive, HER2 negative breast cancer treated with endocrine therapy alone. It has been designed to integrate genomic and clinical information and includes clinico-pathological factors such as tumor size and nodal status. The test is feasible in a decentral setting in molecular pathology laboratories. In this project, we investigated the performance of this test in clinical practice, and performed a retrospective evaluation of its impact on treatment decisions in breast cancer. During one year, EndoPredict assays from 167 patients could be successfully performed. For retrospective evaluation of treatment decisions, a questionnaire was sent to the clinical partner. Regarding the molecular EP class, samples from 56 patients (33.5%) had a low-risk, whereas 111 patients (66.5%) showed a high-risk gene profile. After integration of the clinicopathological factors the combined clinical and molecular score (EPclin) resulted in a low-risk group of 77 patients (46.4%), while 89 (53.6%) had a high risk EPclin score. The EPclin-based estimated median 10-year-risk for metastases with endocrine therapy alone was 11% for the whole cohort. The median handling time averaged three days (range: 0 to 11 days), 59.3% of the tests could be performed in three or less than three days. Comparison of pre- and post-test therapy decisions showed a change of therapy in 37.7% of patients. 16 patients (12.3%) had a change to an additional chemotherapy while 25.4% of patients (n = 33) changed to an endocrine therapy alone. In 73 patients (56.2%) no change of therapy resulted. In 6.1% of patients (n = 8), the patients did not agree to the recommendation of the tumor board. Our results show that the EndoPredict assay could be routinely performed in decentral molecular pathology laboratories and the results markedly change treatment decisions.
Taxanes are regarded as the most effective single agents in the treatment of metastatic breast cancer (MBC). For conventional taxanes, crucial toxicities and impairments in clinical efficacy are related to solvents necessary because of the agents’ hydrophobicity. The mandatory premedication with corticosteroids causes additional side effects. Nab-paclitaxel is a solvent-free colloidal suspension of paclitaxel and human serum albumin that exploits the physiological transport properties of albumin. It is registered as monotherapy with a recommended dose of 260 mg/m2 every 3 weeks for the treatment of patients with MBC, who have failed a first-line treatment of metastatic disease and for whom a standard anthracycline treatment is not indicated. Clinical evidence is available for the registered 3-weekly administration and for alternative weekly schedules in first and further lines of therapy of patients with MBC. During an advisory board meeting, a group of 8 German breast cancer experts reviewed the clinical data of nab-paclitaxel in MBC and discussed how nab-paclitaxel could be used in clinical practice on the basis of the current data.
First-line therapy; Metastatic breast cancer; Chemotherapy; Weekly; nab-Paclitaxel; Paclitaxel; Docetaxel
Prader–Willi syndrome (PWS) is a complex neurogenetic disorder with symptoms that indicate not only hypothalamic, but also a global, central nervous system (CNS) dysfunction. However, little is known about developmental differences in brain structure in children with PWS. Thus, our aim was to investigate global brain morphology in children with PWS, including the comparison between different genetic subtypes of PWS. In addition, we performed exploratory cortical and subcortical focal analyses.
High resolution structural magnetic resonance images were acquired in 20 children with genetically confirmed PWS (11 children carrying a deletion (DEL), 9 children with maternal uniparental disomy (mUPD)), and compared with 11 age- and gender-matched typically developing siblings as controls. Brain morphology measures were obtained using the FreeSurfer software suite.
Both children with DEL and mUPD showed smaller brainstem volume, and a trend towards smaller cortical surface area and white matter volume. Children with mUPD had enlarged lateral ventricles and larger cortical cerebrospinal fluid (CSF) volume. Further, a trend towards increased cortical thickness was found in children with mUPD. Children with DEL had a smaller cerebellum, and smaller cortical and subcortical grey matter volumes. Focal analyses revealed smaller white matter volumes in left superior and bilateral inferior frontal gyri, right cingulate cortex, and bilateral precuneus areas associated with the default mode network (DMN) in children with mUPD.
Children with PWS show signs of impaired brain growth. Those with mUPD show signs of early brain atrophy. In contrast, children with DEL show signs of fundamentally arrested, although not deviant brain development and presented few signs of cortical atrophy. Our results of global brain measurements suggest divergent neurodevelopmental patterns in children with DEL and mUPD.
Prader–Willi syndrome; Neurodevelopmental disorder; Chromosome 15q11-q13; Structural MRI
There is a lack of psychotherapeutic trials of treatments of comorbid depression in cancer patients. Our study determines the efficacy of a manualized short-term psychodynamic psychotherapy and predictors of outcome by personality and quality of the therapeutic relationship.
Eligible breast cancer patients with comorbid depression are assigned to short-term psychodynamic psychotherapy (up to 20 + 5 sessions) or to treatment as usual (augmented by recommendation for counseling center and physician information). We plan to recruit a total of 180 patients (90 per arm) in two centers. Assessments are conducted pretreatment, after 6 (treatment termination) and 12 months (follow-up). The primary outcome measures are reduction of the depression score in the Hospital Anxiety and Depression Scale and remission of depression as assessed by means of the Structured Clinical Interview for DSM IV Disorders by independent, blinded assessors at treatment termination. Secondary outcomes refer to quality of life.
We investigate the efficacy of short-term psychodynamic psychotherapy in acute care and we aim to identify predictors for acceptance and success of treatment.
Breast cancer; Depression; Short-term psychodynamic psychotherapy; Personality; Helping alliance; Quality of life
Inhibitors targeting the cell cycle-regulated aurora kinase A (AURKA) are currently being developed. Here, we examine the prognostic impact of AURKA in node-negative breast cancer patients without adjuvant systemic therapy (n = 766).
AURKA was analyzed using microarray-based gene-expression data from three independent cohorts of node-negative breast cancer patients. In multivariate Cox analyses, the prognostic impact of age, histological grade, tumor size, estrogen receptor (ER), and HER2 were considered.
Patients with higher AURKA expression had a shorter metastasis-free survival (MFS) in the Mainz (HR 1.93; 95% CI 1.34 – 2.78; P < 0.001), Rotterdam (HR 1.95; 95% CI 1.45– 2.63; P<0.001) and Transbig (HR 1.52; 95% CI 1.14–2.04; P=0.005) cohorts. AURKA was also associated with MFS in the molecular subtype ER+/HER2- carcinomas (HR 2.10; 95% CI 1.70–2.59; P<0.001), but not in ER-/HER2- nor in HER2+ carcinomas. In the multivariate Cox regression adjusted to age, grade and tumor size, AURKA showed independent prognostic significance in the ER+/HER2- subtype (HR 1.73; 95% CI 1.24–2.42; P=0.001). Prognosis of patients in the highest quartile of AURKA expression was particularly poor. In addition, AURKA correlated with the proliferation metagene (R=0.880; P<0.001), showed a positive association with grade (P<0.001), tumor size (P<0.001) and HER2 (P<0.001), and was inversely associated with ER status (P<0.001).
AURKA is associated with worse prognosis in estrogen receptor positive breast carcinomas. Patients with the highest AURKA expression (>75% percentile) have a particularly bad prognosis and may profit from therapy with AURKA inhibitors.
Aurora kinase; Node-negative breast cancer; Breast cancer; Prognosis; Aurora kinase inhibitors
EndoPredict (EP) is a clinically validated multianalyte gene expression test to predict distant metastasis in ER-positive, HER2-negative breast cancer treated with endocrine therapy alone. The test is based on the combined analysis of 12 genes in formalin-fixed, paraffin-embedded (FFPE) tissue by reverse transcription-quantitative real-time PCR (RT-qPCR). Recently, it was shown that EP is feasible for reliable decentralized assessment of gene expression. The aim of this study was the analytical validation of the performance characteristics of the assay and its verification in a molecular-pathological routine laboratory.
Gene expression values to calculate the EP score were assayed by one-step RT-qPCR using RNA from FFPE tumor tissue. Limit of blank, limit of detection, linear range, and PCR efficiency were assessed for each of the 12 PCR assays using serial samples dilutions. Different breast cancer samples were used to evaluate RNA input range, precision and inter-laboratory variability.
PCR assays were linear up to Cq values between 35.1 and 37.2. Amplification efficiencies ranged from 75% to 101%. The RNA input range without considerable change of the EP score was between 0.16 and 18.5 ng/μl. Analysis of precision (variation of day, day time, instrument, operator, reagent lots) resulted in a total noise (standard deviation) of 0.16 EP score units on a scale from 0 to 15. The major part of the total noise (SD 0.14) was caused by the replicate-to-replicate noise of the PCR assays (repeatability) and was not associated with different operating conditions (reproducibility). Performance characteristics established in the manufacturer’s laboratory were verified in a routine molecular pathology laboratory. Comparison of 10 tumor samples analyzed in two different laboratories showed a Pearson coefficient of 0.995 and a mean deviation of 0.15 score units.
The EP test showed reproducible performance characteristics with good precision and negligible laboratory-to-laboratory variation. This study provides further evidence that the EP test is suitable for decentralized testing in specialized molecular pathological laboratories instead of a reference laboratory. This is a unique feature and a technical advance in comparison with existing RNA-based prognostic multigene expression tests.
Breast cancer; Prognostic multigene expression test; Analytical validation; PCR; Pathology
Infiltration of plasma cells is associated with better prognosis in breast, lung and colon cancer. Immunoglobulin κ chain (IGKC) is now available as a single, robust immune marker predicting metastasis-free survival and response to chemotherapy. This will facilitate a deeper understanding of the role of the humoral immune system in cancer development.
breast cancer prognosis; colon cancer; humoral immune system; immunoglobulin kappa C; lung cancer; prediction of chemotherapy response; prognosis
Biomarkers of the immune system are currently not used as prognostic factors in breast cancer. We analyzed the association of the B cell/plasma cell marker immunoglobulin kappa C (IGKC) and survival of untreated node-negative breast cancer patients.
Material and Methods
IGKC expression was evaluated by immunostaining in a cohort of 335 node-negative breast cancer patients with a median follow-up of 152 months. The prognostic significance of IGKC for disease-free survival (DFS) and breast cancer-specific overall survival (OS) was evaluated with Kaplan-Meier survival analysis as well as univariate and multivariate Cox analysis adjusted for age at diagnosis, pT stage, histological grade, estrogen receptor (ER) status, progesterone receptor (PR) status, Ki-67 and human epidermal growth factor receptor 2 (HER-2) status.
160 patients (47.7%) showed strong expression of IGKC. Univariate analysis showed that IGKC was significantly associated with DFS (P = 0.017, hazard ratio [HR] = 0.570, 95% confidence interval [CI] = 0.360–0.903) and OS (P = 0.011, HR = 0.438, 95% CI = 0.233–0.822) in the entire cohort. The significance of IGKC was especially strong in ER negative and in luminal B carcinomas. In multivariate analysis IGKC retained its significance independent of established clinical factors for DFS (P = 0.004, HR = 0.504, 95% CI = 0.315–0.804) as well as for OS (P = 0.002, HR = 0.371, 95% CI = 0.196–0.705).
Expression of IGKC has an independent protective impact on DFS and OS in node-negative breast cancer.
We recently reported that nuclear grading in prostate cancer is subject to a strong confirmation bias induced by the tumor architecture. We now wondered whether a similar bias governs nuclear grading in breast carcinoma. An unannounced test was performed at a pathology conference. Pathologists were asked to grade nuclei in a PowerPoint presentation. Circular high power fields of 27 invasive ductal carcinomas were shown, superimposed over low power background images of either tubule-rich or tubule-poor carcinomas. We found (a) that diagnostic reproducibility of nuclear grades was poor to moderate (weighed kappa values between 0.07 and 0.54, 27 cases, 44 graders), but (b) that nuclear grades were not affected by the tumor architecture. We speculate that the categorized grading in breast cancer, separating tubule formation, nuclear pleomorphism, and mitotic figure counts in a combined three tier score, prevents the bias that architecture exerts on nuclear grades in less well-controlled situations.
Confirmation bias; Cancer grading; Nuclear pleomorphism; Architecture; Cognitive psychology
Afatinib (BIBW 2992) is an ErbB-family blocker that irreversibly inhibits signaling from all relevant ErbB-family dimers. Afatinib has demonstrated preclinical activity in human epidermal growth factor receptor HER2 (ErbB2)-positive and triple-negative xenograft models of breast cancer, and clinical activity in phase I studies. This was a multicenter phase II study enrolling patients with HER2-negative metastatic breast cancer progressing following no more than three lines of chemotherapy. No prior epidermal growth factor receptor-targeted therapy was allowed. Patients received 50-mg afatinib once daily until disease progression. Tumor assessment was performed at every other 28-day treatment course. The primary endpoint was clinical benefit (CB) for ≥4 treatment courses in triple-negative (Cohort A) metastatic breast cancer (TNBC) and objective responses measured by Response Evaluation Criteria in Solid Tumors in patients with HER2-negative, estrogen receptor-positive, and/or progesterone receptor-positive breast cancer (Cohort B). Fifty patients received treatment, including 29 patients in Cohort A and 21 patients in Cohort B. No objective responses were observed in either cohort. Median progression-free survival was 7.4 and 7.7 weeks in Cohorts A and B, respectively. Three patients with TNBC had stable disease for ≥4 treatment courses, one of them for 12 courses (median 26.3 weeks; range 18.9–47.9 weeks). The most frequently observed afatinib-associated adverse events (AEs) were gastrointestinal and skin-related side effects, which were manageable by symptomatic treatment and dose reductions. Afatinib pharmacokinetics were comparable to those observed in previously reported phase I trials. In conclusion, afatinib had limited activity in HER2-negative breast cancer. AEs were generally manageable and mainly affected the skin and the gastrointestinal tract.
Electronic supplementary material
The online version of this article (doi:10.1007/s10549-012-2126-1) contains supplementary material, which is available to authorized users.
Afatinib; Metastatic breast cancer; Triple-negative breast cancer; HER2-negative breast cancer; EGFR TKI
Noninvasive biomarkers are urgently needed for early detection of breast cancer since the risk of recurrence, morbidity and mortality are closely related to disease stage at the time of primary surgery. In the past decade, many proteomics-based approaches were developed that utilize the protein profiling of human body fluids or identification of putative biomarkers to obtain more knowledge on the effects of cancer emergence and progression. Herein, we report on an analysis of proteins in the tear fluid from breast carcinoma patients and healthy women using a de novo proteomic approach and 25 mixed samples from each group. This study included 25 patients with primary invasive breast carcinoma and 25 age-matched healthy controls. We performed a MALDI-TOF-TOF-driven semi-quantitative comparison of tear protein levels in cancer (CA) and control (CTRL) using a de novo approach in pooled samples. Over 150 proteins in the tear fluid of CTRL and CA were identified. Using an in-house-developed algorithm we found more than 20 proteins distinctly upregulated or downregulated in the CTRL and CA groups. We identified several proteins that had modified expression in breast cancer patients. These proteins are involved in host immune system pathways (e.g., C1Q1 or S100A8) and different metabolic cascades (ALDH3A or TPI). Further validation of the results in an independent population combined with individual protein profiling of participants is needed to confirm the specificity of our findings and may lead to a better understanding of the pathological mechanism of breast cancer.
breast cancer; biomarker; diagnosis; proteomics; tear fluids