Identification of structural fingerprints for ABCG2 inhibition by using Monte Carlo optimization,Bayesian classification,and structural and physicochemical interpretation (SPCI) analysis

ABSTRACT

The human breast cancer resistance protein (BCRP), one of the members of the large ATP binding cassette (ABC) transporter superfamily, is crucial for resistance against chemotherapeutic agents. Currently, it has been emerged as one of the best biological targets for the designing of small molecule drugs capable of eliminating multidrug resistance in breast cancer. In order to gain insights into the relationship between the molecular structure of compounds and the ABCG2 inhibition, a multi-QSAR approach using different methods was performed on a dataset of 294 ABCG2 inhibitors with diverse scaffolds. The best models obtained by different chemometric methods have the following statistical characteristics: Monte Carlo Optimization-based QSAR (sensitivity = 0.905, specificity = 0.6255, accuracy = 0.756, and MCC = 0.545), Bayesian classification model (sensitivity = 0.735, specificity = 0.775, and concordance = 0.757); structural and physicochemical interpretation analysis-random forest method (balance accuracy = 0.750, sensitivity = 0.810, and specificity = 0.700). Additionally, structural fingerprints modulating the ABCG2 inhibitory properties were identified from the best models of each method and also validated with each other. The current modelling study is an attempt to get a deep insight into the different important structural fingerprints modulating ABCG2 inhibition.     

Molecular insight into the T798M gatekeeper mutation-caused acquired resistance to tyrosine kinase inhibitors in ErbB2-positive breast cancer

Human epidermal growth factor receptor 2 (ErbB2) is an attractive therapeutic target for metastatic breast cancer. The kinase has been clinically observed to harbor a gatekeeper mutation T798M in its active site, which causes acquired resistance to the first-line targeted breast cancer therapy with small-molecule tyrosine kinase inhibitors. Previously, several theories have been proposed to explain the molecular mechanism of gatekeeper mutation-caused drug resistance, such as blocking of inhibitor binding and increasing of ATP affinity. In the current study, the direct binding of three wild type-selective inhibitors (Lapatinib, AEE788 and TAK-285) and two wild type-sparing inhibitors (Staurosporine and Bosutinib) to the wild-type ErbB2 and its T798M mutant are investigated in detail by using rigorous computational analysis and binding affinity assay. Substitution of the polar threonine with a bulky methionine at residue 798 can impair and improve the direct binding affinity of wild type-selective and wild type-sparing inhibitors, respectively. Hindrance effect is responsible for the affinity decrease of wild type-selective inhibitors, while additional nonbonded interactions contribute to the affinity increase of wild type-sparing inhibitors, thus conferring selectivity to the inhibitors for mutant over wild type. The binding affinity of Staurosporine and Bosutinib to ErbB2 kinase domain is improved by 11.9-fold and 2.1-fold upon T798M mutation, respectively. Structural analysis reveals that a nonbonded network of S–π contact interactions (for Staurosporine) or an S-involving halogen bond (for Bosutinib) forms with the sulfide group of mutant Met798 residue.     

Incorporating human behaviour in simulation models of screening for breast cancer

Simulation modelling is widely used in many industries in order to assess and evaluate alternative options and to test strategies or operating rules which are too complex to be modelled analytically. Simulation software has developed its capability in parallel with the growth in computing power since the 1980s. However in practice, the results from the most sophisticated and complex simulation model may not truly reflect what happens in the real world, because such models do not account for human behaviour. For example, in the domain of healthcare simulation is often used to evaluate the outcomes from medical interventions such as new drug treatments. However in reality patients may not complete the course of a prescribed medication, perhaps because they find the side-effects unpleasant. A simulation study designed to evaluate this medication which ignores such behavioural factors may give unreliable results. In this paper we describe a model for screening for breast cancer which includes behavioural factors to model women’s decisions to attend for mammography. The model results indicate that increasing attendance through education or publicity campaigns can be equally as effective as decreasing the intervals between screens. This would have considerable cost implications for healthcare providers.     

The targeted gene (KDRP-CD/TK) therapy of breast cancer mediated by SonoVue and ultrasound irradiation in vitro

Suicide gene therapy has become an effective therapy for breast cancer, and ultrasound targeted microbubble destruction (UTMD) has become a popular topic in the gene therapy field. In this study, MCF-7 cells with the KDR promoter and LSl74T cells without the KDR promoter were transfected with the recombinant plasmid pEGFP-KDRP-CD/TK using UTMD. The recombinant plasmid pEGFP-KDRP-CD/TK was transfected into MCF-7 and LS174T cells successfully with no significant difference in transfection efficiency (p > 0.05). By RT-PCR, the CD/TK fusion gene was shown to be expressed in MCF-7 cells but not expressed in LS174T cells. In a cytotoxicity experiment, transgenic MCF-7 cells were sensitive to the prodrugs 5-FC and GCV. When both 5-FC and GCV were administered, the rate of cellular inhibition was significantly greater than that achieved when only one of the prodrugs was administered (p < 0.001). Moreover, the inhibition rates achieved administering 5-FC, GCV and both 5-FC and GCV were all significantly greater than the gene transfection rate of 21.92 ± 3.64% (p < 0.001). However, transgenic LS174T cells were not sensitive to any prodrug. These results demonstrated that UTMD is a safe, effective and targeted gene delivery system. Also, the KDR promoter can drive expression of the CD/TK double suicide gene target in MCF-7 cells, and the targeted killing effect of the KDRP-CD/TK gene on MCF-7 cells in vitro has good synergy with expression of the CD/TK fusion gene.     

Magnetic Resonance Imaging (MRI) of hormone-induced breast changes in young premenopausal women

Objectives

We conducted a pilot study to identify whether MRI parameters are sensitive to hormone-induced changes in the breast during the natural menstrual cycle and whether changes could also be observed during an oral contraceptive (OC) cycle.

Materials and Methods

The New York University Langone Medical Center Institutional Review Board approved this HIPAA-compliant prospective study. All participants provided written informed consent. Participants were aged 24-31 years.We measured several non-contrast breast MRI parameters during each week of a single menstrual cycle (among 9 women) and OC cycle (among 8 women). Hormones were measured to confirm ovulation and classify menstrual cycle phase among naturally cycling women and to monitor OC compliance among OC users. We investigated how the non-contrast MRI parameters of breast fibroglandular tissue (FGT), apparent diffusion coefficient (ADC), magnetization transfer ratio (MTR), and transverse relaxation time (T2) varied over the natural and the OC cycles.

Results

We observed significant increases in MRI FGT% and ADC in FGT, and longer T2 in FGT in the luteal vs. follicular phase of the menstrual cycle. We did not observe any consistent pattern of change for any of the MRI parameters among women using OCs.

Conclusions

MRI is sensitive to hormone-induced breast tissue changes during the menstrual cycle. Larger studies are needed to assess whether MRI is also sensitive to the effects of exogenous hormones, such as various OC formulations, on the breast tissue of young premenopausal women.     

Conspicuity of bone metastases on fast Dixon-based multisequence whole-body MRI: Clinical utility per sequence

Purpose

The purpose of the study was to evaluate the conspicuity of bone metastases on each of the numerous sequences produced by fast Dixon-based multisequence whole-body (WB) magnetic resonance imaging (MRI) scanning in order to determine the most clinically useful sequences overall and per anatomic region.

Materials and Methods

Twenty-seven breast cancer patients with bone metastases were prospectively studied with fast Dixon-based WB MRI including head/neck, chest, abdominal, pelvic, thigh, calf/feet and either cervical, thoracic and lumbar or cervical/thoracic and thoracic/lumbar regions. Sequences included coronal T2, axial T1 without and with intravenous gadolinium (+ C), sagittal T1 spine + C, each associated fat-only (FO) and fat-saturated (FS) sequence, axial diffusion-weighted imaging (DWI) and short tau inversion recovery (STIR). Blinded reviewers evaluated lesion conspicuity, a surrogate of clinical utility, on a five-point scale per anatomic region. Sequences were compared using analysis of variance, differences were detected with Tukey's honestly significant difference test, and the four sequences with highest mean conspicuity were compared to the remainder overall and per anatomic region.

Results

Overall, a significant lesion conspicuity difference was found (P < .0001), and lesion conspicuity was significantly higher on FS T1 + C, FO T1 + C, T1 + C sagittal and FS T1 + C axial sequences (P < .0001). Per-region results were the same in the head/neck. Other sequences overlapped with these and included the following: chest/abdomen — FO T2, DWI; pelvis — DWI, FO T2; thigh — FS T2, FO T2, FO T1 + C; calf/feet — FS T2, DWI, FO T2, STIR.

Conclusion

Overall, bone lesions were most conspicuous on FS T1 + C sagittal, FO T1 + C sagittal, T1 + C sagittal and FS T1 + C axial fast Dixon WB MRI sequences.     

Alpha 1-antitrypsin: a novel tumor-associated antigen identified in patients with early-stage breast cancer

Several studies have demonstrated that sera from patients with cancer contain antibodies that recognize a unique group of autologous antigens called tumor-associated antigens (TAA). In the current study, we employed an immunoproteomic approach, combining 2DE, Western blot, and MALDI-MS to identify TAA in the sera of patients diagnosed with infiltrating ductal or in situ carcinoma breast cancer. Sera obtained from 25 newly diagnosed patients with stage II breast cancer and 20 healthy volunteers was evaluated for the presence of novel TAA. Alpha 1-antitrypsin (A1AT) antibodies were detected in 24 of 25 patients with breast cancer (96%) and in 2 of 20 controls (10%). Sensitivity of detection of autoantibodies against A1AT in patients with breast cancer was 96%. Our preliminary results suggest that A1AT and autoantibodies against alpha 1 antitrypsin may be useful serum biomarkers for early-stage breast cancer screening and diagnosis.     

Tandem lectin affinity chromatography monolithic columns with surface immobilised concanavalin A, wheat germ agglutinin and Ricinus communis agglutinin-I for capturing sub-glycoproteomics from breast cancer and disease-free human sera

In this study, a liquid-phase separation platform consisting of tandem lectin affinity chromatography was introduced for the selective capturing of sub-glycoproteomics that are affected in cancers, e.g. breast cancer. The platform is comprised of three monolithic columns with surface immobilised lectins including concanavalin A (Con A), wheat germ agglutinin (WGA) and Ricinus communis agglutinin-I (RCA-I). While WGA and Con A have specificities directed towards the core portion of N-glycans on the glycoprotein surface, RCA-I specifically interacts with the non-reducing terminal moieties of the outer chain structures of N-glycans. The effects of the order in which the three lectin columns were arranged in the tandem columns format were evaluated. The most suitable order proved to be WGA → Con A → RCA-I (denoted as WCR) as far as the number of captured proteins was concerned. The WCR tandem columns allowed the capture of 113 and 112 proteins from disease-free and breast cancer sera, respectively, corresponding to 75 and 65 non-redundant proteins, respectively. Using mass spectral count ratios and Q-Q plots yielded a panel of 23 non-redundant differentially expressed proteins (i.e. a panel of 23 candidate markers), which should in principle be more representative of a pathophysiological state than a single marker candidate.     

Crosstalk events in the estrogen signaling pathway may affect tamoxifen efficacy in breast cancer molecular subtypes

Steroid hormones are involved on cell growth, development and differentiation. Such effects are often mediated by steroid receptors. One paradigmatic example of this coupling is the estrogen signaling pathway. Its dysregulation is involved in most tumors of the mammary gland. It is thus an important pharmacological target in breast cancer. This pathway, however, crosstalks with several other molecular pathways, a fact that may have consequences for the effectiveness of hormone modulating drug therapies, such as tamoxifen. For this work, we performed a systematic analysis of the major routes involved in crosstalk phenomena with the estrogen pathway – based on gene expression experiments (819 samples) and pathway analysis (493 samples) – for biopsy-captured tissue and contrasted in two independent datasets with in vivo and in vitro pharmacological stimulation. Our results confirm the presence of a number of crosstalk events across the estrogen signaling pathway with others that are dysregulated in different molecular subtypes of breast cancer. These may be involved in proliferation, invasiveness and apoptosis-evasion in patients. The results presented may open the way to new designs of adjuvant and neoadjuvant therapies for breast cancer treatment.     

Evaluation of Liquid–Liquid Extraction Methods for Determining the Levels of Lipids and Organochlorine Pollutants in Human Milk

Organic chlorine pesticides and polychlorinated biphenyls are organic pollutants that are stored in the adipose tissues of humans. The concentrations of those pollutants in human milk have previously been utilized as a biomarker for monitoring the body burden of lipophilic pollutants in humans. Liquid–liquid extraction methods have been applied to lipids, chlorinated pesticides, and polychlorinated biphenyls from breast milk. In this study, the effectiveness of four extraction methods, which have been widely used to isolate fats and organic pollutants from milk, were compared. The organic solvents included hexane, hexane/acetone (2:1, v/v), ethanol/ethyl ether/hexane (2:3:4, v/v), and methanol/chloroform (1:1, v/v). These results demonstrated that hexane yields the lowest extraction recoveries for lipids (approximately 21.3%) and analytes (approximately 50%). The other three organic solvents demonstrated better performance in extracting the target compounds, with the ethanol/ethyl ether/hexane system showing the optimum efficiency. The optimized system was employed to determine the analytes in human milk.