[Show abstract][Hide abstract] ABSTRACT: Epithelial–mesenchymal transition (EMT) is a key event that is involved in the invasion and dissemination of cancer cells. Although typically considered as having tumour-suppressive properties, transforming growth factor (TGF)-β signalling is altered during cancer and has been associated with the invasion of cancer cells and metastasis. In this study, we report a previously unknown role for the cytoplasmic promyelocytic leukaemia (cPML) tumour suppressor in TGF-β signalling-induced regulation of prostate cancer-associated EMT and invasion. We demonstrate that cPML promotes a mesenchymal phenotype and increases the invasiveness of prostate cancer cells. This event is associated with activation of TGF-β canonical signalling pathway through the induction of Sma and Mad related family 2 and 3 (SMAD2 and SMAD3) phosphorylation. Furthermore, the cytoplasmic localization of promyelocytic leukaemia (PML) is mediated by its nuclear export in a chromosomal maintenance 1 (CRM1)-dependent manner. This was clinically tested in prostate cancer tissue and shown that cytoplasmic PML and CRM1 co-expression correlates with reduced disease-specific survival. In summary, we provide evidence of dysfunctional TGF-β signalling occurring at an early stage in prostate cancer. We show that this disease pathway is mediated by cPML and CRM1 and results in a more aggressive cancer cell phenotype. We propose that the targeting of this pathway could be therapeutically exploited for clinical benefit.
[Show abstract][Hide abstract] ABSTRACT: The expected five-year survival rate from a stage III ovarian cancer diagnosis is a mere 22%; this applies to the 7000 new cases diagnosed yearly in the UK. Stratification of patients with this heterogeneous disease, based on active molecular pathways, would aid a targeted treatment improving the prognosis for many cases. While hundreds of genes have been associated with ovarian cancer, few have yet been verified by peer research for clinical significance. Here, a meta-analysis approach was applied to two carefully selected gene expression microarray datasets. Artificial neural networks, Cox univariate survival analyses and T-tests identified genes whose expression was consistently and significantly associated with patient survival. The rigor of this experimental design increases confidence in the genes found to be of interest. A list of 56 genes were distilled from a potential 37,000 to be significantly related to survival in both datasets with a FDR of 1.39859 × 10−11, the identities of which both verify genes already implicated with this disease and provide novel genes and pathways to pursue. Further investigation and validation of these may lead to clinical insights and have potential to predict a patient’s response to treatment or be used as a novel target for therapy.
[Show abstract][Hide abstract] ABSTRACT: Background and Objectives
Advantages of using cord blood (CB) over other sources of haematopoietic progenitor cells, such as bone marrow, include the ability to cryopreserve and bank the samples until requested for a transplant. Cryopreservation requires the addition of a cryoprotectant to prevent the formation of intracellular ice during freezing. Dimethyl sulphoxide (DMSO) is commonly used at a concentration of 10% (v/v); however, there is evidence to suggest this chemical is toxic to cells as well as to patients after infusion.Materials and Methods
The toxic effects of DMSO were assessed through cell viability and in vitro functional assays in fresh and post-thaw CB samples before determining the maximum exposure time and optimal concentration for cryopreservation.ResultsA dose-dependent toxicity of DMSO was observed in fresh samples with 40% removing all viable and functional haematopoietic progenitor cells (HPC). In fresh and post-thaw analysis, minimal toxic effect was observed when cryopreservation was delayed for up to 1 h after 10% DMSO addition. After thawing, DMSO washout was superior to dilution or unmanipulated when maintained for long periods (advantage observed 1 h after thawing). Finally, the optimum concentration for cryopreserving CB was found to be 7·5 to 10% with detrimental effects observed outside of this range.Conclusion
These results support the use of 7·5–10% as the optimal DMSO concentration and the maximum exposure time should be limited to <1 h prior to freezing and 30 min post-thaw.
[Show abstract][Hide abstract] ABSTRACT: Developing materials that can preferentially select defined cancer cell populations for biological characterization will greatly enhance our understanding of cancer cell growth, differentiation, and invasion. The transitional events between epithelial and mesenchymal phenotypes are particularly crucial, as primary tumors and secondary metastasis are generally epithelial in nature, whereas circulating mesenchymal cells derived from primary epithelial cells appear to facilitate the spread of disease and its resistance to therapy. This study describes an amino-functionalized material, which promotes the enrichment of an epithelial phenotype from a single cell line containing both epithelial and mesenchymal subpopulations of cancer cells. The isolation and transitional control of such subpopulations using functional materials will advance understanding of the disease process, have a significant impact on the downstream development of new targeted cancer therapeutics, and also be applicable to tissue engineering and regenerative medicine.
Full-text · Article · Mar 2015 · Advanced Healthcare Materials
[Show abstract][Hide abstract] ABSTRACT: The aim of the immunotherapy and peptide therapeutics is to destroy cancer cells without eliciting lethal side effects and sustain the long-lasting immune memory. Targeting major histocompatibility complex (MHC) class I and class II epitopes and multiple tumor antigens has met with some clinical success, which is advantageous over single epitope vaccine strategies. Moreover, cancer immunotherapy and peptide therapeutics especially have demonstrated improved responses over that of the conventional treatments in patient groups where treatment options are limited. However, there are still obstacles to combating advanced cancer, where the immune system of the host is compromised. Thus implementing ways of overcoming states of immune tolerance, anergy, or suppression concomitant with a vaccine strategy to enhance adaptive immunity offers an attractive route for clinical intervention. Targeted therapies against defined tumor antigens involving the use of peptide-based vaccination offer potential for the future. Novel therapeutics relies on knowledge of peptide epitopes and awareness of how these can be used to activate appropriate tumor immunity. In this review we discuss many of the tumor antigens that have been discovered in the past two decades and their clinical utility alongside conventional cancer therapies.
[Show abstract][Hide abstract] ABSTRACT: Proliferation is the most fundamental hallmark of cancer. In breast cancer, proliferation is assessed by staining cells for the nuclear antigen Ki-67. Ki-67 has a potential use as a prognostic and predictive factor for therapy in breast cancer. In this study, an Artificial neural network (ANN) using a feedforward multi-layer perceptron back propagation algorithm was applied to facilitate a systems biology viewpoint to identify genes associated with Ki-67 in 3 different breast cancer gene expression studies. Subsequently, a non-reductionist ANN based network was created for the top ten STRING identified genes for Ki-67 to identify the cross linked hubs (UBE2C, KIF2C) in the map. The genes which were found in common among the top 200 probes in the three cohorts and as cross links in the map were hypothesized to be more powerful and found to be significantly associated with prognosis in breast cancer. In future, this work can be expanded to include other proliferation associated targets to map the system in a more detailed way and see the influence of the common markers obtained in drug resistance (endocrine resistance) in breast cancer. The network map constructed, represents a novel non-recursive additive approach to visualize a gene network and provides a dynamic view for the disease of interest.
[Show abstract][Hide abstract] ABSTRACT: Background: The delivery of specific immunotherapies for malignant tumours requires the identification of relevant tumour antigens and
sequences from these which can be used to stimulate protective T cell-mediated immunity. HAGE (DDX43) is a cancer testis antigen
belonging to the DEAD box family of helicases found by our group to be over-expressed in many solid cancers including breast
cancer (Mathieu et al.1), immunogenic (Mathieu et al.2) and to be a biomarker for poor prognosis as well as a predictor of chemotherapy response in breast cancer (Abdel-fatah et
al.3). We propose that HAGE might be a novel immunotherapeutic target for patients bearing breast cancers expressing this antigen.
The aim of this study is to identify strongly immunogenic HAGE-derived sequences which can be used for the development of
a therapeutic vaccine for HAGE positive cancer.
Experimental design: The HAGE-derived sequences were identified and assessed after: (i) using a computer-based epitope predictive tool; (ii)
determining the affinity of the predicted peptide with HLA-A2.1 molecules using a peptide-binding assay; (iii) confirming
the natural endogenous processing of identified peptides using humanised HHDII/DR1 transgenic mice expressing HLA-A2 and HLA-DR1
molecules but no murine Class-I or Class-II molecules.
Results: Two HAGE-derived sequences (a 24 amino acids long and a 30 amino acids long) encompassing several immunogenic epitopes and
exhibiting a broad HLA binding spectrum have been identified and their immunogenicity assessed.
Conclusions: The cancer restricted and elevated expression of HAGE in several cancers makes this particular antigen a promising molecule
on which to base the development of new T cell-based immunotherapeutic strategies. The novel HAGE-derived sequences identified
here might therefore facilitate the development of long peptide-based, broadly applicable cancer vaccines.
References: 1Cancer Immun. 2010 Jan 11;10:2. 2Cancer Immunol Immunother. 2007 Dec; 56(12):1885-95. Epub 2007 May 9. 3Br J Cancer. 2014 May 13;110(10):2450-61. doi: 10.1038/bjc.2014.168. Epub 2014 Apr 22.
Disclosure: All authors have declared no conflicts of interest.
Full-text · Article · Nov 2014 · Annals of Oncology
[Show abstract][Hide abstract] ABSTRACT: Under normal conditions the detachment of anchorage-dependant cells from their extracellular matrix typically induces programmed cell death which is mediated through a pathway referred to as anoikis. However, a resistance to anoikis in cancer enables the migration of cells from the primary tumour and the establishment of aggressive metastatic disease. Although cancer cell aggregation is known to be an important mechanism within anoikis resistance, research into the underlying mechanisms that govern this process remain problematic as commercially available tissue culture material can only sustain 2D monolayer or 3D aggregate/spheroid cultures. This necessitates the development of a system that can accommodate for cancer cell aggregation–disaggregation as a single dynamic process, without the disruption of passaging cells between alternate substrates. This study describes a procedure for modifying tissue culture polystyrene (TCP) to produce a fluoro-silica (FS) surface which preferentially promotes the deposition of a distinct profile of proteins/factors from serum which mediate the transient aggregation of human breast cancer cell lines. This modified surface therefore provides an experimental platform for better understanding cancer cell aggregation–disaggregation events in vitro, and their influence on the establishment of metastatic disease in patients with cancer.
[Show abstract][Hide abstract] ABSTRACT: Background:
HAGE protein is a known immunogenic cancer-specific antigen.
The biological, prognostic and predictive values of HAGE expression was studied using immunohistochemistry in three cohorts of patients with BC (n=2147): early primary (EP-BC; n=1676); primary oestrogen receptor-negative (PER-BC; n=275) treated with adjuvant anthracycline-combination therapies (Adjuvant-ACT); and primary locally advanced disease (PLA-BC) who received neo-adjuvant anthracycline-combination therapies (Neo-adjuvant-ACT; n=196). The relationship between HAGE expression and the tumour-infiltrating lymphocytes (TILs) in matched prechemotherapy and postchemotherapy samples were investigated.
Eight percent of patients with EP-BC exhibited high HAGE expression (HAGE+) and was associated with aggressive clinico-pathological features (Ps<0.01). Furthermore, HAGE+expression was associated with poor prognosis in both univariate and multivariate analysis (Ps<0.001). Patients with HAGE+did not benefit from hormonal therapy in high-risk ER-positive disease. HAGE+and TILs were found to be independent predictors for pathological complete response to neoadjuvant-ACT; P<0.001. A statistically significant loss of HAGE expression following neoadjuvant-ACT was found (P=0.000001), and progression-free survival was worse in those patients who had HAGE+residual disease (P=0.0003).
This is the first report to show HAGE to be a potential prognostic marker and a predictor of response to ACT in patients with BC.
Full-text · Article · Apr 2014 · British Journal of Cancer
[Show abstract][Hide abstract] ABSTRACT: A stage III ovarian cancer diagnosis yields a 22% 5-year survival rate, this applies to over half of the 7000 new cases diagnosed each year in the UK. Stratification of patients with this heterogeneous disease, based on active molecular pathways in their cancer, would aid a targeted treatment and improve prognosis. Hundreds of genes have been significantly associated with ovarian cancer, few have yet been verified. Exploration of published microarray data sets using Artificial Neural Networks confirmed the robustness of PRELP as a biomarker for survival time from stage III ovarian cancer, and generated a new panel of 44 genes that significantly predicted survival length of a blind valida-tion set (p=0.00073).
[Show abstract][Hide abstract] ABSTRACT: Treatment options for patients with advanced prostate cancer remain limited and rarely curative. Prostatic acid phosphatase (PAP) is a prostate-specific protein over-expressed in 95% of prostate tumours. An FDA-approved vaccine for the treatment of advanced prostate disease, PROVENGE® (sipuleucel-T), has been shown to prolong survival, however the precise sequence of the PAP protein responsible for the outcome is unknown. As the PAP antigen is one of the very few prostate-specific antigens for which there is a rodent equivalent with high homology, pre-clinical studies using PAP have the potential to be directly relevant to clinical setting. Here we show three PAP epitopes naturally processed and presented in the context of HHDII/DR1 (114-128, 299-313, and 230-244). The PAP-114-128 epitope elicits CD4(+) and CD8(+) T-cell-specific responses in C57BL/6 mice. Furthermore, when immunised in a DNA vector format (ImmunoBody®), PAP-114-128 prevents and reduces the growth of TRAMP-C1 prostate cancer cell-derived tumours in both prophylactic and therapeutic settings. This anti-tumour effect is associated with infiltration of CD8(+) tumour-infiltrating lymphocytes (TILs) and the generation of high avidity T cells secreting elevated levels of IFN-γ. PAP-114-128 therefore appears to be a highly relevant peptide on which to base vaccines for the treatment of prostate cancer. This article is protected by copyright. All rights reserved.
No preview · Article · Apr 2014 · European Journal of Immunology
[Show abstract][Hide abstract] ABSTRACT: The tumour suppressor PML (promyelocytic leukaemia protein) regulates several cellular pathways involving cell growth, apoptosis, differentiation and senescence. PML also has an important role in the regulation of stem cell proliferation and differentiation. Here, we show the involvement of the helicase HAGE in the transcriptional repression of PML expression in ABCB5+ malignant melanoma-initiating cells (ABCB5+ MMICs), a population of cancer stem cells which are responsible for melanoma growth, progression and resistance to drug-based therapy. HAGE prevents PML gene expression by inhibiting the activation of the JAK-STAT (janus kinase-signal transducers and activators of transcription) pathway in a mechanism which implicates the suppressor of cytokine signalling 1 (SOCS1). Knockdown of HAGE led to a significant decrease in SOCS1 protein expression, activation of the JAK-STAT signalling cascade and a consequent increase of PML expression. To confirm that the reduction in SOCS1 expression was dependent on the HAGE helicase activity, we showed that SOCS1, effectively silenced by small interfering RNA, could be rescued by re-introduction of HAGE into cells lacking HAGE. Furthermore, we provide a mechanism by which HAGE promotes SOCS1 mRNA unwinding and protein expression in vitro. Finally, using a stem cell proliferation assay and tumour xenotransplantation assay in non-obese diabetic/severe combined immunodeficiency mice, we show that HAGE promotes MMICs-dependent tumour initiation and tumour growth by preventing the anti-proliferative effects of interferon-α (IFNα). Our results suggest that the helicase HAGE has a key role in the resistance of ABCB5+ MMICs to IFNα treatment and that cancer therapies targeting HAGE may have broad implications for the treatment of malignant melanoma.
Full-text · Article · Feb 2014 · Cell Death & Disease
[Show abstract][Hide abstract] ABSTRACT: Vaccine-based immunotherapy can increase the overall survival of patients with advanced prostate cancer. However, the efficacy of vaccine-elicited anticancer immune responses is heavily influenced by the physical, nutritional, and psychological status of the patient. Given their importance, these parameters should be carefully considered for the design of future clinical trials testing this immunotherapeutic paradigm in prostate cancer patients.
[Show abstract][Hide abstract] ABSTRACT: Oestrogen receptor (ER) positive (luminal) tumours account for the largest proportion of females with breast cancer. Theirs is a heterogeneous disease presenting clinical challenges in managing their treatment. Three main biological luminal groups have been identified but clinically these can be distilled into two prognostic groups in which Luminal A are accorded good prognosis and Luminal B correlate with poor prognosis. Further biomarkers are needed to attain classification consensus. Machine learning approaches like Artificial Neural Networks (ANNs) have been used for classification and identification of biomarkers in breast cancer using high throughput data. In this study, we have used an artificial neural network (ANN) approach to identify DACH1 as a candidate luminal marker and its role in predicting clinical outcome in breast cancer is assessed.
[Show abstract][Hide abstract] ABSTRACT: The effect of alterations in virulence and transformation by long term in vitro culture of L. mexicana promastigotes on infectivity and immune responses was investigated. Fresh parasite cultures harvested from Balb/c mice were passaged 20 times in vitro. Infectivity was decreased and was completely avirulent after 20 passages. The qPCR results showed a down regulation of GP63, LPG2, CPC, CPB2, CPB2.8, CHT1, LACK and LDCEN3 genes after passage 7 concomitant with a reduced and absence of infectivity by passages 7 and 20, respectively. Parasites at passages 1 and 20 are referred to as virulent and avirulent, respectively. The growth of avirulent and virulent parasite was affected by conditioned media derived from macrophages or monocytes infected with parasites for 2 hours. Giemsa staining showed the failure of avirulent but not virulent parasites to transform to the amastigote stage in infected host cells. with both virulent and avirulent modulating the expression of CCL-22, Tgad51, Cox2, IL-1, IL-10, TGF-β, TNF-α, Rab7, Rab9 and A2 genes; virulent but not avirulent L. mexicana significantly up regulated Th2 associated cytokines, but down regulated Rab7and Rab9 gene expression. In conclusion, a model for L. mexicana is reported, which is of potential value in studying host- parasite interaction. This article is protected by copyright. All rights reserved.
No preview · Article · Jul 2013 · Parasite Immunology