Article

Metabolomic profiling of prostate cancer by matrix assisted laser desorption/ionization-Fourier transform ion cyclotron resonance mass spectrometry imaging using Matrix Coating Assisted by an Electric Field (MCAEF)

Authors:
  • National Institute for Viral Disease Control and Prevention
  • University of Victoria Genome BC Proteomics Centre
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Abstract

In this work, we combined the use of two MALDI matrices (quercetin and 9-aminoacridine), a recently developed new matrix coating technique – matrix coating assisted by an electric field (MCAEF), and matrix-assisted laser desorption/ionization – Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FTICRMS) to detect and image endogenous compounds in the cancerous and non-cancerous regions of three human prostate cancer (stage II) tissue specimens. After three rounds of imaging data acquisitions (i.e., quercetin for positive and negative ion detection and 9-aminoacridine for negative ion detection), and metabolite identification, a total of 1091 metabolites including 1032 lipids and 59 other metabolites were routinely detected and successfully localized. Of these compounds, 250 and 217 were only detected in either the cancerous or the non-cancerous regions respectively, although we cannot rule out the presence of these metabolites at concentrations below the detection limit. In addition, 152 of the other 624 metabolites showed differential distributions (p < 0.05, t-test) between the two regions of the tissues. Further studies on a larger number of clinical specimens will need to be carried out to confirm this large number of apparently cancer-related metabolites. The successful determination of the spatial locations and abundances of these endogenous biomolecules indicated significant metabolism abnormalities – e.g., increased energy charge and under-expression of neutral acyl glycerides, in the prostate cancer samples. To our knowledge, this work has resulted in MALDI-MS imaging of the largest group of metabolites in prostate cancer thus far and demonstrated the importance of using complementary matrices for comprehensive metabolomic imaging by MALDI-MS. This article is part of a Special Issue entitled: MALDI Imaging, edited by Dr. Corinna Henkel and Prof. Peter Hoffmann.

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... Understanding the molecular processes specific to each tissue type may provide novel and clinically important insights into the molecular mechanisms of prostate cancer compared to healthy prostate epithelium. Mass spectrometry imaging (MSI) allows for spatial detection of several different classes of potential cancer markers on tissue sections, including metabolites [2,3], lipids [4,5], peptides [6], glycans [7], and metals [8], which can be matched to corresponding histology images. ...
... Phospholipid mapping by MSI is particularly beneficial due to their high metabolic stability, amphiphilic chemistry, and confinement to membranes, making them robust against diffusion. Several studies using MSI have demonstrated alterations in phospholipid composition in prostate cancer compared to healthy prostate tissue [3,4,15,16,19,23]. ...
... Hence, the percentage of stroma content is not balanced between normal and cancer samples, making stroma a confounding factor for prostate tissue bulk analysis [1]. Previous MSI analysis of prostate tissue have either excluded stroma tissue from data analysis [4,15,23] or combined stroma tissue together with normal epithelial tissue [3]. In the latter case, observed metabolic differences between cancer and normal tissue can be a result of reduced presence of stroma in cancer tissue rather than a difference between cancer and healthy glands. ...
Article
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Background Prostate cancer tissues are inherently heterogeneous, which presents a challenge for metabolic profiling using traditional bulk analysis methods that produce an averaged profile. The aim of this study was therefore to spatially detect metabolites and lipids on prostate tissue sections by using mass spectrometry imaging (MSI), a method that facilitates molecular imaging of heterogeneous tissue sections, which can subsequently be related to the histology of the same section. Methods Here, we simultaneously obtained metabolic and lipidomic profiles in different prostate tissue types using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MSI. Both positive and negative ion mode were applied to analyze consecutive sections from 45 fresh-frozen human prostate tissue samples ( N = 15 patients). Mass identification was performed with tandem MS. Results Pairwise comparisons of cancer, non-cancer epithelium, and stroma revealed several metabolic differences between the tissue types. We detected increased levels of metabolites crucial for lipid metabolism in cancer, including metabolites involved in the carnitine shuttle, which facilitates fatty acid oxidation, and building blocks needed for lipid synthesis. Metabolites associated with healthy prostate functions, including citrate, aspartate, zinc, and spermine had lower levels in cancer compared to non-cancer epithelium. Profiling of stroma revealed higher levels of important energy metabolites, such as ADP, ATP, and glucose, and higher levels of the antioxidant taurine compared to cancer and non-cancer epithelium. Conclusions This study shows that specific tissue compartments within prostate cancer samples have distinct metabolic profiles and pinpoint the advantage of methodology providing spatial information compared to bulk analysis. We identified several differential metabolites and lipids that have potential to be developed further as diagnostic and prognostic biomarkers for prostate cancer. Spatial and rapid detection of cancer-related analytes showcases MALDI-TOF MSI as a promising and innovative diagnostic tool for the clinic.
... MSI identification of prostate cancer-specific metabolites MSI investigation of tissue specimens represents an innovative field in PCa research. The following studies have focused on either the diagnosis or prognosis of PCa by considering either single metabolites or metabolomic profiles quantified from MALDI or DESI-MSI and have reported differences in expression profiles between benign and malignant human prostate tissue [64][65][66][67][68][69][70][71]. A number of metabolites have been identified as potential diagnostic markers for PCa by means of MSI, such as mitogenactivated protein kinase/extracellular signal-regulated kinase (MEKK2), cholesterol sulfate, phosphatidylinositols, and biliverdin reductase B (BVR) [64][65][66]68]. ...
... In a recently published work, endogenous compounds of cancerous and non-cancerous regions in three human prostate cancer tissue specimens were detected and imaged using MALDI-FTICR-MSI [71]. The authors combined two different MALDI matrices (quercetin and 9-aminoacridine) and used the newly developed technique of matrix coating assisted by an electric field [80,81]. ...
... Of these, 152 metabolites showed differentiating distributions between these two regions of tissue, such as an increased energy charge and a lower expression of neutral acyl-glycerides in the cancerous regions. Sixty-two acyl-glycerides were detectable only in non-cancerous regions, which can be interpreted as a result of known low glycolysis in prostate cancer cells and compensatory increased fatty acid beta-oxidation leading to a depletion of these acyl-glycerides in cancerous tissue regions [71,82]. ...
Article
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Background: Prostate cancer (PCa), the most common cancer and second leading cause of cancer death in American men, presents the clinical challenge of distinguishing between indolent and aggressive tumors for proper treatment. PCa presents significant alterations in metabolic pathways that can potentially be measured using techniques like mass spectrometry (MS) or MS imaging (MSI) and used to characterize PCa aggressiveness. MS quantifies metabolomic, proteomic, and lipidomic profiles of biological systems that can be further visualized for their spatial distributions through MSI. Methods: PubMed was queried for all publications relating to MS and MSI in human PCa from April 2007 to April 2017. With the goal of reviewing the utility of MSI in diagnosis and prognostication of human PCa, MSI articles that reported investigations of PCa-specific metabolites or metabolites indicating PCa aggressiveness were selected for inclusion. Articles were included that covered MS and MSI principles, limitations, and applications in PCa. Results: We identified nine key studies on MSI in intact human prostate tissue specimens that determined metabolites which could either differentiate between benign and malignant prostate tissue or indicate PCa aggressiveness. These MSI-detected biomarkers show promise in reliably identifying PCa and determining disease aggressiveness. Conclusions: MSI represents an innovative technique with the ability to interrogate cancer biomarkers in relation to tissue pathologies and investigate tumor aggressiveness. We propose MSI as a powerful adjuvant histopathology imaging tool for prostate tissue evaluations, where clinical translation of this ex vivo technique could make possible the use of MSI for personalized medicine in diagnosis and prognosis of PCa. Moreover, the knowledge provided from this technique can majorly contribute to the understanding of molecular pathogenesis of PCa and other malignant diseases.
... 针 对基质的喷涂方式, 传统喷涂方法包括手动气喷雾 (airbrush) [36] 、电喷雾(electrospray) [37] 、基质升华(sublimation) [38] 等. 手动气喷雾方法具有操作简便、快速 [41] . 相比于传统常规喷涂方式, 利用该技术可直 接原位表征大约1000多种脂类分子 [40] 、50多种核苷类 /能量代谢相关分子 [40] 、近250种蛋白质分子 [39] , 使得 ...
... 针 对基质的喷涂方式, 传统喷涂方法包括手动气喷雾 (airbrush) [36] 、电喷雾(electrospray) [37] 、基质升华(sublimation) [38] 等. 手动气喷雾方法具有操作简便、快速 [41] . 相比于传统常规喷涂方式, 利用该技术可直 接原位表征大约1000多种脂类分子 [40] 、50多种核苷类 /能量代谢相关分子 [40] 、近250种蛋白质分子 [39] , 使得 ...
... Glycerophospholipids, such as phosphatidylcholines (PCs) and phosphatidylethanolamine (PEs), were among the main contributors to region separation. [29][30][31]. It is well known that PCs are the most predominant components of phospholipids for biological membranes; PC metabolism is altered in the onset and development of many tumors [32,33]. ...
... It is well known that PCs are the most predominant components of phospholipids for biological membranes; PC metabolism is altered in the onset and development of many tumors [32,33]. In particular, the elevated levels of PC 32:1 and PC 34:1 have been positively correlated to the cancerous areas of colorectal, prostate, and breast cancer tissues [29][30][31]. Interestingly, PC 32:0 was previously found discriminatory in the Warthin tumor region of benign salivary gland tumor tissues analyzed by MALDI-MSI [20]. ...
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Salivary gland tumors are relatively uncommon neoplasms that represent less than 5% of head and neck tumors, and about 90% are in the parotid gland. The wide variety of histologies and tumor characteristics makes diagnosis and treatment challenging. In the present study, Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) was used to discriminate the pathological regions of patient-derived biopsies of parotid neoplasms by metabolomic and lipidomic profiles. Fresh frozen parotid tissues were analyzed by MALDI time-of-flight (TOF) MSI, both in positive and negative ionization modes, and additional MALDI-Fourier-transform ion cyclotron resonance (FT-ICR) MSI was carried out for metabolite annotation. MALDI-TOF-MSI spatial segmentation maps with different molecular signatures were compared with the histologic annotation. To maximize the information related to specific alterations between the pathological and healthy tissues, unsupervised (principal component analysis, PCA) and supervised (partial least squares-discriminant analysis, PLS-DA) multivariate analyses were performed presenting a 95.00% accuracy in cross-validation. Glycerophospholipids significantly increased in tumor tissues, while sphingomyelins and triacylglycerols, key players in the signaling pathway and energy production, were sensibly reduced. In addition, a significant increase of amino acids and nucleotide intermediates, consistent with the bioenergetics request of tumor cells, was observed. These results underline the potential of MALDI-MSI as a complementary diagnostic tool to improve the specificity of diagnosis and monitoring of pharmacological therapies.
... The enhanced LPC levels in normal tissue may reflect increased activity of lysophospholipase D activity and PC remodelling pathways. Wang and colleagues conducted metabolomic imaging of prostate tissue (n = 3 patients) by MALDI FT-ICR [59], using LC-MS/MS as a structural validation tool. In the study, m/z 534.296 ...
... AMP and m/z 426.022 ADP were diminished consistent with increased ATP flux that is critical for tumour cell proliferation [59]. ...
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Due to advances in the detection and management of prostate cancer over the past 20 years, most cases of localised disease are now potentially curable by surgery or radiotherapy, or amenable to active surveillance without treatment. However, this has given rise to a new dilemma for disease management; the inability to distinguish indolent from lethal, aggressive forms of prostate cancer, leading to substantial overtreatment of some patients and delayed intervention for others. Driving this uncertainty is the critical deficit of novel targets for systemic therapy and of validated biomarkers that can inform treatment decision-making and to select and monitor therapy. In part, this lack of progress reflects the inherent challenge of undertaking target and biomarker discovery in clinical prostate tumours, which are cellularly heterogeneous and multifocal, necessitating the use of spatial analytical approaches. In this review, the principles of mass spectrometry-based lipid imaging and complementary gene-based spatial omics technologies, their application to prostate cancer and recent advancements in these technologies are considered. We put in perspective studies that describe spatially-resolved lipid maps and metabolic genes that are associated with prostate tumours compared to benign tissue and increased risk of disease progression, with the aim of evaluating the future implementation of spatial lipidomics and complementary transcriptomics for prognostication, target identification and treatment decision-making for prostate cancer.
... The authors showed that phosphatidylcholines, phosphatidic acids, phosphatidylserines, phosphatidylinositols, and cardiolipins were overexpressed in GS (4 + 3), suggesting their involvement in the disease progression, and the possibility for them to be utilized as markers of prostate cancer aggressiveness [60]. The results from this study agree with the outcome of another pilot study by Wang and coworkers [61]. ...
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Novel profiling methodologies are redefining the diagnostic capabilities and therapeutic approaches towards more precise and personalized healthcare. Complementary information can be obtained from different omic approaches in combination with the traditional macro- and microscopic analysis of the tissue, providing a more complete assessment of the disease. Mass spectrometry imaging, as a tissue typing approach, provides information on the molecular level directly measured from the tissue. Lipids, metabolites, glycans, and proteins can be used for better understanding imbalances in the DNA to RNA to protein translation, which leads to aberrant cellular behavior. Several studies have explored the capabilities of this technology to be applied to tumor subtyping, patient prognosis, and tissue profiling for intraoperative tissue evaluation. In the future, intercenter studies may provide the needed confirmation on the reproducibility, robustness, and applicability of the developed classification models for tissue characterization to assist in disease management.
... This technique increases both the total ion count in a mass spectrum as well as the signalto-noise ratio (S/N) by up to a factor of 5. This technique has been applied to metabolite analysis in porcine adrenal gland (Wang et al., 2015), a proteomic analysis of prostate-cancer tissue (Wang et al., 2016), and a metabolomic analysis of prostatecancer tissue (Wang et al., 2017). Herein, we applied MCAEF to examine the distinctive metabolite profiles of five tadpole organs and to detect over 1000 metabolites in each the five tissue types with 1700 unique metabolites. ...
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Anuran metamorphosis involves the transformation of an aquatic tadpole into a juvenile frog. This process is completely dependent upon thyroid hormones (THs). Although much research has been focused on changes in gene expression programs during this postembryonic developmental period, transitions in the metabolic profiles are relatively poorly understood. Matrix Assisted Laser Desorption/Ionization-Mass Spectrometry Imaging (MALDI-MSI) is a technique that generates highly multiplexed mass spectra while retaining spatial location information on a thin tissue section. Reconstructed ion heat maps are correlated with morphology of the tissue section for biological interpretation. The present study is the first to use whole-body MALDI-MSI on tadpoles to gain insights into anuran metamorphosis. Approximately 1000 features were detected in each of five tissues examined (brain, eye, liver, notochord, and tail muscle) from premetamorphic North American bullfrog (Rana [Lithobates] catesbeiana) tadpoles. Of these detected metabolites, 1700 were unique and 136 were significantly affected by exposure to 50 nM thyroxine for 48 h. Of the significantly-affected metabolites, 64 features were tentatively identified using the MassTRIX annotation tool. All tissues revealed changes in lipophilic compounds including phosphatidylcholines, phosphatidylinositols, phosphatidylglycerols, phosphatidylethanolamines, and phosphatidylserines. These lipophilic compounds made up the largest portion of significantly-affected metabolites indicating that lipid signaling is a major target of TH action in frog tadpoles.
... The generation of very small crystals (<10 μm) of common matrices resulted in an enhancement of detection sensitivity and a number of LMWC (fatty acids, nucleosides, monophosphate, nucleosides, and Nacetylneuraminic acid) could be detected in cancerous tissues. The same method was employed using other matrices (i.e., quercetin for positive and negative ion detection and 9aminoacridine (9AA) for negative ion detection) for the analysis of endogenous compounds in the cancerous and noncancerous regions of human prostate cancer tissues; 152 metabolites, out of the 1091 overall detected and localized, exhibited differential distributions between the two tissue regions [59]. ...
Article
Since its introduction in the 1980s, matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) has gained a prominent role in the analysis of high molecular weight biomolecules such as proteins, peptides, oligonucleotides, and polysaccharides. Its application to low molecular weight compounds has remained for long time challenging due to the spectral interferences produced by conventional organic matrices in the low m/z window. To overcome this problem, specific sample preparation such as analyte/matrix derivatization, addition of dopants, or sophisticated deposition technique especially useful for imaging experiments, have been proposed. Alternative approaches based on second generation (rationally designed) organic matrices, ionic liquids, and inorganic matrices, including metallic nanoparticles, have been the object of intense and continuous research efforts. Definite evidences are now provided that MALDI MS represents a powerful and invaluable analytical tool also for small molecules, including their quantification, thus opening new, exciting applications in metabolomics and imaging mass spectrometry. This review is intended to offer a concise critical overview of the most recent achievements about MALDI matrices capable of specifically address the challenging issue of small molecules analysis.
... The advent of MS-based imaging techniques provides an alternative approach for in situ monitoring the localization of a drug and its metabolites across tissue sections. Mass spectrometry imaging (MSI) technology has been widely used for the acquisition of spatial distribution information of endogenous chemical components in various specimens such as human, animal and plant tissues, as well as microbial samples [7][8][9][10][11]. Among multiple MSI techniques, matrix-assisted laser desorption/ ionization (MALDI) MSI technology is the most popular MSI modality which provides the most comprehensive mass range from small metabolites to macromolecular proteins, high spatial resolution, high-speed and throughput measurement in contrast to 4-7 day exposure time required in acquiring WBA image [12][13][14]. ...
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Detailed spatio-temporal information on drug distribution in organs is of paramount importance to assess drug clinically-relevant properties and potential side-effects. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) as a label-free and sensitive imaging modality provides an additional means of accurately visualizing drug and its metabolites distributions in tissue sections. However, technical limitations, complex physiochemical environment of surface and low abundance of target drugs make quantitative MALDI imaging of drug and its metabolites quite challenging. Methods: In this study, an internal standard correction strategy was applied for quantitative MALDI imaging of tetrandrine in multiple organs of rats including lung, liver, kidney, spleen, and heart. The feasibility and reliability of the developed quantitative MSI method were validated by conventional liquid chromatography-tandem MS (LC-MS/MS) analysis, and the two methods showed a significant correlation. Results: The quantitative MALDI imaging method met the requirements of specificity, sensitivity and linearity. Tissue-specific spatio-temporal distribution patterns of tetrandrine in different organs were revealed after intravenous administration in the rat. Moreover, demethylated metabolite was detected in liver tissues. Conclusions: The current work illustrates that quantitative MALDI imaging provides an alternative means of accurately addressing the problem of drug and its metabolites distribution in tissues, complementary to traditional LC-MS/MS of tissue homogenates and whole-body autoradiography (WBA). Quantitative spatio-chemical information obtained here can improve our understanding of pharmacokinetics (PK), pharmacodynamics (PD), and potential transient toxicities of tetrandrine in organs, and possibly direct further optimization of drug properties to reduce drug-induced organ toxicity.
... In addition to the above mentioned studies, there were two studies with small sample sizes. Wang and colleagues used a MALDI-FTICR-MS approach to profile three matched, intact tissue pairs and identified differential metabolites using student's t-test [50]. However, they did not highlight or evaluate further any of the detected metabolites. ...
Article
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Prostate cancer (PCa) diagnosis with current biomarkers is difficult and often results in unnecessary invasive procedures as well as over-diagnosis and over-treatment, highlighting the need for novel biomarkers. The aim of this review is to provide a summary of available metabolomics PCa biomarkers, particularly for clinically significant disease. A systematic search was conducted on PubMed for publications from July 2008 to July 2018 in accordance with PRISMA guidelines to report biomarkers with respect to their application in PCa diagnosis, progression, aggressiveness, recurrence, and treatment response. The vast majority of studies report biomarkers with the ability to distinguish malignant from benign prostate tissue with a few studies investigating biomarkers associated with disease progression, treatment response or tumour recurrence. In general, these studies report high dimensional datasets and the number of analysed metabolites often significantly exceeded the number of available samples. Hence, observed multivariate differences between case and control samples in the datasets might potentially also be associated with pre-analytical, technical, statistical and confounding factors. Giving the technical and methodological hurdles, there are nevertheless a number of metabolites and pathways repeatedly reported across various technical approaches, cohorts and sample types that appear to play a predominant role in PCa tumour biology, progression and recurrence.
... So far no in situ method is suitable for the detection of AMP, this is a domain of mass spectrometry, more specifically mass spectrometry imaging. Previous studies showed the feasibility for measuring AMP by MALDI imaging in different kind of tissues [28][29][30][31]. Buck et al. [8] verified the detection of AMP in FFPE tissues by MSMS experiments. ...
Article
Multimodal tissue analyses that combine two or more detection technologies provide synergistic value compared to single methods and are employed increasingly in the field of tissue-based diagnostics and research. Here, we report a technical pipeline that describes a combined approach of HER2/CEP17 fluorescence in situ hybridization (FISH) analysis with MALDI imaging on the very same section of formalin-fixed and paraffin-embedded (FFPE) tissue. FFPE biopsies and a tissue microarray of human gastroesophageal adenocarcinoma were analyzed by MALDI imaging. Subsequently, the very same section was hybridized by HER2/CEP17 FISH. We found that tissue morphology of both, the biopsies and the tissue microarray, was unaffected by MALDI imaging and the HER2 and CEP17 FISH signals were analyzable. In comparison with FISH analysis of samples without MALDI imaging, we observed no difference in terms of fluorescence signal intensity and gene copy number. Our combined approach revealed adenosine monophosphate, measured by MALDI imaging, as a prognostic marker. HER2 amplification, which was detected by FISH, is a stratifier between good and poor patient prognosis. By integrating both stratification parameters on the basis of our combined approach, we were able to strikingly improve the prognostic effect. Combining molecules detected by MALDI imaging with the gene copy number detected by HER2/CEP17 FISH, we found a synergistic effect, which enhances patient prognosis. This study shows that our combined approach allows the detection of genetic and metabolic properties from one very same FFPE tissue section, which are specific for HER2 and hence suitable for prognosis. Furthermore, this synergism might be useful for response prediction in tumors.
... Another common strategy in MSI data analysis is to manually define the ROIs to be compared, guided by an annotated histology image [15][16][17][18]. In general, the ions are selected by means of statistical hypothesis testing and the fold change (FC) calculation of the ion concentrations between ROIs. ...
Article
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... Matrix-assisted laser desorption/ionisation-mass spectrometry imaging (MALDI-MSI) is a powerful method for visualising the spatial distributions of lipids throughout biological tissues [1][2][3]. Its versatility for mapping changing lipid compositions within tissues has been demonstrated in a variety of applications, including oncology [4][5][6], bacterial infections [7,8] and liver disease [9,10], amongst many others. The heterogeneous lipid compositions observed with MSI are ultimately the result of cellular-level lipid metabolism occurring within the cells constituting the tissue sample. ...
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Matrix-assisted laser desorption/ionisation-mass spectrometry imaging (MALDI-MSI) is a powerful technique for visualising the spatial locations of lipids in biological tissues. However, a major challenge in interpreting the biological significance of local lipid compositions and distributions detected using MALDI-MSI is the difficulty in associating spectra with cellular lipid metabolism within the tissue. By-and-large this is due to the typically limited spatial resolution of MALDI-MSI (30–100 μm) meaning individual spectra represent the average spectrum acquired from multiple adjacent cells, each potentially possessing a unique lipid composition and biological function. The use of oversampling is one promising approach to decrease the sampling area and improve the spatial resolution in MALDI-MSI, but it can suffer from a dramatically decreased sensitivity. In this work we overcome these challenges through the coupling of oversampling MALDI-MSI with laser post-ionisation (MALDI-2). We demonstrate the ability to acquire rich lipid spectra from pixels as small as 6 μm, equivalent to or smaller than the size of typical mammalian cells. Coupled with an approach for automated lipid identification, it is shown that MALDI-2 combined with oversampling at 6 μm pixel size can detect up to three times more lipids and many more lipid classes than even conventional MALDI at 20 μm resolution in the positive-ion mode. Applying this to mouse kidney and human brain tissue containing active multiple sclerosis lesions, where 74 and 147 unique lipids are identified, respectively, the localisation of lipid signals to individual tubuli within the kidney and lipid droplets with lesion-specific macrophages is demonstrated.
... Interestingly, the high intensity detection of specific CL species correlated with more severe phenotypes of tumour regions, suggesting CL as a promising biomarker for prostate cancer [69,70]. The technique has also been combined with time-of-flight mass spectrometry (MALDI-TOF-MSI) to measure CL [71]. ...
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Cardiolipin (CL) is a mitochondria-exclusive phospholipid, primarily localised within the inner mitochondrial membrane, that plays an essential role in mitochondrial architecture and function. Aberrant CL content, structure, and localisation have all been linked to impaired mitochondrial activity and are observed in the pathophysiology of cancer and neurological, cardiovascular, and metabolic disorders. The detection, quantification, and localisation of CL species is a valuable tool to investigate mitochondrial dysfunction and the pathophysiological mechanisms underpinning several human disorders. CL is measured using liquid chromatography, usually combined with mass spectrometry, mass spectrometry imaging, shotgun lipidomics, fluorometry, and radiolabelling. This review summarises available methods to analyse CL, with a particular focus on modern mass spectrometry, and evaluates their advantages and limitations. We provide guidance aimed at selecting the most appropriate technique, or combination of techniques, when analysing CL in different model systems, and highlight the clinical contexts in which measuring CL is relevant.
... In the same way, spatial analysis of esophageal cancer showed that metabolite profile can be extremely variable within the cancerous tissue: in particular, metabolites such as amino acids, fatty acids, and nucleosides were found to be unevenly distributed in the the tumoral milieu [114]. Heterogenous metabolite distribution within single tumors was also confirmed in colorectal [115], gastric [116], prostate [117], and papillary thyroid cancers [118]. ...
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Metastasis formation accounts for the majority of tumor-associated deaths and consists of different steps, each of them being characterized by a distinctive adaptive phenotype of the cancer cells. Metabolic reprogramming represents one of the main adaptive phenotypes exploited by cancer cells during all the main steps of tumor and metastatic progression. In particular, the metabolism of cancer cells evolves profoundly through all the main phases of metastasis formation, namely the metastatic dissemination, the metastatic colonization of distant organs, the metastatic dormancy, and ultimately the outgrowth into macroscopic lesions. However, the metabolic reprogramming of metastasizing cancer cells has only recently become the subject of intense study. From a clinical point of view, the latter steps of the metastatic process are very important, because patients often undergo surgical removal of the primary tumor when cancer cells have already left the primary tumor site, even though distant metastases are not clinically detectable yet. In this scenario, to precisely elucidate if and how metabolic reprogramming drives acquisition of cancer-specific adaptive phenotypes might pave the way to new therapeutic strategies by combining chemotherapy with metabolic drugs for better cancer eradication. In this review we discuss the latest evidence that claim the importance of metabolic adaptation for cancer progression.
... Unlike MALDI-MS, MALDI-MSI can be treated statistically, making the identification of biomarkers even easier. Having that in mind, it is clear that MALDI-MSI is a valuable analytical platform in lipidomics [93], as well as in general metabolomics [116]. In many studies, thousands of different metabolites were identified in cancer species, and a few hundreds of them were found to be associated with prostate cancer. ...
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Altered lipid metabolism has been associated with the progression of various cancers, and aberrant expression of enzymes involved in the lipid metabolism has been detected in different stages of cancer. Breast cancer (BC) is one of the cancer types known to be associated with alterations in the lipid metabolism and overexpression of enzymes involved in this metabolism. It has been demonstrated that inhibition of the activity of certain enzymes, such as that of phospholipase A2 in BC cell lines sensitizes these cells and decreases the IC50 values for forthcoming therapy with traditional drugs, such as doxorubicin and tamoxifen. Moreover, other phospholipases, such as phospholipase C and D, are involved in intracellular signal transduction, which emphasizes their importance in cancer development. Finally, BC is assumed to be dependent on the diet and the composition of lipids in nutrients. Despite their importance, analytical approaches that can associate the activity of phospholipases with changes in the lipid composition and distribution in cancer tissues are not yet standardized. In this review, an overview of various analytical platforms that are applied on the study of lipids and phospholipase activity in BC tissues will be given, as well as their association with cancer diagnosis and tumor progression. The methods that are applied to tissues obtained from the BC patients will be emphasized and critically evaluated, regarding their applicability in oncology.
... Therefore, mass spectrometry imaging (MSI), a technique able to spatially resolve a wide range of molecules directly in tissue, has become a rapidly growing methodological approach to analyze a range of diseases [2][3][4][5][6][7], including prostate cancer [8][9][10][11]. Matrix-assisted laser desorption/ionization (MALDI) is the most common MSI method for spatial detection of not only proteins [2, 8-10, 12, 13] and peptides [14], but also for glycans [15,16], metabolites [11,17], lipids [11,18] and recently also for metal elements [3,4]. ...
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MALDI MS imaging (MSI) is a powerful analytical tool for spatial peptide detection in heterogeneous tissues. Proper sample preparation is crucial to achieve high quality, reproducible measurements. Here we developed an optimized protocol for spatially resolved proteolytic peptide detection with MALDI time‐of‐flight MSI of fresh frozen prostate tissue sections. The parameters tested included four different tissue washes, four methods of protein denaturation, four methods of trypsin digestion (different trypsin densities, sprayers, and incubation times), and five matrix deposition methods (different sprayers, settings, and matrix concentrations). Evaluation criteria were the number of detected and excluded peaks, percentage of high mass peaks, signal‐to‐noise ratio, spatial localization, and average intensities of identified peptides, all of which were integrated into a weighted quality evaluation scoring system. Based on these scores, the optimized protocol included an ice‐cold EtOH+H2O wash, a 5 min heating step at 95°C, tryptic digestion incubated for 17h at 37°C and CHCA matrix deposited at a final amount of 1.8 μg/mm2. Including a heat‐induced protein denaturation step after tissue wash is a new methodological approach that could be useful also for other tissue types. This optimized protocol for spatial peptide detection using MALDI MSI facilitates future biomarker discovery in prostate cancer and may be useful in studies of other tissue types. This article is protected by copyright. All rights reserved
... Yin et al., 2016;Dhorabe et al., 2016;Wang et al., 2017b;Hareesh et al., 2016).(Table S2) To verify the inhibition effect of surfactant on the catalytic property, polyvinylpyrrolidone (PVP) was ...
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This work reports a new kind of self-assembled PdCu monolithic aerogels via a mild reduction process, which exhibits highly efficient catalytic reduction activity towards 4-nitrophenol. The enhanced catalytic reduction performance can be contributed the following unique features of PdCu aerogels: 1) the interconnected channels and three-dimensional network provide a platform for accelerating mass transfer during catalysis; 2) metallic aerogels combined with stretching ultrathin nanowires has a large surface area and good crystallinity affording sufficient reactive sites and high atom utilization; 3) the introduction of nonprecious Cu not only drastically cuts down the cost but also attains the excellent catalytic activity due to the bimetallic intrinsic synergetic effect; 4) the self-supporting feature is good for improving the durability of the catalyst. This study pushes a new avenue to develop robust catalysts for heterogeneous catalytic reactions.
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Mass spectrometry imaging (MSI) and mass spectrometry (MS) profiling are two especially suitable methods for recording and visualizing microscopic molecular details. Mass spectrometry imaging of larger molecules, such as proteins and intact peptides, was demonstrated using a gentler imaging technique, termed matrix‐assisted laser/desorption ionization (MALDI). This chapter presents an overview of MSI and MS profiling instrumentation, sample preparation, and data analysis, with a focus on peptide and protein detection. It provides specific examples of MSI and MS profiling in biomarker discovery. Biomarkers are commonly considered to be indicators of a disease state or a therapeutic response. They have proven useful in many fields of research, including fundamental neuroscience, microbiology, and plant biology. Matrix‐assisted laser/desorption ionization‐MSI and MS profiling are valuable tools for characterizing chemical signaling in animal nervous systems, particularly for signaling peptides, which can be both detected and sequenced on tissue.
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Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) is an emerging analytical technique that promises to change tissue-based diagnostics. This article provides a brief introduction to MALDI MSI as well as clinical diagnostic workflows and opportunities to apply this powerful approach. It describes various MALDI MSI applications, from more clinically mature applications such as cancer to emerging applications such as infectious diseases and drug distribution. In addition, it discusses the analytical considerations that need to be considered when bringing these approaches to different diagnostic problems and settings.
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This review is the tenth update of the original article published in 1999 on the application of matrix‐assisted laser desorption/ionization mass spectrometry (MALDI) mass spectrometry to the analysis of carbohydrates and glycoconjugates and brings coverage of the literature to the end of 2018. Also included are papers that describe methods appropriate to glycan and glycoprotein analysis by MALDI, such as sample preparation techniques, even though the ionization method is not MALDI. Topics covered in the first part of the review include general aspects such as theory of the MALDI process, new methods, matrices, derivatization, MALDI imaging, fragmentation and the use of arrays. The second part of the review is devoted to applications to various structural types such as oligo‐ and poly‐saccharides, glycoproteins, glycolipids, glycosides, and biopharmaceuticals. Most of the applications are presented in tabular form. The third part of the review covers medical and industrial applications of the technique, studies of enzyme reactions, and applications to chemical synthesis. The reported work shows increasing use of combined new techniques such as ion mobility and highlights the impact that MALDI imaging is having across a range of diciplines. MALDI is still an ideal technique for carbohydrate analysis and advancements in the technique and the range of applications continue steady progress.
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Introduction Mass spectrometry imaging (MSI) as a label‐free and powerful imaging technique enables in situ evaluation of a tissue metabolome and/or proteome, becoming increasingly popular in the detection of plant endogenous molecules. Objective The characterization of structure and spatial information of endogenous molecules in plants are both very important aspects to better understand the physiological mechanism of plant organism. Methods Matrix‐assisted laser desorption/ionization mass spectrometry imaging (MALDI‐MSI) is a commonly‐used tissue imaging technique, which requires matrix to assist in situ detection of a variety of molecules on the surface of a tissue section. In previous studies, MALDI‐MSI was mostly used for the detection of molecules from animal tissue sections, compared to plant samples due to cell structural limitations, such as plant cuticles, epicuticular waxes, and cell walls. Despite the enormous progress that has been made in tissue imaging, there is still a challenge for MALDI‐MSI suitable for the imaging of endogenous compounds in plants. Results This review summarises the recent advances in MALDI‐MSI, focusing on the application of in situ detection of endogenous molecules in different plant organs, i.e. root, stem, leaf, flower, fruit, and seed. Conclusion Further improvements on instrumentation sensitivity, matrix selection, image processing and sample preparation will expand the application of MALDI‐MSI in plant research.
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Staging of prostate carcinoma provides a standardized method for tumor classification which is based on involvement of the prostate gland, adjacent local structures, regional lymph nodes, and distant sites. Staging information is therefore crucial for clinicians to be able to assess risk of disease progression, to offer therapeutic choices in the individual patient, and to provide population-based prognostic information. Clinical staging, which is based on data obtained prior to first definitive treatment, relies on tumor determination by digital rectal examination, transrectal ultrasonography, other imaging techniques, and serum PSA level, while pathological staging requires histological identification of tumor extent in prostate gland and surrounding tissues. T1 tumors, denoted to clinically unapparent, not palpable or visible by imaging, are diagnosed by transurethral resection of the prostate procedure or needle biopsy. T2 tumors are confined to the organ, are subdivided by involvement in one or both lobes, and are determined by radical prostatectomy procedure. Stage T3 denotes locally advanced tumors that spread beyond the organ's boundaries, and T4 denotes invasion or fixation to the pelvic organs. Despite wide acceptance of the system as a whole, the current 2010 revision of the American Joint Committee on Cancer/Union for International Cancer Control tumor, node and metastasis (TNM 7) appears to contain some controversies, particularly T2 three-tiered subclassification. This review will cover suggested changes to further TNM editions; these changes have been accumulated in the literature in recent years and include items such as lymph node involvement quantification, "vanishing" carcinoma, Gleason score, resection margin status, pretreatment serum PSA level, as well as difficulties the pathologist may encounter in microscopic examination which may hamper accurate stage assessment.
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The availability of human genome sequence has transformed biomedical research over the past decade. However, an equivalent map for the human proteome with direct measurements of proteins and peptides does not exist yet. Here we present a draft map of the human proteome using high-resolution Fourier-transform mass spectrometry. In-depth proteomic profiling of 30 histologically normal human samples, including 17 adult tissues, 7 fetal tissues and 6 purified primary haematopoietic cells, resulted in identification of proteins encoded by 17,294 genes accounting for approximately 84% of the total annotated protein-coding genes in humans. A unique and comprehensive strategy for proteogenomic analysis enabled us to discover a number of novel protein-coding regions, which includes translated pseudogenes, non-coding RNAs and upstream open reading frames. This large human proteome catalogue (available as an interactive web-based resource at http://www.humanproteomemap.org) will complement available human genome and transcriptome data to accelerate biomedical research in health and disease.
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High-resolution matrix-assisted laser desorption/ionization imaging mass spectrometry (HR-MALDI-IMS) is an emerging application for the comprehensive and detailed analysis of the spatial distribution of ionized molecules in situ on tissue slides. HR-MALDI-IMS in negative mode in a mass range of m/z 500-1000 was performed on optimal cutting temperature (OCT) compound-embedded human prostate tissue samples obtained from patients with prostate cancer at the time of radical prostatectomy. HR-MALDI-IMS analysis of the 14 samples in the discovery set identified 26 molecules as highly expressed in the prostate. Tandem mass spectrometry (MS/MS) showed that these molecules included 14 phosphatidylinositols (PIs), 3 phosphatidylethanolamines (PEs) and 3 phosphatidic acids (PAs). Among the PIs, the expression of PI(18:0/18:1), PI(18:0/20:3) and PI(18:0/20:2) were significantly higher in cancer tissue than in benign epithelium. A biomarker algorithm for prostate cancer was formulated by analyzing the expression profiles of PIs in cancer tissue and benign epithelium of the discovery set using orthogonal partial least squares discriminant analysis (OPLS-DA). The sensitivity and specificity of this algorithm for prostate cancer diagnosis in the 24 validation set samples were 87.5 and 91.7%, respectively. In conclusion, HR-MALDI-IMS identified several PIs as being more highly expressed in prostate cancer than benign prostate epithelium. These differences in PI expression profiles may serve as a novel diagnostic tool for prostate cancer.
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Cancer incidence and mortality estimates for 25 cancers are presented for the 40 countries in the four United Nations-defined areas of Europe and for the European Union (EU-27) for 2012. We used statistical models to estimate national incidence and mortality rates in 2012 from recently-published data, predicting incidence and mortality rates for the year 2012 from recent trends, wherever possible. The estimated rates in 2012 were applied to the corresponding population estimates to obtain the estimated numbers of new cancer cases and deaths in Europe in 2012. There were an estimated 3.45 million new cases of cancer (excluding non-melanoma skin cancer) and 1.75 million deaths from cancer in Europe in 2012. The most common cancer sites were cancers of the female breast (464,000 cases), followed by colorectal (447,000), prostate (417,000) and lung (410,000). These four cancers represent half of the overall burden of cancer in Europe. The most common causes of death from cancer were cancers of the lung (353,000 deaths), colorectal (215,000), breast (131,000) and stomach (107,000). In the European Union, the estimated numbers of new cases of cancer were approximately 1.4 million in males and 1.2 million in females, and around 707,000 men and 555,000 women died from cancer in the same year. These up-to-date estimates of the cancer burden in Europe alongside the description of the varying distribution of common cancers at both the regional and country level provide a basis for establishing priorities to cancer control actions in Europe. The important role of cancer registries in disease surveillance and in planning and evaluating national cancer plans is becoming increasingly recognised, but needs to be further advocated. The estimates and software tools for further analysis (EUCAN 2012) are available online as part of the European Cancer Observatory (ECO) (http://eco.iarc.fr). Copyright © 2013 Elsevier Ltd. All rights reserved.
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To identify molecular markers of pathologic response to neoadjuvant paclitaxel/radiation treatment, protein and gene expression profiling were done on pretreatment biopsies. Patients with high-risk, operable breast cancer were treated with three cycles of paclitaxel followed by concurrent paclitaxel/radiation. Tumor tissue from pretreatment biopsies was obtained from 19 of the 38 patients enrolled in the study. Protein and gene expression profiling were done on serial sections of the biopsies from patients that achieved a pathologic complete response (pCR) and compared to those with residual disease, non-pCR (NR). Proteomic and validation immunohistochemical analyses revealed that alpha-defensins (DEFA) were overexpressed in tumors from patients with a pCR. Gene expression analysis revealed that MAP2, a microtubule-associated protein, had significantly higher levels of expression in patients achieving a pCR. Elevation of MAP2 in breast cancer cell lines led to increased paclitaxel sensitivity. Furthermore, expression of genes that are associated with the basal-like, triple-negative phenotype were enriched in tumors from patients with a pCR. Analysis of a larger panel of tumors from patients receiving presurgical taxane-based treatment showed that DEFA and MAP2 expression as well as histologic features of inflammation were all statistically associated with response to therapy at the time of surgery. We show the utility of molecular profiling of pretreatment biopsies to discover markers of response. Our results suggest the potential use of immune signaling molecules such as DEFA as well as MAP2, a microtubule-associated protein, as tumor markers that associate with response to neoadjuvant taxane-based therapy.
Article
Since the benefit of prostate-specific antigen (PSA) screening remains controversial, new non-invasive biomarkers for prostate carcinoma (PCa) are still required. There is evidence that microRNAs (miRNAs) in whole peripheral blood can separate patients with localized prostate cancer from healthy individuals. However, the potential of blood-based miRNAs for the differential diagnosis of PCa and benign prostatic hyperplasia (BPH) has not been tested. We compared the miRNome from blood of PCa and BPH patients and further investigated the influence of the tumor volume, tumor-node-metastasis (TNM) classification, Gleason score, pretreatment risk status, and the pretreatment PSA value on the miRNA pattern. By microarray approach, we identified seven miRNAs that were significantly deregulated in PCa patients compared to BPH patients. Using quantitative real time PCR (qRT-PCR), we confirmed downregulation of hsa-miR-221* (now hsa-miR-221-5p) and hsa-miR-708* (now hsa-miR-708-3p) in PCa compared to BPH. Clinical parameters like PSA level, Gleason score, or TNM status seem to have only limited impact on the overall abundance of miRNAs in patients' blood, suggesting a no influence of these factors on the expression of deregulated miRNAs.
Article
Since 1966, when Donald Gleason first proposed grading prostate cancer based on its histologic architecture, there have been numerous changes in clinical and pathologic practices relating to prostate cancer. Patterns 1 and 2, comprising more than 30% of cases in the original publications by Gleason, are no longer reported on biopsy and are rarely diagnosed on radical prostatectomy. Many of these cases may even have been mimickers of prostate cancer that were described later with the use of contemporary immunohistochemistry. The original Gleason system predated many newly described variants of prostate cancer and our current concept of intraductal carcinoma. Gleason also did not describe how to report prostate cancer on biopsy with multiple cores of cancer or on radical prostatectomy with separate tumor nodules. To address these issues, the International Society of Urological Pathology first made revisions to the grading system in 2005, and subsequently in 2014. Additionally, a new grading system composed of Grade Groups 1 to 5 that was first developed in 2013 at the Johns Hopkins Hospital and subsequently validated in a large multi-institutional and multimodal study, was presented at the 2014 meeting and accepted both by participating pathologists as well as urologists, oncologists, and radiation therapists. In the present study, we describe updates to the grading of prostate cancer along with the new grading system.
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Idiopathic glomerulonephritis (GN), such as membranous glomerulonephritis (MGN), focal segmental glomerulosclerosis (FSGS) and IgA nephropathy (IgAN), represent the most frequent primary Glomerular Kidney Diseases (GKDs) worldwide. Although the renal biopsy currently remains the gold standard for the routine diagnosis of idiopathic GN, the invasiveness and diagnostic difficulty related with this procedure highlight the strong need for new diagnostic and prognostic biomarkers to be translated into less invasive diagnostic tools. Matrix Assisted Laser Desorption/Ionisation Mass Spectrometry Imaging (MALDI-MSI) was applied to fresh frozen bioptic renal tissue from patients with a histological diagnosis of FSGS (n = 6), IgAN, (n = 6) and MGN (n = 7), and from controls (n = 4) in order to detect specific molecular signatures of primary glomerulonephritis. MALDI-MSI was able to generate molecular signatures capable to distinguish between normal kidney and pathological GN, with specific signals (m/z 4025, 4048 and 4963) representing potential indicators of CKD development. Moreover, specific disease-related signatures (m/z 4025 and 4048 for FSGS, m/z 4963 and 5072 for IgAN) were detected. Of these signals, m/z 4048 was identified as α-1-antitrypsin (A1AT) and was shown to be localised to the podocytes within sclerotic glomeruli by immunohistochemistry. A1AT could be one of the markers of podocyte stress that is correlated with the development of FSGS due to both an excessive lossand a hypertrophy of podocytes. This article is protected by copyright. All rights reserved.
Article
In this review, we discuss whether novel techniques in mass spectrometry, from ultrahigh resolution detection to data-independent MS/MS and ion mobility methods, have advanced so far that selectivity and sensitivity of untargeted analyses have indeed reached a point at which hypothesis-driven validation studies can be conducted by accurate mass profiling methods rather than classic triple-quadrupole multiple-reaction monitoring. To this end, we reviewed original MS-based metabolomics and lipidomics papers published mainly in years 2012–2015 with focus on sample extraction, LC separation, MS detection, and data processing.
Article
BACKGROUND Prostate needle biopsy (PNB) is required for the diagnosis of prostate cancer (PCa), but little is known about the frequency and clinical implication of false-negative results. OBJECTIVE To investigate the incidence and clinical impact of minute PCa missed on routine haematoxylin and eosin (H&E) slides, but retrieved by -methylacyl-CoA-racemase (AMACR) immunohistochemistry. METHODSAMACR immunohistochemistry was used to detect PCa missed on H&E slides in a series of consecutive 1,672 PNB including 1,003 patients without evidence of PCa, and 669 patients with PCa meeting pathological criteria for active surveillance (PCAS) under current clinical investigation, including Gleason score 7 (3+4), <33% of biopsies involved by cancer, <50% of any core involved by cancer. Using improved multicore (pre-) embedding techniques a single AMACR immunostain/patient was sufficient to detect missed lesions. RESULTSIn patients without histological evidence of PCa, AMACR immunohistochemistry retrieved minute PCa in 33 of 1,003 patients (3.29%) and atypical small acinar proliferations (ASAP) in 17 of 1,003 patients (1.69%). Among 116 of 669 (17.34%) PCa patients meeting PCAS, detection of additional core(s) involved by cancer was found responsible for disease reclassification in 63 of 116 of patients (54.31%). Limitations include the single-institutional design of the study. CONCLUSIONSPCa missed on routine H&E histology was retrieved by AMACR in 8.91% of PNB, including 17.34% of PCa patients meeting PCAS. 54.31% of them have finally lost their eligibility for active surveillance after detecting additional cores involved by cancer. Underdiagnosis of limited adenocarcinoma on PNB is a matter of concern, but can be prevented by a single AMACR immunostain/patient if improved multicore (pre-) embedding techniques are used. Prostate 76:369-375, 2016. (c) 2015 Wiley Periodicals, Inc.
Article
In this work, we combined a newly developed matrix coating technique -- matrix coating assisted by an electric field (MCAEF) and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to enhance the imaging of peptides and proteins in tissue specimens of human prostate cancer. MCAEF increased the signal-to-noise ratios of the detected proteins by a factor of 2 to 5 and 232 signals were detected within the m/z 3500-37500 mass range on a time-of-flight mass spectrometer and with the sinapinic acid MALDI matrix. Among these species, 3 proteins (S100-A9, S100-A10 and S100-A12) were only observed in the cancerous cell region and 14 proteins, including a fragment of mitogenactivated protein kinase/extracellular signal-regulated kinase kinase kinase 2, a fragment of cAMPregulated phosphoprotein 19, 3 apolipoproteins (C-I, A-I, and A-II), 2 S100 proteins (A6 and A8), β-microseminoprotein, tumor protein D52, α-1-acid glycoprotein 1, heat shock protein β-1, prostate-specific antigen, and 2 unidentified large peptides at m/z 5002.2 and 6704.2, showed significantly differential distributions at the p<0.05 (t-test) level between the cancerous and the non-cancerous regions of the tissue. Among these 17 species, the distributions of apolipoprotein C-I, S100-A6, and S100-A8 were verified by immunohistological staining. In summary, this study resulted in the imaging of the largest group of proteins in prostate cancer tissues by MALDI-MS reported thus far, and is the first to show a correlation between S100 proteins and prostate cancer in a MS imaging study. The successful imaging of the three proteins only found in the cancerous tissues, as well as those showing differential expressions demonstrated the potential of MCAEF-MALDI/MS for the in situ detection of potential cancer biomarkers.
Article
Background Human prostate cancers are highly heterogeneous, indicating a need for various novel biomarkers to predict their prognosis. Lipid metabolism affects numerous cellular processes, including cell growth, proliferation, differentiation, and motility. Direct profiling of lipids in tissue using high-resolution matrix-assisted laser desorption/ionization imaging mass spectrometry (HR-MALDI-IMS) may provide molecular details that supplement tissue morphology.Methods Prostate tissue samples were obtained from 31 patients, with localized prostate cancer who underwent radical prostatectomy. The samples were assessed by HR-MALDI-IMS in positive mode, with the molecules identified by tandem mass spectrometry (MS/MS). The effect of identified molecules on prostate specific antigen recurrence free survival after radical prostatectomy was determined by Cox regression analysis and by the Kaplan–Meier method.ResultsThirteen molecules were found to be highly expressed in prostate tissue, with five being significantly lower in cancer tissue than in benign epithelium. MS/MS showed that these molecules were [lysophosphatidylcholine (LPC)(16:0/OH)+H]+, [LPC(16:0/OH)+Na]+, [LPC(16:0/OH)+K]+, [LPC(16:0/OH)+matrix+H]+, and [sphingomyelin (SM)(d18:1/16:0)+H]+. Reduced expression of LPC(16:0/OH) in cancer tissue was an independent predictor of biochemical recurrence after radical prostatectomy.ConclusionsHR-MALDI-IMS showed that the expression of LPC(16:0/OH) and SM(d18:1/16:0) was lower in prostate cancer than in benign prostate epithelium. These differences in expression of phospholipids may predict prostate cancer aggressiveness, and provide new insights into lipid metabolism in prostate cancer. Prostate © 2015 Wiley Periodicals, Inc.
Article
Significance The absence of appropriate transgenic animal models of renal cell carcinomas (RCCs) has made it difficult to identify and test new therapies for this disease. We developed a new transgenic mouse model of a highly aggressive form of RCC in which tumor growth and regression is conditionally regulated by the MYC oncogene. Using desorption electrospray ionization–mass-spectrometric imaging, we found that certain glycerophosphoglycerols and metabolites of the glutaminolytic pathway were higher in abundance in RCC than in normal kidney tissue. Up-regulation of glutaminolytic genes and proteins was identified by genetic analysis and immunohistochemistry, therefore suggesting that RCC tumors are glutamine addicted. Pharmacological inhibition of glutaminase slowed tumor progression in vivo, which may represent a novel therapeutic route for RCC.
Article
Background: To determine whether prostate-specific antigen (PSA) testing in African American veterans (AAVs) aged 40 to 54 years is associated with high-risk prostate cancer characteristics compared with AAVs aged 55 to 70 years or white veterans (WVs) aged 40 to 54 years. Methods: A total of 231,174 healthy veterans aged 40 to 70 years without clinical evidence of prostate cancer underwent PSA testing between October 1, 2000, and September 30, 2007. Clinicopathologic tumor characteristics were available for 1,044/1,059 AAVs and 1,006/1,971 age-matched WVs diagnosed with prostate cancer after a PSA level>4 ng/ml triggered prostate biopsy. Tumor characteristics of AAVs aged 40 to 54 years were compared with AAVs 55 to 70 years, WVs 40 to 54 years, and WVs 55 to 70 years. Results: Of PSA-tested veterans aged 40 to 54 years diagnosed with prostate cancer, there were no racial differences in prebiopsy PSA levels, prostate cancer grade, or clinical stage at diagnosis. AAVs aged 40 to 54 years were more likely to have ≥ 3 positive cores (P = 0.0229) and were less likely to be active surveillance candidates (P = 0.0340) compared with similarly aged WVs. AAVs aged 55 to 70 years were more likely to have high-grade (P = 0.0204) and higher clinical stage (P = 0.0195) prostate cancer than AAVs aged 40 to 54 years. Conclusions: This large national cohort study suggests that PSA testing at an earlier age for African American men may allow diagnosis of lower risk prostate cancer, potentially reducing disparate outcomes between AAVs and WVs.
Article
Prostate-specific antigen (PSA) has revolutionized the diagnosis and management of men with prostate cancer. Significant advances have been made since the early development of immunoassays. While PSA is useful for staging and monitoring of established disease, it has shown the greatest utility in the realm of early detection realm. PSA is the most important tumor marker; its importance in evaluating men for the possibility of prostate cancer is irrefutable. Enhancing specificity is a pressing need. In this regard, the recognition of the molecular forms of free PSA and complex PSA have shown the most promise and undoubtedly will result in fewer false-positive PSA test results. The salient literature is reviewed and commentary made on the current status of PSA with particular emphasis on methods to enhance its specificity in early detection and applications. Semin. Surg. Oncol. 18:3–9, 2000. © 2000 Wiley-Liss, Inc.
Article
Proteomes are characterized by large protein-abundance differences, cell-type- and time-dependent expression patterns and post-translational modifications, all of which carry biological information that is not accessible by genomics or transcriptomics. Here we present a mass-spectrometry-based draft of the human proteome and a public, high-performance, in-memory database for real-time analysis of terabytes of big data, called ProteomicsDB. The information assembled from human tissues, cell lines and body fluids enabled estimation of the size of the protein-coding genome, and identified organ-specific proteins and a large number of translated lincRNAs (long intergenic non-coding RNAs). Analysis of messenger RNA and protein-expression profiles of human tissues revealed conserved control of protein abundance, and integration of drug-sensitivity data enabled the identification of proteins predicting resistance or sensitivity. The proteome profiles also hold considerable promise for analysing the composition and stoichiometry of protein complexes. ProteomicsDB thus enables navigation of proteomes, provides biological insight and fosters the development of proteomic technology.
Article
Adrenal glands synthesize and release functional zone-specific steroid and catecholamine hormones to regulate mammalian stress responses. Lipids such as sphingolipids have been shown to control the steroid hormone biosynthesis in adrenal glands, indicating their important roles in endocrine organs. Molecular imaging by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a well-established analytical technique for determining both the spatial location and the relative abundances of various lipids on tissue. To better understand the overall roles of different lipid classes that play in the mammalian adrenal glands, it is necessary to comprehensively determine the spatial distributions of various lipids in the different functional zones of adrenal glands. However, the potential of this technique has not been fully reached considering there are thousands of lipid species in a cell or tissue. To achieve this, we used quercetin as a MALDI matrix for negative ion detection of endogenous lipids on tissue sections of porcine adrenal glands by MALDI-Fourier-transform ion cyclotron resonance (FTICR) MS. As a result of these experiments, 409 endogenous compounds were detected in the negative ion mode. Combining both the positive and negative ion detection led to successful determination of the spatial distribution patterns of 555 unique endogenous compounds that were identified as 544 lipid entities and 11 non-lipid metabolites. Many classes of these lipids showed distinct distribution patterns in different functional zones of the adrenal gland. To the best of our knowledge, this work presents the largest group of lipid entities that have been analyzed in a single MS imaging study so far, and comprehensive profiles of the spatial distributions of lipids in porcine adrenal glands are shown here for the first time.
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The sphingolipid metabolites have emerged as a starting point for the development of novel therapeutics for many diseases. However, details of the functions and mechanisms of sphingolipids remain unknown. To better understand the roles of sphingolipids, chemical tools with unique biological and physicochemical properties are needed. In this regard, we previously reported the synthesis of sphingoid base analogues in which the carbon chains are restricted by triple bonds. Here, we have conjugated a fluorescent dye to the polyyne analogues of the sphingoid bases to generate optical probes. Like the parent polyyne-containing sphingoid base, the 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD)-labeled triyne-sphingosine inhibited cancer cell growth far more effectively than did the corresponding sphingosine. NBD-triyne-sphingosine was rapidly incorporated into the cells and displayed broad cytoplasmic distribution. According to the results of a flow cytometric analysis, cancer cells fed with NBD-triyne-sphingosine showed significantly increased fluorescence intensity compared with the NBD-sphingosine treated cells. The metabolism of NBD-triyne-sphingosine was somewhat different from that of NBD-sphingosine. These results indicated that the incorporated rigid polyyne moiety in the sphingoid base altered the physicochemical properties of the sphingolipid, thereby affecting its biological behavior. The higher antiproliferative activity in the SRB assay and the significantly higher fluorescence intensity observed in the flow cytometric analysis are some of the interesting and distinct aspects of NBD-triyne-sphingosine compared to standard NBD-sphingosine probes. Thus, it is believed that the fluorescently labeled polyyne-containing sphingoid base developed in this study will be a useful chemical tool in sphingolipid research.
Article
The discovery of new matrices that are suitable for in situ analysis of low molecular-weight compounds by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is an important technological aspect of tissue imaging. In this work, ten natural flavonoid compounds, including flavone and nine of its mono- or polyhydroxyl-substituted analogues (3-hydroxyflavone, 5-hydroxyflavone, 3,7-dihydroxyflavone, chrysin, 7,3',4'-trihydroxyflavone, fisetin, luteolin, quercetin, and morin) were evaluated as potential MALDI matrices for the profiling and imaging of endogenous lipids in mouse liver, using a Fourier transform ion cyclotron resonance (FTICR) mass spectrometer with a 355-nm Nd:YAG UV laser, in the positive ion mode. When an electronic sprayer was used for matrix coating and with a high-pH (0.1-0.5% ammonia hydroxide) matrix solvent, eight of the ten compounds, all of which had at least one OH group at the C3 or C5 position of the flavone structure, enabled the successful detection of 77 to 161 phospholipids and other lipids. The best results were observed with two penta-OH flavones (i.e., quercertin and morin). Taking quercetin as an example, this matrix showed characteristics superior to those of commonly used MALDI matrices, such as DHB (2,5-dihydroxybenzoic acid), CHCA (α-cyano-4-hydroxycinnamic acid), and 2-mercaptobenzothiazole (2-MBT). These characteristics were: μm-sized matrix crystals, uniform matrix coating, low volatility in the high vacuum (∼10(-7) mbar) source, good chemical stability, low yield of matrix-related ions, low matrix consumption, low power threshold for laser desorption/ionization, and improved safety of handling. The use of quercetin led to improved lipid imaging, with 212 lipids being successfully imaged from rat brain in a single experiment and with asymmetric distributions of some lipids in left and right brain hippocampus being observed for the first time.
Article
An analysis of tissue specimens by matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) in biological and clinical research is studied. In a profiling mode, data are collected from regions of interest defined by tissue pathology, while in imaging mode, the entire sample surface is raster sampled to reproduce an image of the specimen-based ion-specific ion intensities. The complete data set is comprised of many hundreds or thousands of m/z values, and the intensities of each ion can be plotted in a false color display. The resulting set of ion images is used to accurately portray the spatial distribution of the molecules that comprise the sample. After spotting with matrix, the coordinates of the matrix spots are registered to the mass spectrometer and spectra are automatically acquired from each location. Careful planning, attention to detail, and cleanliness in laboratory practices are required to avoid unwanted loss of sensitivity, loss of spatial resolution, or problems with sample stability.
Article
Delahunt B, Miller R J, Srigley J R, Evans A J & Samaratunga H (2012) Histopathology60, 75–86 Gleason grading: past, present and futureIn 1966 Donald Gleason developed his grading and scoring system for prostatic adenocarcinoma. This classification was refined in 1974 and gained almost universal acceptance, being classified as a category 1 prognostic parameter by the College of American Pathologists. Modifications to the classification were recommended at a conference convened by the International Society of Urological Pathology (ISUP) in 2005. This modified classification has resulted in a significant upgrading of tumours, although some studies have shown a greater concordance between needle biopsy and radical prostatectomy scores when compared to classical Gleason (CG) grading. The ISUP consensus conference recommended that for needle biopsies higher tertiary patterns should be incorporated into the final Gleason score, and this has been correlated with biochemical failure, tumour volume and mortality. Recently the validity of including cribriform glands as a component of Gleason pattern 3 has been questioned and it has been recommended that all tumours showing cribriform architecture should be classified as Gleason pattern 4. The recommendations arising from the 2005 Consensus Conference were largely unsupported by validating data, yet this new grading system has achieved widespread usage. It is unfortunate that recent suggestions for further modification are similarly lacking in supporting evidence. In view of this it is recommended that the Modified Gleason Scoring Classification should continue to be utilized in its original (2005) format and that any future alterations should be implemented only when mandated by tumour-related outcome studies.
Article
Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) has emerged as a novel powerful MS methodology that has the ability to generate both molecular and spatial information within a tissue section. Application of this technology as a new type of biochemical lipid microscopy may lead to new discoveries of the lipid metabolism and biomarkers associated with area-specific alterations or damage under stress/disease conditions such as traumatic brain injury or acute lung injury, among others. However there are limitations in the range of what it can detect as compared with liquid chromatography-MS (LC-MS) of a lipid extract from a tissue section. The goal of the current work was to critically consider remarkable new opportunities along with the limitations and approaches for further improvements of MALDI-MSI. Based on our experimental data and assessments, improvements of the spectral and spatial resolution, sensitivity and specificity towards low abundance species of lipids are proposed. This is followed by a review of the current literature, including methodologies that other laboratories have used to overcome these challenges.
Article
Prostate specific antigen (PSA) is serine protease produced at high concentrations by normal and malignant prostatic epithelium. It is mainly secreted into seminal fluid, where it digests the gel forming after ejaculation. Only minor amounts of PSA leak out into circulation from the normal prostate, but the release of PSA is increased in prostatic disease. Thus PSA is a sensitive serum marker for prostate cancer but its specificity is limited by a high frequency of falsely elevated values in men with benign prostatic hyperplasia (BPH). Approximately two-thirds of all elevated values (>4 μg/l) in men over 50 years of age are due to BPH. In serum, most of the PSA immunoreactivity consists of a complex between PSA and α1-antichymotrypsin (PSA-ACT) whereas approximately 5–40% are free. The proportion of PSA-ACT is larger and the free fraction is smaller in prostate cancer than in benign prostatic hyperplasia (BPH). Determination of the proportion of free PSA has become widely used to improve the cancer specificity of PSA especially in men with PSA values in the `grey zone' (4–10 μg/l). PSA also occurs in complexes with other protease inhibitors and determination of these and other markers may further improve the diagnostic accuracy for prostate cancer. Interpretation of the results for many different markers is complicated, but this can be simplified by using statistical methods. The diagnostic accuracy can be further improved by using logistic regression or neural networks to estimate the combined impact of marker results and other findings like digital rectal examination (DRE), transrectal ultrasound (TRUS) and heredity.
Article
Proteomics-based approaches allow us to investigate the biology of cancer beyond genomic initiatives. We used histology-based matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry to identify proteins that predict disease outcome in gastric cancer after surgical resection. A total of 181 intestinal-type primary resected gastric cancer tissues from two independent patient cohorts were analyzed. Protein profiles of the discovery cohort (n = 63) were directly obtained from tumor tissue sections by MALDI imaging. A seven-protein signature was associated with an unfavorable overall survival independent of major clinical covariates. The prognostic significance of three individual proteins identified (CRIP1, HNP-1, and S100-A6) was validated immunohistochemically on tissue microarrays of an independent validation cohort (n = 118). Whereas HNP-1 and S100-A6 were found to further subdivide early-stage (Union Internationale Contre le Cancer [UICC]-I) and late-stage (UICC II and III) cancer patients into different prognostic groups, CRIP1, a protein previously unknown in gastric cancer, was confirmed as a novel and independent prognostic factor for all patients in the validation cohort. The protein pattern described here serves as a new independent indicator of patient survival complementing the previously known clinical parameters in terms of prognostic relevance. These results show that this tissue-based proteomic approach may provide clinically relevant information that might be beneficial in improving risk stratification for gastric cancer patients.
Article
Several significant reasons for the success of matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) applied to lipid imaging have emerged. The first is the high abundance of various lipids in biological tissues because these hydrophobic molecules constitute the external and internal defining membranes of each cell. The mass defect or fractional mass of the molecular ion species provides information about the elemental composition of the desorbed ion from the tissue. MALDI IMS depicts regional localization of specific lipids in many different types of tissues. The validity of these MALDI images, specifically the relationship between MALDI IMS ion intensity and the amount of the lipid actually present in the tissue, although visually intuitive, is in reality affected by a complex combination of ion chemistry, tissue structure, lipid location within the cells of the tissue, and water and salt content.
Article
Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) has improved over the years and is increasingly being used for biomarker discovery directly from human tissue sections. State-of-the-art technology currently enables a resolution down to 20 µm. MSI therefore allows the correlation of spatial and temporal protein expression profiles with distinct morphological features without requiring target-specific reagents, such as antibodies. Several studies have demonstrated the strength of the technology for uncovering new markers that correlate with disease severity as well as prognosis and therapeutic response. This review provides an overview of MALDI imaging functionality and its advantages and disadvantages, and provides a current literature overview of malignancy-based biomarker detection. Further improvements on instrumentation sensitivity, image processing and sample preparation will enable the detection of novel, tissue-specific biomarkers. However, emphasis should be given to large validation studies and/or subsequent identification of differentially observed protein peaks in order to transfer MSI protein profiling and/or novel biomarkers thereof into clinical use.
Article
Matrix assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) is a method that allows the investigation of the molecular content of tissues within its morphological context. Since it is able to measure the distribution of hundreds of analytes at once, while being label free, this method has great potential which has been increasingly recognized in the field of tissue-based research. In the last few years, MALDI-IMS has been successfully used for the molecular assessment of tissue samples mainly in biomedical research and also in other scientific fields. The present article will give an update on the application of MALDI-IMS in clinical and preclinical research. It will also give an overview of the multitude of technical advancements of this method in recent years. This includes developments in instrumentation, sample preparation, computational data analysis and protein identification. It will also highlight a number of emerging fields for application of MALDI-IMS like drug imaging where MALDI-IMS is used for studying the spatial distribution of drugs in tissues.
Article
We identified information critical for patient treatment on prostate needle biopsies diagnosed with prostatic intraepithelial neoplasia or atypical foci suspicious for carcinoma. A search was performed using the MEDLINE database and referenced lists of relevant studies to obtain articles addressing the significance of finding PIN or atypical foci suspicious for carcinoma on needle biopsy. There were certain results concerning PIN. 1) Low grade PIN should not be documented in pathology reports due to poor interobserver reproducibility and a relatively low risk of cancer following re-biopsy. 2) The expected incidence of HGPIN on needle biopsy is between 5% and 8%. 3) Although the diagnosis of HGPIN is subjective, interobserver reproducibility for its diagnosis is fairly high among urological pathologists, and yet only moderate among pathologists without special expertise in prostate pathology. 4) The median risk recorded in the literature for cancer following the diagnosis of HGPIN on needle biopsy is 24.1%, which is not much higher than the risk reported in the literature for repeat biopsy following a benign diagnosis. 5) The majority of publications that compared the risk of cancer in the same study following a needle biopsy diagnosis of HGPIN to the risk of cancer following a benign diagnosis on needle biopsy show no differences between the 2 groups. 6) Clinical and pathological parameters do not help stratify which men with HGPIN are at increased risk for a cancer diagnosis. 7) A major factor contributing to the decreased incidence of cancer following a diagnosis of HGPIN on needle biopsy in the contemporary era is related to increased needle biopsy core sampling, which detects many associated cancers on initial biopsy, such that re-biopsy, even with good sampling, does not detect many additional cancers. 8) It is recommended that men do not need routine repeat needle biopsy within the first year following the diagnosis of HGPIN, while further studies are needed to confirm whether routine repeat biopsies should be performed several years following a HGPIN diagnosis on needle biopsy. There were certain results concerning atypical glands suspicious for carcinoma. 1) An average of 5% of needle biopsy pathology reports are diagnosed as atypical glands suspicious for carcinoma. 2) Cases diagnosed as atypical have the highest likelihood of being changed upon expert review and urologists should consider sending such cases for consultation in an attempt to resolve the diagnosis as definitively benign or malignant before subjecting the patient to repeat biopsy. 3) Ancillary techniques using basal cell markers and AMACR (alpha-methyl-acyl-coenzyme A racemase) can decrease the number of atypical diagnoses, and yet one must use these techniques with caution since there are numerous false-positive and false-negative results. 4) The average risk of cancer following an atypical diagnosis is approximately 40%. 5) Clinical and pathological parameters do not help predict which men with an atypical diagnosis have cancer on repeat biopsy. 6) Repeat biopsy should include increased sampling of the initial atypical site, and adjacent ipsilateral and contralateral sites with routine sampling of all sextant sites. Therefore, it is critical for urologists to submit needle biopsy specimens in a manner in which the sextant location of each core can be determined. 7) All men with an atypical diagnosis need re-biopsy within 3 to 6 months. It is critical for urologists to distinguish between a diagnosis of HGPIN and that of atypical foci suspicious for cancer on needle biopsy. These 2 entities indicate different risks of carcinoma on re-biopsy and different recommendations for followup.
Article
Each year, the American Cancer Society estimates the numbers of new cancer cases and deaths expected in the United States in the current year and compiles the most recent data on cancer incidence, mortality, and survival based on incidence data from the National Cancer Institute, the Centers for Disease Control and Prevention, and the North American Association of Central Cancer Registries and mortality data from the National Center for Health Statistics. A total of 1,596,670 new cancer cases and 571,950 deaths from cancer are projected to occur in the United States in 2011. Overall cancer incidence rates were stable in men in the most recent time period after decreasing by 1.9% per year from 2001 to 2005; in women, incidence rates have been declining by 0.6% annually since 1998. Overall cancer death rates decreased in all racial/ethnic groups in both men and women from 1998 through 2007, with the exception of American Indian/Alaska Native women, in whom rates were stable. African American and Hispanic men showed the largest annual decreases in cancer death rates during this time period (2.6% and 2.5%, respectively). Lung cancer death rates showed a significant decline in women after continuously increasing since the 1930s. The reduction in the overall cancer death rates since 1990 in men and 1991 in women translates to the avoidance of about 898,000 deaths from cancer. However, this progress has not benefitted all segments of the population equally; cancer death rates for individuals with the least education are more than twice those of the most educated. The elimination of educational and racial disparities could potentially have avoided about 37% (60,370) of the premature cancer deaths among individuals aged 25 to 64 years in 2007 alone. Further progress can be accelerated by applying existing cancer control knowledge across all segments of the population with an emphasis on those groups in the lowest socioeconomic bracket.
Article
The global burden of cancer continues to increase largely because of the aging and growth of the world population alongside an increasing adoption of cancer-causing behaviors, particularly smoking, in economically developing countries. Based on the GLOBOCAN 2008 estimates, about 12.7 million cancer cases and 7.6 million cancer deaths are estimated to have occurred in 2008; of these, 56% of the cases and 64% of the deaths occurred in the economically developing world. Breast cancer is the most frequently diagnosed cancer and the leading cause of cancer death among females, accounting for 23% of the total cancer cases and 14% of the cancer deaths. Lung cancer is the leading cancer site in males, comprising 17% of the total new cancer cases and 23% of the total cancer deaths. Breast cancer is now also the leading cause of cancer death among females in economically developing countries, a shift from the previous decade during which the most common cause of cancer death was cervical cancer. Further, the mortality burden for lung cancer among females in developing countries is as high as the burden for cervical cancer, with each accounting for 11% of the total female cancer deaths. Although overall cancer incidence rates in the developing world are half those seen in the developed world in both sexes, the overall cancer mortality rates are generally similar. Cancer survival tends to be poorer in developing countries, most likely because of a combination of a late stage at diagnosis and limited access to timely and standard treatment. A substantial proportion of the worldwide burden of cancer could be prevented through the application of existing cancer control knowledge and by implementing programs for tobacco control, vaccination (for liver and cervical cancers), and early detection and treatment, as well as public health campaigns promoting physical activity and a healthier dietary intake. Clinicians, public health professionals, and policy makers can play an active role in accelerating the application of such interventions globally.
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