Choline phospholipid metabolism is altered in a wide variety of cancers. The choline metabolite profile of tumors and cancer cells is characterized by an elevation of phosphocholine and total ...choline-containing compounds. Noninvasive magnetic resonance spectroscopy can be used to detect this elevation as an endogenous biomarker of cancer, or as a predictive biomarker for monitoring tumor response to novel targeted therapies. The enzymes directly causing this elevation, such as choline kinase, phospholipase C and phospholipase D may provide molecular targets for anticancer therapies. Signal transduction pathways that are activated in cancers, such as those mediated by the receptor tyrosine kinases breakpoint cluster region-abelson (Bcr-Abl), c-KIT or epidermal growth factor receptor (EGFR), correlate with the alterations in choline phospholipid metabolism of cancers, and also offer molecular targets for specific anticancer therapies. This review summarizes recently discovered molecular targets in choline phospholipid metabolism and signal transduction pathways, which may lead to novel anticancer therapies potentially being monitored by magnetic resonance spectroscopy techniques.
Notch signaling can promote tumorigenesis in the nervous system and plays important roles in stem‐like cancer cells. However, little is known about how Notch inhibition might alter tumor metabolism, ...particularly in lesions arising in the brain. The gamma‐secretase inhibitor MRK003 was used to treat glioblastoma neurospheres, and they were subdivided into sensitive and insensitive groups in terms of canonical Notch target response. Global metabolomes were then examined using proton magnetic resonance spectroscopy, and changes in intracellular concentration of various metabolites identified which correlate with Notch inhibition. Reductions in glutamate were verified by oxidation‐based colorimetric assays. Interestingly, the alkylating chemotherapeutic agent temozolomide, the mTOR‐inhibitor MLN0128, and the WNT inhibitor LGK974 did not reduce glutamate levels, suggesting that changes to this metabolite might reflect specific downstream effects of Notch blockade in gliomas rather than general sequelae of tumor growth inhibition. Global and targeted expression analyses revealed that multiple genes important in glutamate homeostasis, including glutaminase, are dysregulated after Notch inhibition. Treatment with an allosteric inhibitor of glutaminase, compound 968, could slow glioblastoma growth, and Notch inhibition may act at least in part by regulating glutaminase and glutamate.
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Glutamate metabolism may play a key role in brain tumor growth. These authors investigated the effects of blocking Notch, an important developmental pathway for some stem cells. They tested the change in levels of various metabolites in brain tumor cells when Notch was blocked. They found a drop in glutamate levels, a change that did not occur when they blocked Notch in human neural stem cells nor did they observe this loss of glutamate when they suppressed other signaling pathways. Perhaps, the authors suggest, Notch regulates tumor growth via glutamate metabolism, and hindering glutaminase could be a useful therapeutic avenue.
Hypoxic areas are a common feature of rapidly growing malignant tumors and their metastases and are typically spatially heterogeneous. Hypoxia has a strong impact on tumor cell biology and ...contributes to tumor progression in multiple ways. To date, only a few molecular key players in tumor hypoxia, such as hypoxia-inducible factor-1 (HIF-1), have been discovered. The distribution of biomolecules is frequently heterogeneous in the tumor volume and may be driven by hypoxia and HIF-1α. Understanding the spatially heterogeneous hypoxic response of tumors is critical. Mass spectrometric imaging (MSI) provides a unique way of imaging biomolecular distributions in tissue sections with high spectral and spatial resolution. In this paper, breast tumor xenografts grown from MDA-MB-231-HRE-tdTomato cells, with a red fluorescent tdTomato protein construct under the control of a hypoxia response element (HRE)-containing promoter driven by HIF-1α, were used to detect the spatial distribution of hypoxic regions. We elucidated the 3D spatial relationship between hypoxic regions and the localization of lipids and proteins by using principal component analysis–linear discriminant analysis (PCA-LDA) on 3D rendered MSI volume data from MDA-MB-231-HRE-tdTomato breast tumor xenografts. In this study, we identified hypoxia-regulated proteins active in several distinct pathways such as glucose metabolism, regulation of actin cytoskeleton, protein folding, translation/ribosome, splicesome, the PI3K-Akt signaling pathway, hemoglobin chaperone, protein processing in endoplasmic reticulum, detoxification of reactive oxygen species, aurora B signaling/apoptotic execution phase, the RAS signaling pathway, the FAS signaling pathway/caspase cascade in apoptosis, and telomere stress induced senescence. In parallel, we also identified colocalization of hypoxic regions and various lipid species such as PC(16:0/18:0), PC(16:0/18:1), PC(16:0/18:2), PC(16:1/18:4), PC(18:0/18:1), and PC(18:1/18:1), among others. Our findings shed light on the biomolecular composition of hypoxic tumor regions, which may be responsible for a given tumor’s resistance to radiation or chemotherapy.
Although aberrant glycosylation is recognized as a hallmark of cancer, glycosylation in clinical breast cancer (BC) metastasis has not yet been studied. While preclinical studies show that the ...glycocalyx coating of cancer cells is involved in adhesion, migration, and metastasis, glycosylation changes from primary tumor (PT) to various metastatic sites remain unknown in patients.
We investigated N-glycosylation profiles in 17 metastatic BC patients from our rapid autopsy program. Primary breast tumor, lymph node metastases, multiple systemic metastases, and various normal tissue cores from each patient were arranged on unique single-patient tissue microarrays (TMAs). We performed mass spectrometry imaging (MSI) combined with extensive pathology annotation of these TMAs, and this process enabled spatially differentiated cell-based analysis of N-glycosylation patterns in metastatic BC.
N-glycan abundance increased during metastatic progression independently of BC subtype and treatment regimen, with high-mannose glycans most frequently elevated in BC metastases, followed by fucosylated and complex glycans. Bone metastasis, however, displayed increased core-fucosylation and decreased high-mannose glycans. Consistently, N-glycosylated proteins and N-glycan biosynthesis genes were differentially expressed during metastatic BC progression, with reduced expression of mannose-trimming enzymes and with elevated EpCAM, N-glycan branching, and sialyation enzymes in BC metastases versus PT.
We show in patients that N-glycosylation of breast cancer cells undergoing metastasis occurs in a metastatic site-specific manner, supporting the clinical importance of high-mannose, fucosylated, and complex N-glycans as future diagnostic markers and therapeutic targets in metastatic BC.
NIH grants R01CA213428, R01CA213492, R01CA264901, T32CA193145, Dutch Province Limburg "LINK", European Union ERA-NET TRANSCAN2-643638.
Multiple sclerosis (MS) is a neurodegenerative disease, wherein aberrant immune cells target myelin-ensheathed nerves. Conventional magnetic resonance imaging (MRI) can be performed to monitor damage ...to the central nervous system that results from previous inflammation; however, these imaging biomarkers are not necessarily indicative of active, progressive stages of the disease. The immune cells responsible for MS are first activated and sensitized to myelin in lymph nodes (LNs). Here, we present a new strategy for monitoring active disease activity in MS, chemical exchange saturation transfer (CEST) MRI of LNs.
We studied the potential utility of conventional (T2-weighted) and CEST MRI to monitor changes in these LNs during disease progression in an experimental autoimmune encephalomyelitis (EAE) model. We found CEST signal changes corresponded temporally with disease activity. CEST signals at the 3.2 ppm frequency during the active stage of EAE correlated significantly with the cellular (flow cytometry) and metabolic (mass spectrometry imaging) composition of the LNs, as well as immune cell infiltration into brain and spinal cord tissue. Correlating primary metabolites as identified by matrix-assisted laser desorption/ionization (MALDI) imaging included alanine, lactate, leucine, malate, and phenylalanine.
Taken together, we demonstrate the utility of CEST MRI signal changes in superficial cervical LNs as a complementary imaging biomarker for monitoring disease activity in MS. CEST MRI biomarkers corresponded to disease activity, correlated with immune activation (surface markers, antigen-stimulated proliferation), and correlated with LN metabolite levels.
Abstract Collagen 1 (Col1) fibers play an important role in tumor interstitial macromolecular transport and cancer cell dissemination. Our goal was to understand the influence of Col1 fibers on water ...diffusion, and to examine the potential of using noninvasive diffusion tensor imaging (DTI) to indirectly detect Col1 fibers in breast lesions. We previously observed, in human MDA-MB-231 breast cancer xenografts engineered to fluoresce under hypoxia, relatively low amounts of Col1 fibers in fluorescent hypoxic regions. These xenograft tumors together with human breast cancer samples were used here to investigate the relationship between Col1 fibers, water diffusion and anisotropy, and hypoxia. Hypoxic low Col1 fiber containing regions showed decreased apparent diffusion coefficient (ADC) and fractional anisotropy (FA) compared to normoxic high Col1 fiber containing regions. Necrotic high Col1 fiber containing regions showed increased ADC with decreased FA values compared to normoxic viable high Col1 fiber regions that had increased ADC with increased FA values. A good agreement of ADC and FA patterns was observed between in vivo and ex vivo images. In human breast cancer specimens, ADC and FA decreased in low Col1 containing regions. Our data suggest that a decrease in ADC and FA values observed within a lesion could predict hypoxia, and a pattern of high ADC with low FA values could predict necrosis. Collectively the data identify the role of Col1 fibers in directed water movement and support expanding the evaluation of DTI parameters as surrogates for Col1 fiber patterns associated with specific tumor microenvironments as companion diagnostics and for staging.
Magnetic resonance studies from the last 10 years have conclusively demonstrated that choline metabolism is altered in a wide variety of cancers. In cancer, the choline metabolite profile is ...characterized by an elevation of phosphocholine and total choline-containing compounds. This elevation is increasingly being used as an endogenous biomarker of cancer. Importantly, the enzymes and pathways resulting in these distinct alterations in phosphocholine and total choline may provide novel molecular targets for specific, targeted anticancer therapies. In this article, we have summarized some of the magnetic resonance spectroscopy and positron emission tomography techniques that are currently available, or will be in the near future, for choline metabolism-based diagnosis, staging and therapy assessment in cancer patients. This review also outlines currently known molecular alterations that cause the aberrant choline metabolite profile in cancers and concludes with a summary of recent research findings that may, in the future, lead to novel anticancer therapies targeting choline metabolism.
Imaging mass spectrometry (IMS) is a promising technology which allows for detailed analysis of spatial distributions of (bio)molecules in organic samples. In many current applications, IMS relies ...heavily on (semi)automated exploratory data analysis procedures to decompose the data into characteristic component spectra and corresponding abundance maps, visualizing spectral and spatial structure. The most commonly used techniques are principal component analysis (PCA) and independent component analysis (ICA). Both methods operate in an unsupervised manner. However, their decomposition estimates usually feature negative counts and are not amenable to direct physical interpretation. We propose probabilistic latent semantic analysis (pLSA) for non-negative decomposition and the elucidation of interpretable component spectra and abundance maps. We compare this algorithm to PCA, ICA, and non-negative PARAFAC (parallel factors analysis) and show on simulated and real-world data that pLSA and non-negative PARAFAC are superior to PCA or ICA in terms of complementarity of the resulting components and reconstruction accuracy. We further combine pLSA decomposition with a statistical complexity estimation scheme based on the Akaike information criterion (AIC) to automatically estimate the number of components present in a tissue sample data set and show that this results in sensible complexity estimates.
Phosphocholine (PC) and total choline (tCho) are increased in malignant breast tumors. In this study, we combined magnetic resonance spectroscopic imaging (MRSI), mass spectrometry (MS) imaging, and ...pathologic assessment of corresponding tumor sections to investigate the localization of choline metabolites and cations in viable versus necrotic tumor regions in the nonmetastatic MCF-7 and the highly metastatic MDA-MB-231 breast cancer xenograft models. In vivo three-dimensional MRSI showed that high tCho levels, consisting of free choline (Cho), PC, and glycerophosphocholine (GPC), displayed a heterogeneous spatial distribution in the tumor. MS imaging performed on tumor sections detected the spatial distributions of individual PC, Cho, and GPC, as well as sodium (Na+) and potassium (K+), among many others. PC and Cho intensity were increased in viable compared with necrotic regions of MDA-MB-231 tumors, but relatively homogeneously distributed in MCF-7 tumors. Such behavior may be related to the role of PC and PC-related enzymes, such as choline kinase, choline transporters, and others, in malignant tumor growth. Na+ and K+ colocalized in the necrotic tumor areas of MDA-MB-231 tumors, whereas in MCF-7 tumors, Na+ was detected in necrotic and K+ in viable tumor regions. This may be attributed to differential Na+/K+ pump functions and K+ channel expressions. Principal component analysis of the MS imaging data clearly identified different tumor microenvironmental regions by their distinct molecular signatures. This molecular information allowed us to differentiate between distinct tumor regions and tumor types, which may, in the future, prove clinically useful in the pathologic assessment of breast cancers.
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