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Ganapathy V, Thangaraju M, Prasad PD.. Nutrient transporters in cancer: relevance to Warburg hypothesis and beyond. Pharmacol Ther 121: 29-40
Abstract
Tumor cells have an increased demand for nutrients; this demand is met by increased availability of nutrients through vasculogenesis and by enhanced cellular entry of nutrients through upregulation of specific transporters. This review focuses on three groups of nutrient transporters relevant to cancer: glucose transporters, lactate transporters, and amino acid transporters. Tumor cells enhance glucose uptake via induction of GLUT1 and SGLT1, and coordinate the increased entry of glucose with increased glycolysis. Since enhanced glycolysis in cancer is associated with lactate production, tumor cells must find a way to eliminate lactic acid to prevent cellular acidification. This is achieved by the upregulation of MCT4, a H+-coupled lactate transporter. In addition, the Na+-coupled lactate transporter SMCT1 is silenced in cancer. SMCT1 also transports butyrate and pyruvate, which are inhibitors of histone deacetylases. The silencing of SMCT1 occurs in cancers of a variety of tissues. Re-expression of SMCT1 in cancer cell lines leads to growth arrest and apoptosis in the presence of butyrate or pyruvate, suggesting that the transporter may function as a tumor suppressor. Tumor cells meet their amino acid demands by inducing xCT/4F2hc, LAT1/4F2hc, ASCT2, and ATB0,+. xCT/4F2hc is related primarily to glutathione status, protection against oxidative stress, and cell cycle progression, whereas the other three transporters are related to amino acid nutrition. Pharmacologic blockade of LAT1/4F2hc, xCT/4F2hc, or ATB0,+ leads to inhibition of cancer cell growth. Since tumor cells selectively regulate these nutrient transporters to support their rapid growth, these transporters have potential as drug targets for cancer therapy.
In this work, we design and synthesize 2,2'-(7,9-dimethyl-2,4,6,8-tetraoxo-6,7,8,9-tetrahydropyrimido[5,4-g]pteridine-1,3(2H,4H)-diyl)bis(N,N-bis(2-chloroethyl)acetamide) (PT-MCA) as a novel DNA intercalator and potential antitumor agent. Electrochemical analysis reveals the redox process of PT-MCA on the electrode surface. The bioelectrochemical sensors are obtained by modifying the surface of GCE with calf thymus DNA (ctDNA), poly (dG), poly (dA), and G-quadruplex, respectively. The DNA oxidative damage induced by PT-MCA is investigated by comparing the peak intensity change of dGuo and dAdo and monitoring the peaks of the oxidation products of guanine and/or adenine (8-oxoGua and/or 2,8-oxoAde). UV-vis absorption and fluorescence spectra and gel electrophoresis are further employed to understand the intercalation of PT-MCA into DNA base pairs. Moreover, PT-MCA is proved to exhibit stronger anti-proliferation activity than mitoxantrone against both 4T1 and B16-F10 cancer cells. At last, the oxidative damage of PT-MCA toward ctDNA is not interfered by the coexistence of ions and also can be detected in real serums.
This chapter aims to familiarize readers with the basics of correlative imaging, the strengths and shortcomings of various imaging modalities, and how the correlation among them leads to better understanding of pathologies. It also aims to familiarize nuclear physicians and radiologists with the basic principles of tomographic techniques utilized in correlative/fusion imaging. After the advent of positron emission tomography (PET)‐computed tomography (CT) in the 1990s, the initial PET‐CT acquisition protocols utilized CT as a fast transmission source for attenuation correction, with little additional information for anatomic localization. PET‐CT scanners allow excellent fusion of PET and CT images and improve lesion localization and interpretation accuracy. The chapter consists of the pictorial demonstration of clinical case examples where fusion imaging of FDG PET and CT resulted in accurate diagnosis of pathology and its extent that would have been otherwise difficult to reach. Most of the newer single‐photon emission tomography systems use statistical iterative reconstruction for creating images.
Molecular imaging has witnessed a great progress in the field of oncology over the past few decades. Radiolabeled amino acid (AA) tracers are particularly helpful in the areas where the utility of 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography with computed tomography imaging has been limited such as in evaluating brain tumors, neuroendocrine tumors (NETs), and prostate cancer. Radiolabeled AA tracers such as 6-[18F]-L-fluoro-L-3, 4-dihydroxyphenylalanine (18F-FDOPA), 18F-fluoro-ethyl-tyrosine (18F-FET), and 11C-methionine have found wide applications in brain tumors, which, unlike 18F-FDG, concentrate in the tumor tissue to a greater extent than that in normal brain tissue by providing accurate information about tumor volume and boundaries. 18F-FDOPA is also useful in evaluating NETs. Tracers such as 18F-FACBC (Fluciclovine) and anti-1-amino-2-[18F]fluorocyclopentyl-1-carboxylic acid (18F-FACPC) are used in imaging of prostate cancer and provide valuable information of locoregional, recurrent, and metastatic disease. This review highlights AA tracers and their major applications in imaging, viz., in evaluating brain tumors, NETs, and prostate cancer.
The Nobel Prize Winner (1931) Dr. Otto H. Warburg had established that the primary energy source of the cancer cell is aerobic glycolysis (the Warburg effect). He also postulated the hypothesis about "the prime cause of cancer", which is a matter of debate nowadays. Contrary to the hypothesis, his discovery was recognized entirely. However, the discovery had almost vanished in the heat of battle about the hypothesis. The prime cause of cancer is essential for the prevention and diagnosis, yet the effects that inf luence tumor growth are more important for cancer treatment. Due to the Warburg effect, a large amount of data has been accumulated on biochemical changes in the cell and the organism as a whole. Due to the Warburg effect, the recovery of normal biochemistry and oxygen respiration and the restoration of the work of mitochondria of cancer cells can inhibit tumor growth and lead to remission. Here, we review the current knowledge on the inhibition of abnormal glycolysis, neutralization of its consequences, and normalization of biochemical parameters, as well as recovery of oxygen respiration of a cancer cell and mitochondrial function from the point of view of classical biochemistry and organic chemistry.
Lung cancer is the leading cause of cancer deaths globally; 1 in 16 people are diagnosed with lung cancer in their lifetime. Microtubules, a critical cytoskeletal assembly, have an essential role in cell division. Interference with the microtubule assembly leads to genetic instability during mitosis and cancer cell death. Currently, available antimitotic drugs such as vincas and taxanes are limited due to side effects such as alopecia, myelosuppression, and drug resistance. Noscapine, an opium alkaloid, is a tubulin-binding agent and can alter the microtubule assembly, causing cancer cell death. Amino acids are fundamental building blocks for protein synthesis, making them essential for the biosynthesis of cancer cells. However, the ability of amino acids in drug transportation has yet to be exploited in developing noscapine analogues as a potential drug candidate for cancer. Hence, in the present study, we have explored the ninth position of noscapine by introducing a hydroxymethylene group using the Blanc reaction and further coupled it with a series of amino acids to construct five target conjugates in good yields. The synthesized amino acid conjugate molecules were biologically evaluated against the A549 lung cancer cell line, among which the noscapine-tryptophan conjugate showed IC50 = 32 μM, as compared to noscapine alone (IC50 = 73 μM). Morphological changes in cancer cells, cell cycle arrest in the G1 phase, and ethidium bromide/acridine orange staining indicated promising anticancer properties. Molecular docking confirmed strong binding to tubulin, with a score of -41.47 kJ/mol with all 3D coordinates and significant involvement of molecular forces, including the hydrogen bonds and hydrophobic interactions. Molecular dynamics simulations demonstrated a stable binding of noscapine-tryptophan conjugate for a prolonged time (100 ns) with the involvement of free energy through the reaction coordinates analyses, solving the bioavailability of parent noscapine to the body.
Neoadjuvant chemotherapy (NACT) is offered to patients with operable or inoperable breast cancer (BC) to downstage the disease. Clinical responses to NACT may vary depending on a few known clinical and biological features, but the diversity of responses to NACT is not fully understood. In this study, 80 women had their metabolite profiles of pre-treatment sera analyzed for potential NACT response biomarker candidates in combination with immunohistochemical parameters using Nuclear Magnetic Resonance (NMR). Sixty-four percent of the patients were resistant to chemotherapy. NMR, hormonal receptors (HR), human epidermal growth factor receptor 2 (HER2), and the nuclear protein Ki67 were combined through machine learning (ML) to predict the response to NACT. Metabolites such as leucine, formate, valine, and proline, along with hormone receptor status, were discriminants of response to NACT. The glyoxylate and dicarboxylate metabolism was found to be involved in the resistance to NACT. We obtained an accuracy in excess of 80% for the prediction of response to NACT combining metabolomic and tumor profile data. Our results suggest that NMR data can substantially enhance the prediction of response to NACT when used in combination with already known response prediction factors.
(1) Background: Many transporters of the SLC22 family (e.g., OAT1, OAT3, OCT2, URAT1, and OCTN2) are highly expressed in the kidney. They transport drugs, metabolites, signaling molecules, antioxidants, nutrients, and gut microbiome products. According to the Remote Sensing and Signaling Theory, SLC22 transporters play a critical role in small molecule communication between organelles, cells and organs as well as between the body and the gut microbiome. This raises the question about the potential role of SLC22 transporters in cancer biology and treatment. (2) Results: In two renal cell carcinoma RNA-seq datasets found in TCGA, KIRC and KIRP, there were multiple differentially expressed (DE) SLC22 transporter genes compared to normal kidney. These included SLC22A6, SLC22A7, SLC22A8, SLC22A12, and SLC22A13. The patients with disease had an association between overall survival and expression for most of these DE genes. In KIRC, the stratification of patient data by pathological tumor characteristics revealed the importance of SLC22A2, SLC22A6, and SLC22A12 in disease progression. Interaction networks combining the SLC22 with ADME genes supported the centrality of SLC22 transporters and other transporters (ABCG2, SLC47A1) in disease progression. (3) Implications: The fact that many of these genes are uric acid transporters is interesting because altered uric acid levels have been associated with kidney cancer. Moreover, these genes play key roles in processing metabolites and chemotherapeutic compounds, thus making them potential therapeutic targets. Finally, our analyses raise the possibility that current approaches may undertreat certain kidney cancer patients with low SLC22 expression and only localized disease while possibly overtreating more advanced disease in patients with higher SLC22 expression. Clinical studies are needed to investigate these possibilities.
Breast cancer (BC) is a malignant tumor that seriously endangers health in women. BC, like other cancers, is accompanied by metabolic reprogramming. Among energy metabolism-related pathways, BC exhibits enhanced glycolysis, tricarboxylic acid (TCA) cycle, pentose phosphate pathway (PPP), glutamate metabolism, and fatty acid metabolism activities. These pathways facilitate the proliferation, growth and migration of BC cells. The progression of BC is closely related to the alterations in the activity or expression level of several metabolic enzymes, which are regulated by the intrinsic factors such as the key signaling and transcription factors. The metabolic reprogramming in the progression of BC is attributed to the aberrant expression of the signaling and transcription factors associated with the energy metabolism pathways. Understanding the metabolic mechanisms underlying the development of BC will provide a druggable potential for BC treatment and drug discovery.
Few prospective cohort trials have investigted the effect of pretreatment nutritional and inflammatory status on the clinical outcome of patients with cancer and optimal performance status and assessed the interplay between nutrition, inflammation, body composition, and circulating metabolites before treatment. Here, 50 patients with locally advanced head and neck squamous cell carcinoma (LAHNSCC) and Eastern Cooperative Oncology Group performance status (ECOG PS) ≤ 2 were prospectively recruited along with 43 healthy participants. Before concurrent chemoradiotherapy, compared with healthy controls, the cancer group showed lower levels of histidine, leucine, and phenylalanine and had low values in anthropometric and body composition measurements; however, the group displayed higher ornithine levels, more malnutrition, and severe inflammation. Pretreatment advanced Glasgow prognostic score (1 and 2) status was the sole prognostic factor for 3-year mortality rate and was associated with age and serum histidine levels in patients with cancer. Thus, even at the same tumor stage and ECOG PS, patients with LAHNSCC, poor nutrition, and high inflammation severity at baseline may have inferior survival outcomes than those with adequate nutrition and low inflammation severity. Assessment of pretreatment nutritional and inflammatory status should be included in the enrollment criteria in future studies.
Compared to normal cells, cancer cells generate ATP mainly through aerobic glycolysis, which promotes tumorigenesis and tumor progression. Long non-coding RNAs (LncRNAs) are a class of transcripts longer than 200 nucleotides with little or without evident protein-encoding function. LncRNAs are involved in the ten hallmarks of cancer, interestingly, they are also closely associated with aerobic glycolysis. However, the mechanism of this process is non-transparent to date. Demonstrating the mechanism of lncRNAs regulating tumorigenesis and tumor progression through aerobic glycolysis is particularly critical for cancer therapy, and may provide novel therapeutic targets or strategies in cancer treatment. In this review, we discuss the role of lncRNAs and aerobic glycolysis in tumorigenesis and tumor progression, and further explore their interaction, in hope to provide a novel therapeutic target for cancer treatment.
HCC is well known for low glycolysis in the tumors, whereas hypoxia induces glycolytic phenotype and tumor progression. This study was conducted to evaluate the expression of SLCs in human HCCs and investigated whether extracellular nutrient administration related to SLCs in low-glycolytic HCC can prevent hypoxic tumor progression. SLCs expression was screened according to the level of glycolysis in HCCs. Then, whether extracellular nutrient treatment can affect hypoxic tumor progression, as well as the mechanisms, were evaluated in an in vitro cell line and an in vivo animal model. Low-glycolytic HCCs showed high SLC13A5/NaCT and SLC16A1/MCT1 but low SLC2A1/GLUT1 and HIF1α/HIF1α expression. Especially, high SLC13A5 expression was significantly associated with good overall survival in the Cancer Genome Atlas (TCGA) database. In HepG2 cells with the highest NaCT expression, extracellular citrate treatment upon hypoxia induced HIF1α degradation, which led to reduced glycolysis and cellular proliferation. Finally, in HepG2-animal models, the citrate-treated group showed smaller tumor with less hypoxic areas than the vehicle-treated group. In patients with HCC, SLC13A5/NaCT is an important SLC, which is associated with low glycolysis and good prognosis. Extracellular citrate treatment induced the failure of metabolic adaptation to hypoxia and tumor growth inhibition, which can be a potential therapeutic strategy in HCCs.
We investigated risk factors for treatment interruption (TI) in patients with locally advanced head and neck squamous-cell carcinoma (LAHNSCC) following concurrent chemoradiotherapy (CCRT), under the provision of recommended calorie and protein intake; we also evaluated the associations between clinicopathological variables, calorie and protein supply, nutrition–inflammation biomarkers (NIBs), total body composition change (TBC), and a four-serum-amino-acid metabolite panel (histidine, leucine, ornithine, and phenylalanine) among these patients. Patients with LAHNSCC who completed the entire planned CCRT course and received at least 25 kcal/kg/day and 1 g of protein/kg/day during CCRT were prospectively recruited. Clinicopathological variables, anthropometric data, blood NIBs, CCRT-related factors, TBC data, and metabolite panels before and after treatment were collected; 44 patients with LAHNSCC were enrolled. Nine patients (20.4%) experienced TIs. Patients with TIs experienced greater reductions in hemoglobin, serum levels of albumin, uric acid, histidine, and appendicular skeletal mass, and suffered from more grade 3/4 toxicities than those with no TI. Neither increased daily calorie supply (≥30 kcal/kg/day) nor feeding tube placement was correlated with TI. Multivariate analysis showed that treatment-interval changes in serum albumin and histidine levels, but not treatment toxicity, were independently associated with TI. Thus, changes in serum levels of albumin and histidine over the treatment course could cause TI in patients with LAHNSCC following CCRT.
Background: Cluster of differentiation 98 heavy chain (CD98hc) is a transmembrane protein, which functions both as a coreceptor of ß-integrins, enhancing intracellular integrin-dependent downstream signaling, and as a transporter of branched-chain and aromatic amino acids. As such, it is pivotal in cell cycle regulation and protection of oxidative, nutritional and DNA replication stress. Overexpression of CD98hc occurs widely in cancer cells and is associated with poor clinical prognosis. The role of CD98hc in pancreatic cancer remains to be elucidated. The aim of this study was to determine the expression of CD98hc in pancreatic ductal adenocarcinoma and to define its potential functional role in cancer cell biology. Methods: Immunohistochemical staining for CD98hc was performed on 222 tissue samples of patients with pancreatic ductal adenocarcinoma. The pancreatic cancer cell lines PANC-1 and BxPC-3 were used to determine the effect of CD98hc expression on cancer cell behavior using cell adhesion, cell trans-migration and cell spreading assays. Flow cytometry was performed to study the rate of apoptosis after detachment or serum starvation. shRNA-lentiviral constructs were used to knock down or reconstitute full length or mutated CD98hc. Results: Up to 20% of pancreatic ductal adenocarcinomas express CD98hc in the acinar cells (13%) and islet cells (20%) embedded in tumor tissue. Although expression of CD98hc in tumor tissue was not associated with a particular tumor stage or grade, our data show a trend towards longer overall survival of pancreatic cancer patients without CD98hc expression as compared to those with immunohistochemical positivity. In vitro downregulation of CD98hc in the pancreatic cancer cell lines PANC-1 and BxPC-3 significantly inhibits cell proliferation (p<0.05), self-renewal (p<0.05) and anchorage-independent growth (p<0.05). Conclusion: CD98hc is expressed in a remarkable percentage of pancreatic ductal adenocarcinomas. Due to its important role in cell behavior and malignant cell transformation, it may be a promising molecular target for potential new therapeutic approaches in pancreatic cancer in the future.
Despite all precautionary actions and the possibility of using vaccinations to counteract infections caused by human papillomaviruses (HPVs), HPV-related cancers still account for approximately 5% of all carcinomas. Worldwide, many women are still excluded from adequate health care due to their social position and origin. Therefore, immense efforts in research and therapy are still required to counteract the challenges that this disease entails. The special thing about an HPV infection is that it is not only able to trick the immune system in a sophisticated way, but also, through genetic integration into the host genome, to use all the resources available to the host cells to complete the replication cycle of the virus without activating the alarm mechanisms of immune recognition and elimination. The mechanisms utilized by the virus are the metabolic, immune, and hormonal signaling pathways that it manipulates. Since the virus is dependent on replication enzymes of the host cells, it also intervenes in the cell cycle of the differentiating keratinocytes and shifts their terminal differentiation to the uppermost layers of the squamocolumnar transformation zone (TZ) of the cervix. The individual signaling pathways are closely related and equally important not only for the successful replication of the virus but also for the onset of cervical cancer. We will therefore analyze the effects of HPV infection on metabolic signaling, as well as changes in hormonal and immune signaling in the tumor and its microenvironment to understand how each level of signaling interacts to promote tumorigenesis of cervical cancer.
Sodium-glucose transporter 2 (SGLT2) inhibitors are antidiabetic drugs affecting SGLT2. Recent studies have shown various cancers expressing SGLT2, and SGLT2 inhibitors attenuating tumor proliferation. We evaluated the antitumor activities of canagliflozin, a SGLT2 inhibitor, on glioblastoma (GBM). Three GBM cell lines, U251MG (human), U87MG (human), and GL261 (murine), were used. We assessed the expression of SGLT2 of GBM through immunoblotting, specimen-use, cell viability assays, and glucose uptake assay with canagliflozin. Then, we assessed phosphorylation of AMP-activated protein kinase (AMPK), p70 S6 kinase, and S6 ribosomal protein by immunoblotting. Concentrations of 5, 10, 20, and 40 μM canagliflozin were used in these tests. We also evaluated cell viability and immunoblotting using U251MG with siRNA knockdown of SGLT2. Furthermore, we divided the mice into vehicle group and canagliflozin group. The canagliflozin group was administrated with 100 mg/kg of canagliflozin orally for 10 days starting from the third days post-GBM transplant. The brains were removed and the tumor volume was evaluated using sections. SGLT2 was expressed in GBM cell and GBM allograft mouse. Canagliflozin administration at 40 μM significantly inhibited cell proliferation and glucose uptake into the cell. Additionally, canagliflozin at 40 μM significantly increased the phosphorylation of AMPK and suppressed that of p70 S6 kinase and S6 ribosomal protein. Similar results of cell viability assays and immunoblotting were obtained using siRNA SGLT2. Furthermore, although less effective than in vitro, the canagliflozin group significantly suppressed tumor growth in GBM-transplanted mice. This suggests that canagliflozin can be used as a potential treatment for GBM.
Iron homeostasis is altered in tumours in response to a perturbation in the expression of iron-dependent proteins. Therefore, iron chelators make cancerous cells more vulnerable to iron deficiency. Compounds having thiosemicarbazide scaffold with the ability to metal complex formation have the potential to act as anticancer. A series of thiosemicarbazide derivatives were designed, synthesized successfully and their cytotoxicity was then tested on some cancerous as well as laboratory normal model systems by using colorimetric assay based on WST-1 reagent. According to the cytotoxicity results, some compounds showed high toxicity effect on both the cancerous and healthy cell lines. The results of toxicity assays on U87 and A549 cell lines showed the survivability less than 50% at all concentrations higher than 10 ppm for all the synthesized compounds. The MCF-7 cell line approximately exhibited the same behaviour and had survivability less than 60%. The 3T3 in compared with HUVEC cell line showed a completely different behaviour against the synthesized compounds and had survivability more than 50%. The selectivity index was also measured and based on the study results this could be concluded that the cytotoxicity profile of the synthesized compounds on 3T3 cell line shows a significant difference, indicating a good anticancer effect for these compounds.
Purpose:
Recently, glucose and amino acid transporters have gradually become a hot topic in thyroid gland biology and cancer research. We aimed to investigate the expressions of glucose transporter 1 (GLUT1) and glutamine transporter 2 (ASCT2) in papillary thyroid carcinoma (PTC) and their clinical significance and relation to HCV-related hepatitis.
Patients and methods:
Screening 202 TC tissue samples against the selection criteria using a propensity-score matched analysis to adjust for age, sex, side of tumor, histopathological variants, TNM staging system, and the positivity for HCV yielded 51 matched (17 HCV positive and 34 HCV negative) PTC samples. The expressions of GLUT1 and ASCT2 expressions were detected by immunohistochemical staining. Kaplan-Meier survival curves were generated for disease-free and overall survival, and multivariate Cox regression analysis was applied to identify predictors for mortality.
Results:
Of 51 thyroid cancer tissues, 85% showed positive GLUT1 cytoplasmic staining, and 26% had a high expression score. All thyroid cancer specimens demonstrated ASCT2 cytoplasmic staining with membranous accentuation. Of these, 78% showed a high expression score, and 22% showed weak staining. On stratifying the study cohort based on the HCV status, HCV negative cohort showed a significantly higher immunoreactivity score for GLUT1 (p = 0.004) but not ASCT2 (p = 0.94) than HCV positive group. The expressions of the studied transporters showed no significant associations with the prognostic features of PTC nor the disease-free/overall survival.
Conclusion:
GLUT1 and ASCT2 immunohistochemical staining showed positive expression with variable intensity in nearly 85% and 100% of PTC tissue samples compared to normal ones, respectively. Furthermore, GLUT1 protein expression, not ASCT2, showed a higher immunoreactivity score in PTC patients who are negative for HCV than cancer patients with positive HCV. Meanwhile, the expression of both protein markers was not associated with the clinicopathological characteristics of the studied PTC patients. Further large-scale multicenter studies are recommended to validate the present findings.
Background
Hepatocellular carcinoma (HCC) is the most common primary liver cancer in the world. Although great advances in HCC diagnosis and treatment have been achieved, due to the complicated mechanisms in tumor development and progression, the prognosis of HCC is still dismal. Recent studies have revealed that the Warburg effect is related to the development, progression and treatment of various cancers; however, there have been a few explorations of the relationship between glycolysis and HCC prognosis.
Methods
mRNA expression profiling was downloaded from public databases. Gene set enrichment analysis (GSEA) was used to explore glycolysis-related genes (GRGs), and the LASSO method and Cox regression analysis were used to identify GRGs related to HCC prognosis and to construct predictive models associated with overall survival (OS) and disease-free survival (DFS). The relationship between the predictive model and the tumor mutation burden (TMB) and tumor immune microenvironment (TIME) was explored. Finally, real-time PCR was used to validate the expression levels of the GRGs in clinical samples and different cell lines.
Results
Five GRGs (ABCB6, ANKZF1, B3GAT3, KIF20A and STC2) were identified and used to construct gene signatures to predict HCC OS and DFS. Using the median value, HCC patients were divided into low- and high-risk groups. Patients in the high-risk group had worse OS/DFS than those in the low-risk group, were related to higher TMB and were associated with a higher rate of CD4+ memory T cells resting and CD4+ memory T cells activated. Finally, real-time PCR suggested that the five GRGs were all dysregulated in HCC samples compared to adjacent normal samples.
Conclusions
We identified five GRGs associated with HCC prognosis and constructed two GRGs-related gene signatures to predict HCC OS and DFS. The findings in this study may contribute to the prediction of prognosis and promote HCC treatment.
Amino acid transporters are expressed in mammalian cells not only in the plasma membrane but also in intracellular membranes. The conventional function of these transporters is to transfer their amino acid substrates across the lipid bilayer; the direction of the transfer is dictated by the combined gradients for the amino acid substrates and the co-transported ions (Na+, H+, K+ or Cl−) across the membrane. In cases of electrogenic transporters, the membrane potential also contributes to the direction of the amino acid transfer. In addition to this expected traditional function, several unconventional functions are known for some of these amino acid transporters. This includes their role in intracellular signaling, regulation of acid–base balance, and entry of viruses into cells. Such functions expand the biological roles of these transporters beyond the logical amino acid homeostasis. In recent years, two additional unconventional biochemical/metabolic processes regulated by certain amino acid transporters have come to be recognized: macropinocytosis and obesity. This adds to the repertoire of biological processes that are controlled and regulated by amino acid transporters in health and disease. In the present review, we highlight the unusual involvement of selective amino acid transporters in macropinocytosis (SLC38A5/SLC38A3) and diet-induced obesity/metabolic syndrome (SLC6A19/SLC6A14/SLC6A6).
Common capabilities of thyroid malignant cells are accelerating metabolism and increasing glucose uptake to optimize energy supply for growth. In tumor cells, keeping the power load required for cell survival is essential and glucose transporters are capable of promoting this task. GLUT-1 and GLUT3 are promising goals for the development of anti-cancer strategies. The lack of oncosuppressors has dominant effect on the membrane expression of GLUT1 and glucose uptake. Overexpression of hypoxia-inducing factors, in thyroid cancer, modulates the expression of some glucose transporter genes. Although the physiology of the thyroid gland has been excellently explained, metabolic regulation in thyroid cancer is inevitable. In this section, we investigated the proliferation pathways of pivotal regulators and signal molecules around GLUT regulation in thyroid cancer, including PTEN, p53, MicroRNA, iodide, BRAF, HIF-1, PI3K-Akt, TSH, c-Myc, and AMPK. Impaired energy regulation and cell metabolism are the most critical symptoms of most cancers. As a result, understanding the mechanisms of glucose transport in the normal and pathological tissues of the thyroid may be very crucial and offer tremendous insights into the science of analysis and remedy of thyroid disease.
The unique ability to adapt and thrive in inhospitable, stressful tumor microenvironments (TME) also renders cancer cells resistant to traditional chemotherapeutic treatments and/or novel pharmaceuticals. Cancer cells exhibit extensive metabolic alterations involving hypoxia, accelerated glycolysis, oxidative stress, and increased extracellular ATP that may activate ancient, conserved prion adaptive response strategies that exacerbate multidrug resistance (MDR) by exploiting cellular stress to increase cancer metastatic potential and stemness, balance proliferation and differentiation, and amplify resistance to apoptosis. The regulation of prions in MDR is further complicated by important, putative physiological functions of ligand-binding and signal transduction. Melatonin is capable of both enhancing physiological functions and inhibiting oncogenic properties of prion proteins. Through regulation of phase separation of the prion N-terminal domain which targets and interacts with lipid rafts, melatonin may prevent conformational changes that can result in aggregation and/or conversion to pathological, infectious isoforms. As a cancer therapy adjuvant, melatonin could modulate TME oxidative stress levels and hypoxia, reverse pH gradient changes, reduce lipid peroxidation, and protect lipid raft compositions to suppress prion-mediated, non-Mendelian, heritable, but often reversible epigenetic adaptations that facilitate cancer heterogeneity, stemness, metastasis, and drug resistance. This review examines some of the mechanisms that may balance physiological and pathological effects of prions and prion-like proteins achieved through the synergistic use of melatonin to ameliorate MDR, which remains a challenge in cancer treatment.
The prevalence of obesity and type 2 diabetes is higher in French Guiana compared to mainland France. These metabolic disorders are associated with an increased risk of cancer. One of the factors involved is hyperinsulinemia that promotes the action of glucose transporter 1 (GLUT-1). The objective of this study is to characterize the expression of GLUT-1 in breast cancers cells in diabetic and obese patients compared to those who are not and to describe the clinical and histological prognostic factors of breast cancer in this population. We conducted a monocentric study including patients with breast cancer diagnosed between 2014 and 2020. Patients were classified into three groups: diabetes, obesity, and control group. The GLUT-1 expression was assessed by immunohistochemistry. In total, 199 patients were included in this study. The median age was 53.5 years, and the median tumor size was 2.8 cm. Luminal A was the most frequent molecular type (58.1%), followed by the triple-negative type (19.9%). The breast cancer in our population was characterized by a younger age at diagnosis, more aggressive molecular types, and larger tumor size. Thus, we suggest the advancement of the age of breast cancer screening in this territory. A total of 144 patients (31 diabetes, 22 obese, and 91 control group) were included for the study of GLUT-1 expression. Overexpression of GLUT-1 was observed in 60.4% of cases and in all carcinoma in situ lesions. GLUT-1 overexpression was associated with more aggressive cancers. This overexpression is correlated with high histological grade, high proliferation index, and aggressive molecular types. Our study found no difference in GLUT-1 expression between the diabetic or obese patients and the control group. These results highlight the potential role of GLUT-1 as a tumor metabolic prognostic marker and also as an interesting target therapy, independently of patient metabolic disorder.
A critical obstacle to the successful treatment of colorectal cancer (CRC) is chemoresistance. Chemoresistant CRC cells contribute to treatment failure by providing a mechanism of drug lethargy and modifying chemoresistance-associated molecules. The gut microbiota provide prophylactic and therapeutic effects by targeting CRC through anticancer mechanisms. Among them, Lactobacillus plantarum contributes to the health of the host and is clinically effective in treating CRC. This study confirmed that 5-fluorouracil (5-FU)-resistant CRC HCT116 (HCT116/5FUR) cells acquired butyrate-insensitive properties. To date, the relationship between 5-FU-resistant CRC and butyrate resistance has not been elucidated. Here, we demonstrated that the acquisition of butyrate resistance in HCT116/5FUR cells was strongly correlated with the inhibition of the expression and function of SMCT1, a major transporter of butyrate in colonocytes. L. plantarum-cultured cell-free supernatant (LP) restored the functional expression of SMCT1 in HCT116/5FUR cells, leading to butyrate-induced antiproliferative effect and apoptosis. These results suggest that LP has a synergistic effect on the SMCT1/butyrate-mediated tumor suppressor function and is a potential chemosensitizer to overcome dual 5-FU and butyrate resistance in HCT116 cells.
Cystic echinococcosis (CE) is a zoonotic parasitic disease caused by infection with the larvae of Echinococcus granulosus sensu lato ( s.l. ) cluster. It is urgent to identify novel drug targets and develop new drug candidates against CE. Glucose transporter 1 (GLUT1) is mainly responsible for the transmembrane transport of glucose to maintain its constant cellular availability and is a recent research hotspot as a drug target in various diseases. However, the role of GLUT1 in E. granulosus s.l. (EgGLUT1) was unknown. In this study, we cloned a conserved GLUT1 homology gene (named EgGLUT1-ss) from E. granulosus sensu stricto ( s.s. ) and found EgGLUT1-ss was crucial for glucose uptake and viability by the protoscoleces of E. granulosus s.s. WZB117, a GLUT1 inhibitor, inhibited glucose uptake by E. granulosus s.s. and the viability of the metacestode in vitro . In addition, WZB117 showed significant therapeutic activity in E. granulosus s.s. -infected mice: a 10 mg/kg dose of WZB117 significantly reduced the number and weight of parasite cysts ( P < 0.05) as efficiently as the reference drug, albendazole. Our results demonstrate that EgGLUT1-ss is crucial for glucose uptake by the protoscoleces of E. granulosus s.s. , and its inhibitor WZB117 has a therapeutic effect on CE.
Background:
Amide proton transfer weighted imaging (APTw), intravoxel incoherent motion (IVIM), and positron emission tomography (PET) imaging all have the potential to characterize solitary pulmonary lesions (SPLs).
Purpose:
To compare APTw and IVIM with PET imaging for distinguishing between benign and malignant SPLs and their subtypes.
Study type:
Prospective.
Population:
Ninety-five patients, 78 with malignant SPLs (including 48 with adenocarcinoma [AC] and 17 with squamous cell carcinoma [SCC]), and 17 with benign SPLs.
Field strength/sequence:
Fast spin-echo (FSE) T2WI, FSE APTw and echo-planar imaging IVIM, MR-base attenuation correction (MRAC), and PET imaging on a 3-T whole-body PET/MR system.
Assessment:
The magnetization transfer ratio asymmetry (MTRasym) at 3.5 ppm, diffusion coefficient (D), pseudo diffusion coefficient (D*), perfusion fraction (f), and the maximum standardized uptake value (SUVmax) were analyzed.
Statistical tests:
Individual sample t-test, Delong test, Pearson's correlation analysis, and area under the receiver operating characteristic curve (AUC). P < 0.05 indicated statistical significance.
Results:
The MTRasym and SUVmax were significantly higher, and D was significantly lower in the malignant group (3.3 ± 2.6 [%], 7.8 ± 5, and 1.2 ± 0.3 [×10-3 mm2 /second]) compared to the benign group (-0.3 ± 1.6 [%], 3.1 ± 3.8, and 1.6 ± 0.3 [×10-3 mm2 /second]). The MTRasym and D were significantly lower, and SUVmax was significantly higher in the SCC group (0.8 ± 1.0 [%], 1.0 ± 0.2 [×10-3 mm2 /second] than in the AC group (4.1 ± 2.6 [%], 1.3 ± 0.3 [×10-3 mm2 /second], 6.7 ± 4.6). Besides, the combination (AUC = 0.964) of these three methods showed higher diagnostic efficacy than any individual method (AUC = 0.917, 0.851, 0.82, respectively) in identifying malignant and benign SPLs. However, APTw showed better diagnostic efficacy than the combination of three methods or PET imaging alone in distinguishing SCC and AC groups (AUC = 0.934, 0.781, 0.725, respectively).
Data conclusion:
APTw, IVIM, and PET imaging are all effective methods to distinguish benign and malignant SPLs and their subtypes. APTw is potentially more capable than PET imaging of distinguishing lung SCC from AC.
Level of evidence:
2 TECHNICAL EFFICACY: Stage 2.
Metabolic reprogramming in cancer necessitates increased amino acid uptake, which is accomplished by upregulation of specific amino acid transporters. However, not all tumors rely on any single amino acid transporter for this purpose. Here we report on the differential upregulation of the amino acid transporter SLC38A5 in triple-negative breast cancer (TNBC). The upregulation is evident in TNBC tumors, conventional and patient-derived xenograft TNBC cell lines, and a mouse model of spontaneous TNBC mammary tumor. The upregulation is confirmed by functional assays. SLC38A5 is an amino acid-dependent Na+/H+ exchanger which transports Na+ and amino acids into cells coupled with H+ efflux. Since cell-surface Na+/H+ exchanger is an established inducer of macropinocytosis, an endocytic process for cellular uptake of bulk fluid and its components, we examined the impact of SLC38A5 on macropinocytosis in TNBC cells. We found that the transport function of SLC38A5 is coupled to induction of macropinocytosis. Surprisingly, the transport function of SLC38A5 is inhibited by amilorides, the well-known inhibitors of Na+/H+ exchanger. Downregulation of SLC38A5 in TNBC cells attenuates serine-induced macropinocytosis and reduces cell proliferation significantly as assessed by multiple methods, but does not induce cell death. The Cancer Genome Atlas database corroborates SLC38A5 upregulation in TNBC. This represents the first report on the selective expression of SLC38A5 in TNBC and its role as an inducer of macropinocytosis, thus revealing a novel, hitherto unsuspected, function for an amino acid transporter that goes beyond amino acid delivery but is still relevant to cancer cell nutrition and proliferation.
Cancer is a complex and dynamic disease with an outcome that depends on a strict crosstalk between tumor cells and other components in tumor microenvironment, namely, tumor-infiltrating immune cells, fibroblasts, cancer stem cells, adipocytes, and endothelial cells. Within the tumor microenvironment, macrophages and T-lymphocytes appear to be key effectors during the several steps of tumor initiation and progression. Tumor cells, through the release of a plethora of signaling molecules, can induce immune tolerance, by avoiding immune surveillance, and inhibit immune cells cytotoxic functions. Furthermore, as the tumor grows, tumor microenvironment reveals a series of dysfunctional conditions that potentiate a polarization of harmful humoral Th2 and Th17, an upregulation of Treg cells, and a differentiation of macrophages into the M2 subtype, which contribute to the activation of several signaling pathways involving important tissue biomarkers (COX-2, EGFR, VEGF) implicated in cancer aggressiveness and poor clinical outcomes. In order to maintain the tumor growth, cancer cells acquire several adaptations such as neovascularization and metabolic reprogramming. An extensive intracellular production of lactate and protons is observed in tumor cells as a result of their high glycolytic metabolism. This contributes not only for the microenvironment pH alteration but also to shape the immune response that ultimately impairs immune cells capabilities and effector functions. In this chapter, the complexity of tumor microenvironment, with special focus on macrophages, T-lymphocytes, and the impact of lactate efflux, was reviewed, always trying to demonstrate the strong similarities between data from studies of humans and dogs, a widely proposed model for comparative oncology studies.
Glucosamine (Gln) is a universal amino sugar, present in animals, plants, and microbes as a major component of various structural polymers, i.e. chitin, chitosan, and peptidoglycans. Besides structural applications, several experimental and clinical trials have shown its tremendous applications in the health sector especially in the case of cancer, osteoarthritis (OA), allergy, and inflammation-associated complications. Wider and safer applications for Gln increased its volumetric demand and market share exponentially. Different methods of Gln production such as direct extraction from sea waste (chitin) and plants biomass have been reported. However, phytoextraction and chemical hydrolysis of chitinous wastes are suffered from limited yield and high production costs. In recent years, microbial/enzymatic hydrolysis has taken over the market due to economic and eco-friendly production. In addition, genetically modified bacterial fermentation is also under development stage. In this review, recent progress related to Gln production from different resources and its various applications is reviewed in detail.
PEPT1 is a proton-coupled peptide transporter that is upregulated in PDAC cell lines and PDXs, with little expression in normal pancreas. However, the relevance of this upregulation to cancer progression and the mechanism of upregulation have not been investigated. Herein, we show that PEPT1 is not just upregulated in a large panel of PDAC cell lines and PDXs but is also functional and transport-competent. PEPT2, another proton-coupled peptide transporter, is also overexpressed in PDAC cell lines and PDXs, but is not functional due to its intracellular localization. Using glibenclamide as a pharmacological inhibitor of PEPT1, we demonstrate in cell lines in vitro and mouse xenografts in vivothat inhibition of PEPT1 reduces the proliferation of the cancer cells. These findings are supported by genetic knockdown of PEPT1 with shRNA, wherein the absence of the transporter significantly attenuates the growth of cancer cells, both in vitro and in vivo, suggesting that PEPT1 is critical for the survival of cancer cells. We also establish that the tumor-derived lactic acid (Warburg effect) in the tumor microenvironment supports the transport function of PEPT1 in the maintenance of amino acid nutrition in cancer cells by inducing MMPs and DPPIV to generate peptide substrates for PEPT1 and by generating a H+ gradient across the plasma membrane to energize PEPT1. Taken collectively, these studies demonstrate a functional link between PEPT1 and extracellular protein breakdown in the tumor microenvironment as a key determinant of pancreatic cancer growth, thus identifying PEPT1 as a potential therapeutic target for PDAC.
Immune escape is one of the hallmarks of cancer. While metabolic reprogramming provides survival advantage to tumor cancer cells, accumulating data also suggest such metabolic rewiring directly affects the activation, differentiation and function of immune cells, particularly in the tumor microenvironment. Understanding how metabolic reprogramming affects both tumor and immune cells, as well as their interplay, is therefore critical to better modulate tumor immune microenvironment in the era of cancer immunotherapy. In this review, we discuss alterations in several essential metabolic pathways in both tumor and key immune cells, provide evidence on their dynamic interaction, and propose innovative strategies to improve cancer immunotherapy via the modulation of metabolic pathways.
An enhanced requirement for nutrients is a hallmark property of cancer cells. Here, we optimized an in vivo genetic screening strategy in acute myeloid leukemia (AML), which led to the identification of the myo-inositol transporter SLC5A3 as a dependency in this disease. We demonstrate that SLC5A3 is essential to support a myo-inositol auxotrophy in AML. The commonality among SLC5A3-dependent AML lines is the transcriptional silencing of ISYNA1, which encodes the rate-limiting enzyme for myo-inositol biosynthesis, inositol-3-phosphate synthase 1. We use gain- and loss-of-function experiments to reveal a synthetic lethal genetic interaction between ISYNA1 and SLC5A3 in AML, which function redundantly to sustain intracellular myo-inositol. Transcriptional silencing and DNA hypermethylation of ISYNA1 occur in a recurrent manner in human AML patient samples, in association with IDH1/IDH2 and CEBPA mutations. Our findings reveal myo-inositol as a nutrient dependency in AML caused by the aberrant silencing of a biosynthetic enzyme.
Significance
We show how epigenetic silencing can provoke a nutrient dependency in AML by exploiting a synthetic lethality relationship between biosynthesis and transport of myo-inositol. Blocking the function of this solute carrier may have therapeutic potential in an epigenetically defined subset of AML.
This article is highlighted in the In This Issue feature, p. 275
Background
Glycolysis affects tumor growth, invasion, chemotherapy resistance, and the tumor microenvironment. In this study, we aimed to construct a glycolysis-related prognostic model for ovarian cancer and analyze its relationship with the tumor microenvironment’s immune cell infiltration.
Methods
We obtained six glycolysis-related gene sets for gene set enrichment analysis (GSEA). Ovarian cancer data from The Cancer Genome Atlas (TCGA) database and two Gene Expression Omnibus (GEO) datasets were divided into two groups after removing batch effects. We compared the tumor environments' immune components in high-risk and low-risk groups and analyzed the correlation between glycolysis- and immune-related genes. Then, we generated and validated a predictive model for the prognosis of ovarian cancer using the glycolysis-related genes.
Results
Overall, 27/329 glycolytic genes were associated with survival in ovarian cancer, 8 of which showed predictive value. The tumor cell components in the tumor microenvironment did not differ between the high-risk and low-risk groups; however, the immune score differed significantly between groups. In total, 13/24 immune cell types differed between groups, including 10 T cell types and three other immune cell types. Eight glycolysis-related prognostic genes were related to the expression of multiple immune-related genes at varying degrees, suggesting a relationship between glycolysis and immune response.
Conclusions
We identified eight glycolysis-related prognostic genes that effectively predicted survival in ovarian cancer. To a certain extent, the newly identified gene signature was related to the tumor microenvironment, especially immune cell infiltration and immune-related gene expression. These findings provide potential biomarkers and therapeutic targets for ovarian cancer.
Background
Lung branching morphogenesis is characterized by epithelial-mesenchymal interactions that ultimately define the airway conducting system. Throughout this process, energy and structural macromolecules are necessary to sustain the high proliferative rates. The extensive knowledge of the molecular mechanisms underlying pulmonary development contrasts with the lack of data regarding the embryonic lung metabolic requirements. Here, we studied the metabolic profile associated with the early stages of chicken pulmonary branching.
Methods
In this study, we used an ex vivo lung explant culture system and analyzed the consumption/production of extracellular metabolic intermediates associated with glucose catabolism (alanine, lactate, and acetate) by ¹ H-NMR spectroscopy in the culture medium. Then, we characterized the transcript levels of metabolite membrane transporters ( glut1 , glut3 , glut8 , mct1 , mct3 , mct4 , and mct8 ) and glycolytic enzymes ( hk1 , hk2 , pfk1 , ldha , ldhb , pdha , and pdhb ) by qPCR. ldha and ldhb mRNA spatial localization was determined by in situ hybridization. Proliferation was analyzed by directly assessing DNA synthesis using an EdU-based assay. Additionally, we performed western blot to analyze LDHA and LDHT protein levels. Finally, we used a Clark-Type Electrode to assess the lung explant's respiratory capacity.
Results
Glucose consumption decreases, whereas alanine, lactate, and acetate production progressively increase as branching morphogenesis proceeds. mRNA analysis revealed variations in the expression levels of key enzymes and transporters from the glycolytic pathway. ldha and ldhb displayed a compartment-specific expression pattern that resembles proximal–distal markers. In addition, high proliferation levels were detected at active branching sites. LDH protein expression levels suggest that LDHB may account for the progressive rise in lactate. Concurrently, there is a stable oxygen consumption rate throughout branching morphogenesis.
Conclusions
This report describes the temporal metabolic changes that accompany the early stages of chicken lung branching morphogenesis. Overall, the embryonic chicken lung seems to shift to a glycolytic lactate-based metabolism as pulmonary branching occurs. Moreover, this metabolic rewiring might play a crucial role during lung development.
Cancer cells need a constant supply of nutrients. SLC6A14, an amino acid transporter B0,+ (ATB0,+) that is upregulated in many cancers, transports all but acidic amino acids. In its exit from the endoplasmic reticulum (ER), it is recognized by the SEC24C subunit of coatomer II (COPII) for further vesicular trafficking to the plasma membrane. SEC24C has previously been shown to be phosphorylated by protein kinase B/AKT, which is hyper-activated in cancer; therefore, we analyzed the influence of AKT on SLC6A14 trafficking to the cell surface. Studies on overexpressed and endogenous transporters in the breast cancer cell line MCF-7 showed that AKT inhibition with MK-2206 correlated with a transient increase of the transporter in the plasma membrane, not resulting from the inhibition of ER-associated protein degradation. Two-dimensional electrophoresis demonstrated the decreased phosphorylation of SLC6A14 and SEC24C upon AKT inhibition. A proximity ligation assay confirmed this conclusion: AKT inhibition is correlated with decreased SLC6A14 phosphothreonine and SEC24C phosphoserine. Augmented levels of SLC6A14 in plasma membrane led to increased leucine transport. These results show that the inactivation of AKT can rescue amino acid delivery through SLC6A14 trafficking to the cell surface, supporting cancer cell survival. The regulation of the ER export of the amino acid transporter seems to be a novel function of AKT.
Reprogramed cellular metabolism is one of the most significant hallmarks of cancer. All cancer cells exhibit increased demand for specific amino acids, and become dependent on either an exogenous supply or upregulated de novo synthesis. The resultant enhanced availability of amino acids supports the reprogramed metabolic pathways and fuels the malignant growth and metastasis of cancers by providing energy and critical metabolic intermediates, facilitating anabolism, and activating signaling networks related to cell proliferation and growth. Therefore, pharmacologic blockade of amino acid entry into cancer cells is likely to have a detrimental effect on cancer cell growth. Here we developed a nanoplatform ([email protected]) to therapeutically target two transporters, SLC6A14 (ATB0,+) and SLC7A5 (LAT1), that are known to be essential for the sustenance of amino acid metabolism in most cancers. The [email protected] uses tryptophan to guide SLC6A14-targeted delivery of JPH203, a high-affinity inhibitor of SLC7A5. In the process, SLC6A14 is also down-regulated. We tested the ability of this strategy to synergize with the anticancer efficacy of lapatinib, an inhibitor of EGFR/HER1/HER2-assocated kinase. These studies show that blockade of amino acid entry amplifies the anticancer effect of lapatinib via interference with mTOR signaling, promotion of apoptosis, and suppression of cell proliferation and metastasis. This represents the first study to evaluate the impact of amino acid starvation on the anticancer efficacy of widely used kinase inhibitor.
Amino acid (AA) metabolism plays an important role in many cellular processes including energy production, immune function, and purine and pyrimidine synthesis. Cancer cells therefore require increased AA uptake and undergo metabolic reprogramming to satisfy the energy demand associated with their rapid proliferation. Like many other cancers, myeloid leukemias are vulnerable to specific therapeutic strategies targeting metabolic dependencies. Herein, our review provides a comprehensive overview and TCGA data analysis of biosynthetic enzymes required for non-essential AA synthesis and their dysregulation in myeloid leukemias. Furthermore, we discuss the role of the general control nonderepressible 2 (GCN2) and-mammalian target of rapamycin (mTOR) pathways of AA sensing on metabolic vulnerability and drug resistance.
Serine threonine tyrosine kinase 1 (STYK1)/novel oncogene with kinase domain (NOK) has been demonstrated to promote cell carcinogenesis and tumorigenesis, as well as to strengthen cellular aerobic glycolysis, which is considered to be a defining hallmark of cancer. As the carriers of glucose into cells, glucose transporters (GLUTs) are important participants in cellular glucose metabolism and even tumorigenesis. However, to the best of our knowledge, the role of GLUTs in biological events caused by STYK1/NOK has not yet been reported. The present study assessed GLUT3 as a key transporter, and glucose consumption and lactate production assays revealed that downregulation of GLUT3 impaired STYK1/NOK-induced augmented glucose uptake and lactate production, and RT-qPCR and western blotting confirmed that GLUT3 knockdown attenuated the STYK1/NOK-induced increase in the expression levels of key enzymes implicated in glycolysis. Furthermore, MTT and Transwell assays demonstrated that STYK1/NOK-triggered cell proliferation and migration were also markedly decreased following knockdown of GLUT3. To the best of our knowledge, the present study is the first to demonstrate that GLUT3 serves a prominent role in STYK1/NOK-driven aerobic glycolysis and cell proliferation characteristics. These findings may provide a clue for the investigation of the oncogenic activity of STYK1/NOK and for the identification of potential tumor therapy targets associated with GLUT3.
The glutaminolysis and serine-glycine-one-carbon pathways represent metabolic reactions that are reprogramed and upregulated in cancer; these pathways are involved in supporting the growth and proliferation of cancer cells. Glutaminolysis participates in the production of lactate, an oncometabolite, and also in anabolic reactions leading to the synthesis of fatty acids and cholesterol. The serine-glycine-one-carbon pathway is involved in the synthesis of purines and pyrimidines and the control of the epigenetic signature (DNA methylation, histone methylation) in cancer cells. Methionine is obligatory for most of the methyl-transfer reactions in the form of S-adenosylmethionine; here, too, the serine-glycine-one-carbon pathway is necessary for the resynthesis of methionine following the methyl-transfer reaction. Glutamine, serine, glycine, and methionine are obligatory to fuel these metabolic pathways. The first three amino acids can be synthesized endogenously to some extent, but the need for these amino acids in cancer cells is so high that they also have to be acquired from extracellular sources. Methionine is an essential amino acid, thus making it necessary for cancer cells to acquire this amino acid solely from the extracellular milieu. Cancer cells upregulate specific amino acid transporters to meet this increased demand for these four amino acids. SLC6A14 and SLC38A5 are the two transporters that are upregulated in a variety of cancers to mediate the influx of glutamine, serine, glycine, and methionine into cancer cells. SLC6A14 is a Na+/Cl- -coupled transporter for multiple amino acids, including these four amino acids. In contrast, SLC38A5 is a Na+-coupled transporter with rather restricted specificity towards glutamine, serine, glycine, and methionine. Both transporters exhibit unique functional features that are ideal for the rapid proliferation of cancer cells. As such, these two amino acid transporters play a critical role in promoting the survival and growth of cancer cells and hence represent novel, hitherto largely unexplored, targets for cancer therapy.
Cervical cancer is responsible for around 5% of all human cancers worldwide. It develops almost exclusively from an unsolved, persistent infection of the squamocolumnar transformation zone between the endo- and ecto-cervix with various high-risk (HR) human papillomaviruses (HPVs). The decisive turning point on the way to persistent HPV infection and malignant transformation is an immune system weakened by pathobionts and oxidative stress and an injury to the cervical mucosa, often caused by sexual activities. Through these injury and healing processes, HPV viruses, hijacking activated keratinocytes, move into the basal layers of the cervical epithelium and then continue their development towards the distal prickle cell layer (Stratum spinosum). The microbial microenvironment of the cervical tissue determines the tissue homeostasis and the integrity of the protective mucous layer through the maintenance of a healthy immune and metabolic signalling. Pathological microorganisms and the resulting dysbiosis disturb this signalling. Thus, pathological inflammatory reactions occur, which manifest the HPV infection. About 90% of all women contract an HPV infection in the course of their lives. In about 10% of cases, the virus persists and cervical intra-epithelial neoplasia (CIN) develops. Approximately 1% of women with a high-risk HPV infection incur a cervical carcinoma after 10 to 20 years. In this non-systematic review article, we summarise how the sexually and microbial mediated pathogenesis of the cervix proceeds through aberrant immune and metabolism signalling via CIN to cervical carcinoma. We show how both the virus and the cancer benefit from the same changes in the immune and metabolic environment.
Most cancer cells employ overexpression of glucose transports (GLUTs) to satisfy glucose demand ("Sweet Tooth") for increased aerobic glycolysis rates. GLUT1, one of the most widely expressed GLUTs in numerous cancers, was identified as a prognosis-related biomarker of gastric cancer via tissue array analysis. Herein, a "Sweet Tooth"-oriented SN38 prodrug delivery nanoplatform (Glu-SNP) was developed for targeted gastric cancer therapy. For this purpose, a SN38-derived prodrug (PLA-SN38) was synthesized by tethering 7-ethyl-10-hydroxycamptothecin (SN38) to biocompatible polylactic acid (PLA) with the appropriate degree of polymerization (n = 44). The PLA-SN38 conjugate was further assembled with glycosylated amphiphilic lipid to obtain glucosamine-decorated nanoparticles (Glu-SNP). Glu-SNP exhibited potent antitumor efficiency both in vitro and in vivo through enhanced cancer cell-specific targeting associated with the overexpression of GLUT1, which provides a promising approach for gastric cancer therapy.
A glucose-based vector for targeting cancer cells conjugated to a tris(methylpyridyl)amine (tpa) ligand to generate targeted chaperone and caging complexes for active anticancer agents is described. The ligand, tpa(CONHPEGglucose)1, inhibits hexokinase, suggesting that it will be phosphorylated in the cell. A Co(III) complex incorporating this ligand and coumarin-343 hydroximate (C343ha), [Co(C343ha){tpa(CONHPEGglucose)1}]Cl, is shown to exhibit glucose-dependent cellular accumulation in DLD-1 colon cancer cells. Cellular accumulation of [Co(C343ha){tpa(CONHPEGglucose)1}]+ is slower than for the glucose null and glucosamine analogues, and the glucose complex also exhibits a lower ability to inhibit antiproliferative activity. Distributions of cobalt (X-ray fluorescence) and C343ha (visible light fluorescence) in DLD-1 cancer cell spheroids are consistent with uptake of [Co(C343ha){tpa(CONHPEGglucose)1}]+ by rapidly dividing cells, followed by release and efflux of C343ha and trapping of the Co{tpa(CONHPEGglucose)1} moiety. The Co{tpa(CONHPEGglucose)1} moiety is shown to have potential for the caged and targeted delivery of highly toxic anticancer agents.
The development and progression of colorectal cancer (CRC) have been associated with genetic and epigenetic alterations and more recently with changes in cell metabolism. Amino acid transporters are key players in tumor development, and it is described that tumor cells upregulate some AA transporters in order to support the increased amino acid (AA) intake to sustain the tumor additional needs for tumor growth and proliferation through the activation of several signaling pathways. LAT1 and ASCT2 are two AA transporters involved in the regulation of the mTOR pathway that has been reported as upregulated in CRC. Some attempts have been made in order to develop therapeutic approaches to target these AA transporters, however none have reached the clinical setting so far. MiRNA-based therapies have been gaining increasing attention from pharmaceutical companies and now several miRNA-based drugs are currently in clinical trials with promising results. In this review we combine a bioinformatic approach with a literature review in order to identify a miRNA profile with the potential to target both LAT1 and ASCT2 with potential to be used as a therapeutic approach against CRC.
The positron emission tomography (PET) molecular imaging technique has gained its universal value as a remarkable tool for medical diagnosis and biomedical research. Carbon-11 is one of the promising radiotracers that can report target-specific information related to its pharmacology and physiology to understand the disease status. Currently, many of the available carbon-11 (t1/2 = 20.4 min) PET radiotracers are heterocyclic derivatives that have been synthesized using carbon-11 inserted different functional groups obtained from primary and secondary carbon-11 precursors. A spectrum of carbon-11 PET radiotracers has been developed against many of the upregulated and emerging targets for the diagnosis, prognosis, prediction, and therapy in the fields of oncology, cardiology, and neurology. This review focuses on the carbon-11 radiochemistry and various target-specific PET molecular imaging agents used in tumor, heart, brain, and neuroinflammatory disease imaging along with its associated pathology.
As with most pharmaceutical developments, the vast majority of nuclear medicine procedures that are applied to pediatric disorders and investigations utilize radiopharmaceuticals and imaging methods developed for adults. This chapter summarizes the well‐established or clinically approved nuclear medicine procedures but focuses on studies more commonly performed in pediatric populations. It also summarizes the initial development of key radiopharmaceuticals used in pediatric imaging and therapy, citing relevant previous reviews, and present representative studies and applications in young populations. Many radiotracers, labeled with either single‐photon‐ or positron‐emitting radionuclides, have been used to probe a wide variety of pediatric disorders. In pediatric oncology, [¹⁸F]fluoro‐2‐deoxy‐d‐glucose positron emission tomography/computed tomography is frequently used in the management of disease, particularly in the selection of biopsy sites, tumor staging, risk stratification, radiation treatment planning, monitoring of response to treatment, and surveillance of tumors.
This chapter enlightens the development and application of dye-based as well as genetically encoded nanosensors for metabolites related to various diseases, their comparison, and usefulness in understanding the metabolic pathways by using novel high throughput assays. Previously dye-based fluorescent probes have been constructed for the in vivo detection of cellular molecules. To study the real-time localization and monitoring of metabolites in live cells, genetically encoded fluorescent nanosensors are steadily used as an elected method in comparison with chemical probes that are intrinsically dependent on effective delivery into the cell and sometimes causes the toxicity in the cells. Several methods have been reported to detect cancer and neurodegenerative diseases. However, FRET-based disease-specific biomarker detection is a preferable method because of its high sensitivity and selectivity. By using FRET-based genetically encoded nanosensor, we can detect some of the metabolites which are related to several diseases like cancer, Alzheimer, and diabetes. In humans, lack of reactive oxygen species (ROS) causes certain autoimmune diseases and excess production of ROS causes cardiovascular and respiratory diseases which can be detected in real time and non-invasively by using a nanosensor. Protein recognition-based genetically encoded sensors are projected to discover many research, medical, and environmental applications. Many human problems can be solved by the utilization of nanosensors which can also be helpful to authenticate the medicines to their principle components. Thus, sensor biology can be revolutionary for human welfare in the future with the help of FRET-based tools.
A major goal of cancer genomics is to identify all genes that play critical roles in carcinogenesis. Most approaches focused on genes positively selected for mutations that drive carcinogenesis and neglected the role of negative selection. Some studies have actually concluded that negative selection has no role in cancer evolution. We have re-examined the role of negative selection in tumor evolution through the analysis of the patterns of somatic mutations affecting the coding sequences of human genes. Our analyses have confirmed that tumor suppressor genes are positively selected for inactivating mutations, oncogenes, however, were found to display signals of both negative selection for inactivating mutations and positive selection for activating mutations. Significantly, we have identified numerous human genes that show signs of strong negative selection during tumor evolution, suggesting that their functional integrity is essential for the growth and survival of tumor cells.
The role of the amino acid transporters ASCT2 and LAT1 in cancer has been explored throughout the years. In this review, we report their impact on the hallmarks of cancer, as well as their clinical significance. Overall, both proteins have been associated with cell death resistance through dysregulation of caspases and sustainment of proliferative signaling through mTOR activation. Furthermore, ASCT2 appears to play an important role in cellular energetics regulation, whereas LAT1 expression is associated with angiogenesis and invasion and metastasis activation. The molecular impact of these proteins on the hallmarks of cancer translates into various clinical applications and both transporters have been identified as prognostic factors in many types of cancer. Concerning their role as therapeutic targets, efforts have been undertaken to synthesize competitive or irreversible ASCT2 and LAT1 inhibitors. However, JHP203, a selective inhibitor of the latter, is, to the best of our knowledge, the only compound included in a Phase 1 clinical trial. In conclusion, considering the usefulness of ASCT2 and LAT1 in a variety of cancer-related pathways and cancer therapy/diagnosis, the development and testing of novel inhibitors for these transporters that could be evaluated in clinical trials represents a promising approach to cancer prognosis improvement.
Background:
Glutamine metabolism is considered an important metabolic phenotype of proliferating tumor cells.
Objective:
The objective of this study was to investigate the expression of glutamine metabolism-related and amino acid transporter proteins in adrenal cortical neoplasms (ACNs) and pheochromocytomas (PCCs) in the adrenal gland.
Methods:
A tissue microarray was constructed for 132 cases of ACN (115 cases of adrenal cortical adenoma and 17 cases of adrenal cortical carcinoma) and 189 cases of PCC. Immunohistochemical staining for glutamine metabolism-related proteins GLS1 and GDH and amino acid transporter proteins SLC1A5, SLC7A5, and SLC7A11 as well as SDHB was performed and compared with clinicopathologic parameters.
Results:
The expression levels of GLS (p < 0.001), SLC7A5 (p = 0.049), and SDHB (p = 0.007) were higher in ACN than in PCC, whereas the expression levels of SLC1A5 (p < 0.001) and SLC7A11 (p < 0.001) were higher in PCC than in ACN. In ACN, GLS positivity was associated with a higher Fuhrman grade (p = 0.009), and SLC1A5 positivity was associated with SDHB positivity (p = 0.004) and a clear cell proportion < 25% (p = 0.010). SDHB negativity was also associated with tumor cell necrosis (p = 0.007). In PCC, SLC7A11 positivity was associated with nonnorepinephrine type (p = 0.008). In Kaplan-Meier analysis, patients with GLS positivity (p = 0.039) and SDHB negativity (p = 0.005) had significantly shorter overall survival in ACN. In PCC patients with a GAPP score ≥ 3, GLS positivity (p = 0.001) and SDHB positivity (p = 0.001) were associated with shorter disease-free survival, whereas GLS positivity (p = 0.004) was also associated with shorter overall survival.
Conclusions:
The expression of glutamine metabolism-related and amino acid transporter proteins in ACN and PCC is distinct and associated with prognosis.
BACKGROUND
Cancer cell growth is an energy-related process supported by an increased glucose metabolism. The objective of this study was to investigate the association of GLUT-1 with response to chemotherapy and outcome in patients with ovarian carcinoma.METHODS
Histologic sections of formalin fixed, paraffin embedded specimens from 113 primary ovarian carcinomas were stained for GLUT-1 by using polyclonal GLUT-1 antibody (Dako Co., Carpinteria, CA) and the labeled streptavidin biotin procedure. Intensity of GLUT-1 staining was compared with disease free survival (DFS), chemotherapy response, and other clinicopathologic characteristics.RESULTSGLUT-1 cytoplasmic membrane staining was observed in 89 of 104 (85.6%) malignant tumors. Poorly differentiated tumors showed a trend to overexpress the GLUT-1 protein compared with the more differentiated counterparts (27.6% vs. 8.7%; P = 0.08). Patients who experienced a complete clinical response to chemotherapy were more frequently GLUT-1 positive than GLUT-1 negative (80% vs. 51.5%; P = 0.036). In multivariate analysis of advanced stage disease, residual tumor (P = 0.0001) and high GLUT-1 expression levels (P = 0.028) were the only independent variables that maintained a significant association with response to chemotherapy (P = 0.0001; chi-square = 38.13). In the subgroup of Stage III–IV (International Federation of Gynecology and Obstetrics patients showing a complete clinical response, GLUT-1 overexpression was associated with a shorter DFS. The median time to progression was 30 months in GLUT-1 strongly positive cases (> 50% of cancer cells positive) versus 60 months in GLUT-1 weakly positive cases (≤ 50% of cancer cells positive; P = 0.024).CONCLUSIONSGLUT-1 status is an independent prognostic factor of response to chemotherapy in advanced stage ovarian carcinoma. Moreover, patients overexpressing GLUT-1 show a significantly shorter DFS. These results suggest that the assessment of GLUT-1 status may provide clinically useful prognostic information in patients with ovarian carcinoma. Cancer 2001;92:1144–50. © 2001 American Cancer Society.
ATB(0,+) [SLC6A14 (solute carrier family 6 member 14)] is an Na(+)/Cl(-)-coupled amino acid transporter whose expression is upregulated in cancer. 1-Methyltryptophan is an inducer of immune surveillance against tumour cells through its ability to inhibit indoleamine dioxygenase. In the present study, we investigated the role of ATB(0,+) in the uptake of 1-methyltryptophan as a potential mechanism for entry of this putative anticancer drug into tumour cells. These studies show that 1-methyltryptophan is a transportable substrate for ATB(0,+). The transport process is Na(+)/Cl(-)-dependent with an Na(+)/Cl(-)/1-methyltryptophan stoichiometry of 2:1:1. Evaluation of other derivatives of tryptophan has led to identification of alpha-methyltryptophan as a blocker, not a transportable substrate, for ATB(0,+). ATB(0,+) can transport 18 of the 20 proteinogenic amino acids. alpha-Methyltryptophan blocks the transport function of ATB(0,+) with an IC(50) value of approximately 250 muM under conditions simulating normal plasma concentrations of all these 18 amino acids. These results suggest that alpha-methyltryptophan may induce amino acid deprivation in cells which depend on the transporter for their amino acid nutrition. Screening of several mammary epithelial cell lines shows that ATB(0,+) is expressed robustly in some cancer cell lines, but not in all; in contrast, non-malignant cell lines do not express the transporter. Treatment of ATB(0,+)-positive tumour cells with alpha-methyltryptophan leads to suppression of their colony-forming ability, whereas ATB(0,+)-negative cell lines are not affected. The blockade of ATB(0,+) in these cells with alpha-methyltryptophan is associated with cell cycle arrest. These studies reveal the potential of ATB(0,+) as a drug target for cancer chemotherapy.
Human colon cancer cells and primary colon cancer silence the gene coding for LDH (lactate dehydrogenase)-B and up-regulate the gene coding for LDH-A, resulting in effective conversion of pyruvate into lactate. This is associated with markedly reduced levels of pyruvate in cancer cells compared with non-malignant cells. The silencing of LDH-B in cancer cells occurs via DNA methylation, with involvement of the DNMTs (DNA methyltransferases) DNMT1 and DNMT3b. Colon cancer is also associated with the expression of pyruvate kinase M2, a splice variant with low catalytic activity. We have shown recently that pyruvate is an inhibitor of HDACs (histone deacetylases). Here we show that pyruvate is a specific inhibitor of HDAC1 and HDAC3. Lactate has no effect on any of the HDACs examined. Colon cancer cells exhibit increased HDAC activity compared with non-malignant cells. HDAC1 and HDAC3 are up-regulated in colon cancer cells and in primary colon cancer, and siRNA (small interfering RNA)-mediated silencing of HDAC1 and HDAC3 in colon cancer cells induces apoptosis. Colon cancer cells silence SLC5A8, the gene coding for a Na(+)-coupled pyruvate transporter. Heterologous expression of SLC5A8 in the human colon cancer cell line SW480 leads to inhibition of HDAC activity when cultured in the presence of pyruvate. This process is associated with an increase in intracellular levels of pyruvate, increase in the acetylation status of histone H4, and enhanced cell death. These studies show that cancer cells effectively maintain low levels of pyruvate to prevent inhibition of HDAC1/HDAC3 and thereby to evade cell death.
The type 1 glucose transporter (GLUT1) gene encodes an integral membrane glycoprotein responsible for facilitating transfer of glucose across plasma membrane and is rapidly activated by serum, growth factors, and by oncogenic transformation. To elucidate the molecular mechanisms of regulation of GLUT1 gene expression, we isolated and characterized the mouse GLUT1 gene. DNA elements regulating transcription of the gene were analyzed in transient expression assays after transfection of NIH/3T3 cells with a low background chloramphenicol acetyltransferase (CAT) vector system pSVOOCAT. We identified two enhancer elements; the first one is located 2.7 kilobases upstream of the cap site of the gene which contains the homologous sequences with two 12-O-tetradecanoylphorbol-13-acetate-responsive elements (TREs), a serum response element, a cyclic AMP-responsive element (CRE) and three GC boxes, and the second one is located in the second intron of the gene which contains the homologous sequences with two TREs and one CRE. With the promoter alone the transcription of the gene is activated by src, only slightly activated by ras and is not activated by serum and platelet-derived growth factor. When the gene is accompanied by one of these enhancers, the transcription is activated by all these stimuli.
Hypoxia-inducible factor 1 (HIF-1) is a basic helix-loop-helix transcription factor which is expressed when mammalian cells are subjected to hypoxia and which activates transcription of genes encoding erythropoietin, vascular endothelial growth factor, and other proteins that are important for maintaining oxygen homeostasis. Previous studies have provided indirect evidence that HIF-1 also regulates transcription of genes encoding glycolytic enzymes. In this paper we characterize hypoxia response elements in the promoters of the ALDA, ENO1, and Ldha genes. We demonstrate that HIF-1 plays an essential role in activating transcription via these elements and show that although absolutely necessary, the presence of a HIF-1 binding site alone is not sufficient to mediate transcriptional responses to hypoxia. Analysis of hypoxia response elements in the ENO1 and Ldha gene promoters revealed that each contains two functionally-essential HIF-1 sites arranged as direct and inverted repeats, respectively. Our data establish that functional hypoxia-response elements consist of a pair of contiguous transcription factor binding sites at least one of which contains the core sequence 5'-RCGTG-3' and is recognized by HIF-1. These results provide further evidence that the coordinate transcriptional activation of genes encoding glycolytic enzymes which occurs in hypoxic cells is mediated by HIF-1.
Analysis of rat, pre-T cell 'Nb2 lymphoma' sublines, manifesting different degrees of malignant progression, can indicate phenotypic changes potentially useful as therapeutic targets. In this study, the prolactin (cytokine)-dependent Nb2-11 and autonomous Nb2-SFJCD1 sublines were compared for in vitro thiol growth requirements. Whereas Nb2-11 culture growth depended on 2-mercaptoethanol (2-ME; 33-100 microM), Nb2-SFJCD1 cells were 2-ME-independent. This difference stemmed from differential uptake of exogenous L-cystine, critically required for proliferation. Uptake of 35S-L-cystine (10 microCi/ml; 40 microM) showed Nb2-11 cells had low cystine uptake capability; 2-ME enhanced cystine uptake to growth-sustaining levels. Nb2-SFJCD1 cells did not require 2-ME due to intrinsic, 11-fold higher cystine uptake via the x(c)- cystine/glutamate transport system. In absence of 2-ME, monosodium glutamate abrogated Nb2-SFJCD1 proliferation by specifically inhibiting cystine uptake (85% at 10 mM). Elevated glutathione (GSH) levels were not essential for growth of either line as shown with L-buthionine-(S,R)-sulfoximine (0.1-4 mM) treatment. The cyst(e)ine requirement therefore did not primarily involve maintenance of normal GSH levels, reported critical for T lymphocyte replication. These and other results suggest increased cystine uptake capability constitutes another potential step in progression of T cell cancers which is not coupled to cytokine autonomy or metastatic ability development. The x(c)- transport system apparently provides a novel target for T cell cancer therapy. Its inhibition would suppress cystine uptake by certain progressed cells, and also interfere with cystine uptake, and subsequent cysteine release, by eg macrophages, thought to have a role in cysteine delivery to lymphoid cells.
We previously reported that grading of GLUT-1 glucose transporter expression was related closely to FDG accumulation in FDG PET in human cancers. But in this strong GLUT-1 expression group, there was an enormous range of standardized uptake values (SUVs) within them.
To evaluate other factors determining the FDG PET uptake, FDG PET was performed in 36 preoperative patients (mean age 62.0 yr) suspected of having pancreatic tumors, including 33 malignant and 3 benign neoplastic tumors. FDG uptake at 50 min after injection of 185 MBq 18F-FDG with > 5 hr fasting condition was semiquantitatively analyzed as SUVs. The GLUT-1 expression was studied by immunohistochemistry of paraffin sections from these tumors after the operation using the antiGLUT-1 antibody. The number of tumor cells within a 5- x 5-mm square was counted manually using x200 magnification photographs and was graded immunohistochemically as strong, weak or negative.
In all 36 cases there were 3 cases of GLUT-1 negative, 8 of GLUT-1 weak positive and 25 of GLUT-1 strong positive. In all cases, the total number of tumor cells had no significant value for SUVs. Among 33 GLUT-1 positive cases, the number of GLUT-1 positive tumor cells correlated significantly with SUVs (p < 0.01). Only in 25 strong grade cases, the number of GLUT-1 strong positive tumor cells had a more significant value for SUVs (p < 0.005). Computational multivariate analysis using multiple regression for SUVs was performed evaluating the five variables as follows: tumor size, GLUT-1 immunohistochemical grading, number of total tumor cells, number of total GLUT-1 positive tumor cells and number of GLUT-1 strong positive cells. This analysis revealed that only the variable, the number of GLUT-1 strong positive cells, had a significant regression coefficient for SUVs (standard regression coefficient = 0.855, p < 0.0001).
These data indicate that GLUT-1 expression plays an essential role in higher FDG accumulation in pancreatic tumor FDG PET, and the cellularity has a significant influence on SUVs only in the condition of GLUT-1 strong positive expression.
We used mouse hepatoma (Hepa1c1c7) cells to study the role of the serine/threonine kinase Akt in the induction of GLUT1 gene expression. In order to selectively turn on the Akt kinase cascade, we expressed a hydroxytamoxifen-regulatable form of Akt (myristoylated Akt1 estrogen receptor chimera (MER-Akt1)) in the Hepa1c1c7 cells; we verified that hydroxytamoxifen stimulates MER-Akt1 activity to a similar extent as the activation of endogenous Akt by insulin. Our studies reveal that stimulation of MER-Akt1 by hydroxytamoxifen induces GLUT1 mRNA and protein accumulation to levels comparable to that induced by insulin; therefore, activation of the Akt cascade suffices to induce GLUT1 gene expression in this cell system. Furthermore, expression of a kinase-inactive Akt mutant partially inhibits the response of the GLUT1 gene to insulin. Additional studies reveal that the induction of GLUT1 mRNA by Akt and by insulin reflects increased mRNA synthesis and not decreased mRNA degradation. Our findings imply that the GLUT1 gene responds to insulin at the transcriptional level and that Akt mediates a step in the activation of GLUT1 gene expression in this system.
A Na(+)-dependent neutral and cationic amino acid transport system (B(0+)) plays an important role in many cells and tissues; however, the molecular basis for this transport system is still unknown. To identify new transporters, the expressed sequence tag database was queried, and cDNA fragments with sequence similarity to the Na(+)/Cl(-)-dependent neurotransmitter transporter family were identified. Based on these sequences, rapid amplification of cDNA ends of human mammary gland cDNA was used to obtain a cDNA of 4.5 kilobases (kb). The open reading frame encodes a 642-amino acid protein named amino acid transporter B(0+). Human ATB(0+) (hATB(0+)) is a novel member of the Na(+)/Cl(-)-dependent neurotransmitter transporter family with the highest sequence similarity to the glycine and proline transporters. Northern blot analysis identified transcripts of approximately 4.5 kb and approximately 2 kb in the lung. Another tissue survey suggests expression in the trachea, salivary gland, mammary gland, stomach, and pituitary gland. Electrophysiology and radiolabeled amino acid uptake measurements were used to functionally characterize the transporter expressed in Xenopus oocytes. hATB(0+) was found to transport both neutral and cationic amino acids, with the highest affinity for hydrophobic amino acids and the lowest affinity for proline. Amino acid transport was Na(+) and Cl(-)-dependent and was attenuated in the presence of 2-aminobicyclo-[2.2.1]-heptane-2-carboxylic acid, a system B(0+) inhibitor. These characteristics are consistent with system B(0+) amino acid transport. Thus, hATB(0+) is the first cloned B(0+) amino acid transporter.
Resistant starch (RS) is starch and products of its small intestinal digestion that enter the large bowel. It occurs for various reasons including chemical structure, cooking of food, chemical modification, and food mastication. Human colonic bacteria ferment RS and nonstarch polysaccharides (NSP; major components of dietary fiber) to short-chain fatty acids (SCFA), mainly acetate, propionate, and butyrate. SCFA stimulate colonic blood flow and fluid and electrolyte uptake. Butyrate is a preferred substrate for colonocytes and appears to promote a normal phenotype in these cells. Fermentation of some RS types favors butyrate production. Measurement of colonic fermentation in humans is difficult, and indirect measures (e.g., fecal samples) or animal models have been used. Of the latter, rodents appear to be of limited value, and pigs or dogs are preferable. RS is less effective than NSP in stool bulking, but epidemiological data suggest that it is more protective against colorectal cancer, possibly via butyrate. RS is a prebiotic, but knowledge of its other interactions with the microflora is limited. The contribution of RS to fermentation and colonic physiology seems to be greater than that of NSP. However, the lack of a generally accepted analytical procedure that accommodates the major influences on RS means this is yet to be established.
Cancer cell growth is an energy-related process supported by an increased glucose metabolism. The objective of this study was to investigate the association of GLUT-1 with response to chemotherapy and outcome in patients with ovarian carcinoma.
Histologic sections of formalin fixed, paraffin embedded specimens from 113 primary ovarian carcinomas were stained for GLUT-1 by using polyclonal GLUT-1 antibody (Dako Co., Carpinteria, CA) and the labeled streptavidin biotin procedure. Intensity of GLUT-1 staining was compared with disease free survival (DFS), chemotherapy response, and other clinicopathologic characteristics.
GLUT-1 cytoplasmic membrane staining was observed in 89 of 104 (85.6%) malignant tumors. Poorly differentiated tumors showed a trend to overexpress the GLUT-1 protein compared with the more differentiated counterparts (27.6% vs. 8.7%; P = 0.08). Patients who experienced a complete clinical response to chemotherapy were more frequently GLUT-1 positive than GLUT-1 negative (80% vs. 51.5%; P = 0.036). In multivariate analysis of advanced stage disease, residual tumor (P = 0.0001) and high GLUT-1 expression levels (P = 0.028) were the only independent variables that maintained a significant association with response to chemotherapy (P = 0.0001; chi-square = 38.13). In the subgroup of Stage III-IV (International Federation of Gynecology and Obstetrics patients showing a complete clinical response, GLUT-1 overexpression was associated with a shorter DFS. The median time to progression was 30 months in GLUT-1 strongly positive cases (> 50% of cancer cells positive) versus 60 months in GLUT-1 weakly positive cases (< or = 50% of cancer cells positive; P = 0.024).
GLUT-1 status is an independent prognostic factor of response to chemotherapy in advanced stage ovarian carcinoma. Moreover, patients overexpressing GLUT-1 show a significantly shorter DFS. These results suggest that the assessment of GLUT-1 status may provide clinically useful prognostic information in patients with ovarian carcinoma.
Increased glucose transport is a common characteristic of most tumors. To examine the role of elevated glucose uptake in lung cancer, we performed PCR amplification of 2 facilitative glucose transporter genes (GLUT1 and GLUT3) and immunohistochemical staining for GLUT1, proliferating cell nuclear antigen (PCNA), and sialyl Lewis × (sLex) on tumor specimens from 327 patients with lung cancer who underwent surgical resection from 1980 to 1993. To evaluate the relationship between GLUT, α‐2,3‐sialyltransferase (ST), and α‐1,3‐fucosyltransferase (Fuc‐T) genes, PCR amplification of the ST3N and Fuc‐TVII also was performed. Amplification of GLUT1 was significantly greater than that of GLUT3. GLUT1 and GLUT3 amplification correlated with PCNA staining (p < 0.01). In addition, GLUT1 amplification correlated with the grading of sLex staining as well as with the grading of GLUT1 staining (p < .03, p < 0.01). GLUT1 was co‐amplified with ST3N and Fuc‐TVII genes, which are involved in the synthesis of sLex (p < 0.01). The survival of patients whose tumors showed GLUT1 amplification was significantly shorter than that of patients whose tumors did not (p < 0.01). In a multivariate analysis of survival, GLUT1 remained a statistically significant prognostic factor. Our results suggest that GLUT1 amplification may participate in sLex synthesis as well as in proliferation, and may be of prognostic value in lung cancer. Int. J. Cancer74:189–192, 1997. © 1997 Wiley‐Liss, Inc.
Amino acids perform a variety of functions in mammalian cells. They not only serve as the building blocks for protein synthesis but also play additional important roles in neurotransmission, production and storage of metabolic energy, nitrogen metabolism, and synthesis of hormones, purine and pyrimidine nucleotides, and glutathione. The intracellular pool of amino acids is derived not only from endogenous production (biosynthesis as well as protein degradation) but also from transfer across the plasma membrane. Due to their ionic nature, amino acids do not permeate biological membranes by diffusion to any great extent. Transfer across biological membranes therefore requires participation of specific transport proteins. This transfer process does not necessarily have to serve only the entry of amino acids into cells. This process also mediates the exit of certain amino acids in a tissue-specific manner. In a number of cases, the transfer involves exchange across the membrane, a process that couples the entry of certain amino acids into cells and the exit of different amino acids from the cells. A multitude of transport systems is known to facilitate the transfer of amino acids across the plasma membrane in mammalian cells. These transport systems exhibit variable but in many instances overlapping substrate specificity and are not expressed uniformly in all cell types. The differential expression of amino acid transport systems coupled with their variable substrate specificity has physiological significance because different tissue types vary markedly in their requirements for specific amino acids to suit their specific physiological functions. In addition to the differences in substrate specificity, the amino acid transport systems also differ in their energetics and hence in their transport mechanism. Some transport systems are passive, functioning either as uniporters or exchangers without involving any driving force, whereas others are active, driven by transmembrane ion gradients. In many cases, multiple ion gradients are involved in the energization process, thus enhancing the concentrative capacity of the transport systems.
Redox state is a term used widely in the research field of free radicals and oxidative stress. Unfortunately, it is used as a general term referring to relative changes that are not well defined or quantitated. In this review we provide a definition for the redox environment of biological fluids, cell organelles, cells, or tissue. We illustrate how the reduction potential of various redox couples can be estimated with the Nernst equation and show how pH and the concentrations of the species comprising different redox couples influence the reduction potential. We discuss how the redox state of the glutathione disulfide-glutathione couple (GSSG/2GSH) can serve as an important indicator of redox environment. There are many redox couples in a cell that work together to maintain the redox environment; the GSSG/2GSH couple is the most abundant redox couple in a cell. Changes of the half-cell reduction potential (E(hc)) of the GSSG/2GSH couple appear to correlate with the biological status of the cell: proliferation E(hc) approximately -240 mV; differentiation E(hc) approximately -200 mV; or apoptosis E(hc) approximately -170 mV. These estimates can be used to more fully understand the redox biochemistry that results from oxidative stress. These are the first steps toward a new quantitative biology, which hopefully will provide a rationale and understanding of the cellular mechanisms associated with cell growth and development, signaling, and reductive or oxidative stress.
Cancer cells have increased amino acid transport and retention. Our aim was to determine whether the neutral amino acid transporter ASCT2 is expressed in colorectal adenocarcinomas (CRC), and whether its expression is of biological Significance. Western blot analysis was performed on 4 cases of CRC. Immunohistochemical staining was performed on sections of 63 CRC, and the percent of positive cells was determined Survival analysis was performed using the Kaplan-Meier method. Western blot analysis showed a 50 kDa protein, corresponding to ASCT2, in 3 of the 4 CRC samples. Immunohistostaining showed lack of ASCT2 immunoreactivity in 41% of the cases. ASCT2 immunoreactivity was present in 1-25% of the cancer cells in 24% of the cases, 26-50% of the cells in 13%, and in >50% of the cells in 22% of the cases. Patient survival decreased with increased percentage of ASCT2-positive cancer cells (p = 0.0002). These results idicate that ASCT2 is expressed in a significant number of CRC, and that ASCT2 expression is associated with aggressive biological behavior. Larger studies with multivariate analysis are needed to determine whether ASCT2 expression is an independent prognostic marker in CRC.
slc5a8 and slc5a12 represent the high affinity and low affinity Na+/lactate co-transporters, respectively, in the kidney. Here we show that these transporters are expressed in the apical membrane
of the proximal tubular cells in mouse kidney, indicating that these transporters are likely to mediate the first step in
the renal reabsorption of lactate. Interestingly, the renal expression of both transporters is almost completely ablated in
mice homozygous for the deletion of the transcription factor c/ebpδ. This effect is tissue-specific since the expression of
the transporters is not affected in non-renal tissues. The functional role of C/EBPδ in the expression of SLC5A8 and SLC5A12 is demonstrable in HEK293 cells in reporter assays using gene-specific promoters. The ablation of the transporters in the
kidney is accompanied by a marked increase in urinary excretion of lactate as well as a decrease in blood levels of lactate
in c/ebpδ–/– mice. These data provide evidence for an obligatory role for slc5a8 and slc5a12 in the renal absorption of lactate. In addition,
we show that urinary excretion of urate is significantly elevated in c/ebpδ–/– mice even though the expression of URAT1, the transporter responsible for the apical membrane uptake of urate in renal proximal
tubule, is not altered. These data provide in vivo evidence for the functional coupling between lactate reabsorption and urate reabsorption in the kidney. Thus, the fortuitous
double knock-out of slc5a8 and slc5a12 in kidney in c/ebpδ–/– mice reveals the physiologic role of these transporters in the renal handling of lactate and urate.
BACKGROUND
Malignant cells exhibit increased glycolytic metabolism, and in many cases increased glucose transporter gene expression. The authors hypothesized that GLUT1 glucose transporter expression is increased in colorectal carcinoma, and that the degree of expression might have prognostic significance.METHODSGLUT1 glucose transporter immunostaining was studied in normal colon and benign colon adenomas and in 112 colorectal carcinomas from patients for whom long term clinical outcome was known.RESULTSGLUT1 immunostaining was absent in normal colorectal epithelium and tubular adenomas, and absent or only weakly apparent in tubulovillous adenomas. The majority of carcinomas (101 of 112; 90%) had GLUT1 immunostaining. Tumors from 92 patients had low GLUT1 expression (< 50% of cells were GLUT1 positive) and 19 of these patients (21%) died of disease during follow-up. In contrast, tumors from 20 patients had high GLUT1 expression (> 50% of cells were GLUT1 positive) and 9 of these patients (45%) died of disease during follow-up. Disease specific mortality was greater in patients with high GLUT1 tumors (relative risk of 2.4; P = 0.02). In a multivariate analysis to assess whether high GLUT1 staining correlated with increased mortality independently of Dukes stage, the risk of death from colon carcinoma in the group with high GLUT1 staining was 2.3 times that in the group with low GLUT1 staining, a difference that approached statistical significance (P = 0.07).CONCLUSIONSGLUT1 glucose transporter expression is associated strongly with neoplastic progression in the colon, and assessment of the extent of GLUT1 immunostaining in colorectal carcinoma identifies patients with a poorer prognosis. Cancer 1998;83:34-40. © 1998 American Cancer Society.
• Oligonucleotide primers based on the human heart monocarboxylate transporter (MCT1) cDNA sequence were used to isolate a 544 bp cDNA product from human colonic RNA by reverse transcription-polymerase chain reaction (RT-PCR). The sequence of the RT-PCR product was identical to that of human heart MCT1. Northern blot analysis using the RT-PCR product indicated the presence of a single transcript of 3.3 kb in mRNA isolated from both human and pig colonic tissues. Western blot analysis using an antibody to human MCT1 identified a specific protein with an apparent molecular mass of 40 kDa in purified and well-characterized human and pig colonic lumenal membrane vesicles (LMV). • Properties of the colonic lumenal membranel-lactate transporter were studied by the uptake of L-[U-14C]lactate into human and pig colonic LMV.l-lactate uptake was stimulated in the presence of an outward-directed anion gradient at an extravesicular pH of 5.5. Transport ofl-lactate into anion-loaded colonic LMV appeared to be via a proton-activated, anion exchange mechanism. • l-lactate uptake was inhibited by pyruvate, butyrate, propionate and acetate, but not by Cl− and SO42−. The uptake ofl-lactate was inhibited by phloretin, mercurials and α-cyano-4-hydroxycinnamic acid (4-CHC), but not by the stilbene anion exchange inhibitors, 4,4′-diisothiocyanostilbene-2,2′-disulphonic acid (DIDS) and 4-acetamido-4′-isothiocyanostilbene-2,2′-disulphonic acid (SITS). • The results indicate the presence of a MCT1 protein on the lumenal membrane of the colon that is involved in the transport ofl-lactate as well as butyrate across the colonic lumenal membrane. Western blot analysis showed that the abundance of this protein decreases in lumenal membrane fractions isolated from colonic carcinomas compared with that detected in the normal healthy colonic tissue.
Background and Objectives
It has been said that amino acid transporters play an important role in supplying nutrition to cells and for cell proliferation. In this study, we examined whether LAT1 and 4F2hc are closely related to tumor growth.Methods
Rat colon cancer cells (RCN-9) were injected into the spleen of 12 male rats (inbred F344/DuCrj). In each rat, liver samples including tumor lesions were immunostained with anti-LAT1 and anti-4F2hc antibodies. The staining area of LAT1 and 4F2hc tumor lesions was calculated by computer analysis.ResultsSixty-eight tumor nodules were observed in 12 livers. Out of the 68 tumor nodules, 36 nodules (52.9%) indicated a positive staining of LAT1 and 32 (47.1%) had a negative staining of LAT1. However, the LAT1 expression was scarcely detected in non-tumor areas. In terms of the 4F2hc expression, there were 56 nodules (82.4%) with 4F2hc positive and 12 (17.6) with 4F2hc negative. In addition, the expression of 4F2hc in non-tumor areas was almost the same as the expression of 4F2hc in tumor lesions. The average tumor size of the group with LAT1 positive and 4F2hc positive (n = 31) was 0.845 ± 0.232 mm2, which was significantly larger than that of the group with LAT1 negative and 4F2hc negative group (n = 7) (0.090 ± 0.028 mm2) or the group with LAT1 positive and 4F2hc negative (n = 5) (0.097 ± 0.025 mm2), respectively (P = 0.0017, P = 0.007).ConclusionLAT1 was related to tumor growth. We think that LAT1 can possibly enhance its ability to promote tumor growth in cooperation with 4F2hc. J. Surg. Oncol. 2001;78:265–272. © 2001 Wiley-Liss, Inc.
Background. The diagnosis of breast cancer based on nipple discharge, often the only clinical manifestation of early breast cancer, is currently unsatisfactory. Because M subunits of lactate dehydrogenase (LDH) have been noted to increase in cancer tissue, the author assessed the value of using LDH isozyme patterns in nipple discharge for the diagnosis of breast cancer.Methods. LDH isozyme levels in (1) nipple discharge of patients diagnosed as having breast cancer, intraductal papilloma, mastopathy, drug-induced nipple discharge, mastitis, or benign nipple discharge; (2) control samples of normal nipple discharge (milk) 6 days, 1–5 months, and 6 months to 2 years postpartum; (3) the serum of patients presenting with nipple discharge; and (4) normal and cancerous breast tissue extracts were measured using a Ciba-Corning LDH isozyme system (Ciba Corning Diagnostic Corp., Tokyo, Japan).Results. LDH isozyme levels in the nipple discharge of patients with benign breast diseases displayed various patterns. Levels in the nipple discharge of patients with breast cancer, including noninvasive carcinoma, tended to increase in ascending order from LDH1 to LDH5. This pattern was similar to that in breast cancer tissue and was unrelated to the pattern in serum.Conclusion. LDH isozyme assay of nipple discharge may be a useful technique for providing a supporting diagnosis of breast cancer.
SIRT1 impacts upon diverse cellular processes via its roles in the determination of chromatin structure, chromatin remodelling and gene expression. This review covers the recent discoveries linking SIRT1 with the regulation of mammalian metabolism and considers ways in which abnormal metabolism in disease may, in turn, impact upon SIRT1 because of SIRT1's functional dependency upon NAD.
Diverse signalling pathways are integrated to regulate energy metabolism and homeostasis. Such pathways involve intracellular networks and mitochondria, and also intercellular signalling within and between tissues to co-ordinate adaptive metabolic responses within the organism as a whole. Here, we outline the recent studies exploring the regulatory links between SIRT1 and mitochondrial biogenesis, cellular redox and associated metabolic pathways, and angiogenesis/Notch signalling. These links are effected by the SIRT1-mediated deacetylation of transcriptional regulators and enzymes with key roles in metabolism.
SIRT1 activity is directly coupled with homeostasis and metabolism. SIRT1 is also a metabolic sensor. It follows that disease-related metabolic abnormalities are likely to impinge upon SIRT1 functioning. Disease-related functions of SIRT1, in their turn, offer potential targets for the development of novel SIRT1-based therapies. In cancer, for example, the survival function of SIRT1 may reflect abnormal cancer metabolism and identifies SIRT1 as a target for anticancer therapy.
SLC5A8, expressed predominantly in the colon, is a Na(+)-coupled transporter for short-chain fatty acids. In this paper, we report on the characterization of butyrate transport by SLC5A8 and the relevance of SLC5A8-mediated butyrate transport to colon cancer.
SLC5A8 transports butyrate via a Na(+)-dependent electrogenic process. Na(+) activation of the transport process exhibits sigmoidal kinetics, indicating involvement of more than one Na(+) in the activation process. SLC5A8 is silenced in colon cancer in humans, in a mouse model of intestinal/colon cancer, and in colon cancer cell lines. The tumor-associated silencing of SLC5A8 involves DNA methylation by DNA methyltransferase 1. Reexpression of SLC5A8 in colon cancer cells leads to apoptosis but only in the presence of butyrate. SLC5A8-mediated entry of butyrate into cancer cells is associated with inhibition of histone deacetylation. The changes in gene expression in SLC5A8/butyrate-induced apoptosis include upregulation of pro-apoptotic genes and downregulation of anti-apoptotic genes. In addition, the expression of phosphatidylinositol-3-kinase subunits is affected differentially, with downregulation of p85alpha and upregulation of p55alpha and p50alpha.
These studies show that SLC5A8 mediates the tumor-suppressive effects of the bacterial fermentation product butyrate in the colon.
L-Type amino acid transporter 1 (LAT1) is highly expressed in cancer cells to support their continuous growth and proliferation. We have examined the effect of 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH), an inhibitor of system L amino acid transporters, and the mechanism by which BCH suppresses cell growth in cancer cells.
The effect of BCH and the mechanism of BCH on cell growth suppression in cancer cells were examined using amino acid transport measurement, MTT assay, DNA fragmentation analysis, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay and immunoblotting.
BCH inhibited L-leucine transport in a concentration-dependent manner, and it inhibited cell growth in a time-dependent manner in KB human oral epidermoid carcinoma cells, Saos2 human osteogenic sarcoma cells and C6 rat glioma cells. The formation of a DNA ladder was observed, and the number of TUNEL-positive cells was increased with BCH treatment. Furthermore, the proteolytic processing of caspase-3 in KB and C6 cells and of caspase-7 in KB, Saos2 and C6 cells was increased by BCH treatment.
These results suggest that the inhibition of LAT1 activity by BCH leads to apoptotic cancer cell death by inducing intracellular depletion of neutral amino acids necessary for cancer cell growth.
This introductory article to the review series entitled "The Cancer Cell's Power Plants as Promising Therapeutic Targets" is written while more than 20 million people suffer from cancer. It summarizes strategies to destroy or prevent cancers by targeting their energy production factories, i.e., "power plants." All nucleated animal/human cells have two types of power plants, i.e., systems that make the "high energy" compound ATP from ADP and P( i ). One type is "glycolysis," the other the "mitochondria." In contrast to most normal cells where the mitochondria are the major ATP producers (>90%) in fueling growth, human cancers detected via Positron Emission Tomography (PET) rely on both types of power plants. In such cancers, glycolysis may contribute nearly half the ATP even in the presence of oxygen ("Warburg effect"). Based solely on cell energetics, this presents a challenge to identify curative agents that destroy only cancer cells as they must destroy both of their power plants causing "necrotic cell death" and leave normal cells alone. One such agent, 3-bromopyruvate (3-BrPA), a lactic acid analog, has been shown to inhibit both glycolytic and mitochondrial ATP production in rapidly growing cancers (Ko et al., Cancer Letts., 173, 83-91, 2001), leave normal cells alone, and eradicate advanced cancers (19 of 19) in a rodent model (Ko et al., Biochem. Biophys. Res. Commun., 324, 269-275, 2004). A second approach is to induce only cancer cells to undergo "apoptotic cell death." Here, mitochondria release cell death inducing factors (e.g., cytochrome c). In a third approach, cancer cells are induced to die by both apoptotic and necrotic events. In summary, much effort is being focused on identifying agents that induce "necrotic," "apoptotic" or apoptotic plus necrotic cell death only in cancer cells. Regardless how death is inflicted, every cancer cell must die, be it fast or slow.
Estimation of lactate dehydrogenase (LDH) isoenzymes in the serum and aqueous humor was carried out in 15 cases of benign ocular tumour, 15 cases of malignant tumor and 15 normal cases. Cases of both sexes aged between 1 year and 75 years were included. LDH, isoenzymes specially LDH4 and LDH5 are higher and LDH1 and LDH2 lower in sera of patients with malignant tumor specially retinoblastoma as compared to benign tumor cases and control cases. LDH isoenzymes in aqueous humor are significantly higher and show a characteristic pattern in retinoblastoma cases, the concentration was presumably too low in the control, malignant tumor other than retinoblastoma and benign tumor cases as its fractionation was not possible.
Elevation of glucose transport is an alteration common to most virally induced tumors. Rat fibroblasts transformed with wild-type
or a temperature-sensitive Fujinami sarcoma virus (FSV) were studied in order to determine the mechanisms underlying the increased
transport. Five- to tenfold increases in total cellular glucose transporter protein in response to transformation were accompanied
by similar increases in transporter messenger RNA levels. This, in turn, was preceded by an absolute increase in the rate
of glucose transporter gene transcription within 30 minutes after shift of the temperature-sensitive FSV-transformed cells
to the permissive temperature. The transporter messenger RNA levels in transformed fibroblasts were higher than those found
in proliferating cells maintained at the nonpermissive temperature. The activation of transporter gene transcription by transformation
represents one of the earliest known effects of oncogenesis on the expression of a gene encoding a protein of well-defined
function.
An accelerated rate of glucose transport is among the most characteristic biochemical markers of cellular transformation.
To study the molecular mechanism by which transporter activity is altered, cultured rodent fibroblasts transfected with activated
myc, ras, or src oncogenes were used. In myc-transfected cells, the rate of 2-deoxy-D-glucose uptake was unchanged. However,
in cells transfected with activated ras and src oncogenes, the rate of glucose uptake was markedly increased. The increased
transport rate in ras- and src-transfected cells was paralleled by a marked increase in the amount of glucose transporter
protein, as assessed by immunoblots, as well as by a markedly increased abundance of glucose transporter messenger RNA. Exposure
of control cells to the tumor-promoting phorbol ester 12-O-tetradecanoyl phorbol-13-acetate (TPA) for 18 hours had a similar
effect of increasing the rate of glucose transport and the abundance of transporter messenger RNA. For ras, src, and TPA,
the predominant mechanism responsible for activation of the transport system is increased expression of the structural gene
encoding the glucose transport protein.
The activities of six different enzymes were compared in 29 normal, 34 dysplastic, and 80 cancerous (both primary and metastatic) human breast tissues; in MCF-7 cells; and in primary rat mammary tumors. Benign lesions generally showed enzyme activities similar to those of normal breast tissues. Malignant tumors had significantly increased activities of lactate dehydrogenase (LDH), malate dehydrogenase (MDH), fructose-bisphosphate aldolase, hexokinase (HK), pyruvate kinase (PK), and creatine kinase. Enzyme activity in the malignant tumor was always higher than that in apparently normal or fibrocystic tissue from the same patient. Enzyme activities did not correlate with the levels of estrogen and progesterone receptors. LDH, MDH, and HK were elevated to a similar extent in all the tissues examined. Conversely, PK was elevated to a much greater extent in cancerous tissues, particularly in MCF-7 cells. The elevated activities of these enzymes may have diagnostic potential, especially when tumor tissue and apparently normal tissue from the same patient are compared.
Isozyme patterns of 23 different enzymes were compared in normal, benign, and malignant breast tissues; in MCF-7 cells; and in organoids of normal human breast tissue. Benign lesions generally showed isozyme patterns similar to those of normal tissues. Lactate dehydrogenase isozyme 5 was significantly increased in malignant tumors; MCF-7 cells had only lactate dehydrogenase (L-lactate:NAD oxidoreductase; EC 1.1.1.27). The mitochondrial form of malate dehydrogenase was also significantly increased in human malignant tumors; this was especially evident when comparing tumor and apparently uninvolved breast tissue from the same patient. The K4 isozyme of pyruvate kinase was the major form in most malignant breast tumors, but in only 41% of normal tissues, 30% of fibrocystic disease specimens, and 46% of fibroadenomas. A more anodal band of pyruvate kinase, probably a K3M or K3Kpm hybrid, predominated in most normal and benign tissues, but in only 63% of primary and 56% of secondary tumors. All specimens had predominantly creatine kinase BB, aldolase A4, and hexokinase I. Traces of aldolase A3C and of hexokinase II were observed in some tumors. None of the tumors had the Regan variant of alkaline phosphatase. The isozymes of lactate and malate dehydrogenases and of pyruvate kinase appear to be the most promising as putative tumor markers.
The diagnosis of breast cancer based on nipple discharge, often the only clinical manifestation of early breast cancer, is currently unsatisfactory. Because M subunits of lactate dehydrogenase (LDH) have been noted to increase in cancer tissue, the author assessed the value of using LDH isozyme patterns in nipple discharge for the diagnosis of breast cancer.
LDH isozyme levels in (1) nipple discharge of patients diagnosed as having breast cancer, intraductal papilloma, mastopathy, drug-induced nipple discharge, mastitis, or benign nipple discharge; (2) control samples of normal nipple discharge (milk) 6 days, 1-5 months, and 6 months to 2 years postpartum; (3) the serum of patients presenting with nipple discharge; and (4) normal and cancerous breast tissue extracts were measured using a Ciba-Corning LDH isozyme system (Ciba Corning Diagnostic Corp., Tokyo, Japan).
LDH isozyme levels in the nipple discharge of patients with benign breast diseases displayed various patterns. Levels in the nipple discharge of patients with breast cancer, including noninvasive carcinoma, tended to increase in ascending order from LDH1 to LDH5. This pattern was similar to that in breast cancer tissue and was unrelated to the pattern in serum.
LDH isozyme assay of nipple discharge may be a useful technique for providing a supporting diagnosis of breast cancer.
Breast cancers have been shown to have increased glucose uptake and utilization, and to express the facilitative glucose transporter Glut1. The aim of this study was to determine the biological significance of Glut1 expression in breast cancer.
Paraffin sections of 118 breast cancers were immunostained with antibody to Glut1. The percent of Glut1-positive cancer cells in each tumor was correlated with known prognostic markers, and with patient outcome.
Glut1 was expressed in 42% of the tumors. Glut1 immunoreactivity correlated positively with the proliferative activity as determined by Ki-67 immunostaining, and with the total histologic score, and showed negative correlation with bcl-2 immunostaining. There was no correlation between the percent of Glut1-immunoreactive cancer cells and estrogen receptor status, tumor size, or lymph node status.
1) Glut1 expression is increased in breast cancers with higher grade and proliferative activity, and 2) glucose transport in the majority of breast cancers may be mediated by a glucose transporters other than Glut1.
Fermentation, the process whereby anaerobic bacteria break down carbohydrates to short-chain (C2-C6) fatty acids (SCFAs), is an important function of the large bowel. SCFAs constitute approximately two-thirds of the colonic anion concentration (70-130 mmol/l), mainly as acetate, propionate, and butyrate. Gastroenterologists have, in spite of these facts, addressed this scientific field surprisingly late, in contrast to veterinarians, for whom the fermentative production of SCFAs has been acknowledged as a principal mechanism of intestinal digestion in plant-eating animals for decades. Interest in the effects of SCFA production on the human organism has been growing rapidly in the last 10 years, because gastrointestinal functions and beneficial effects are associated with these acids. SCFAs are of major importance in the understanding of the physiological function of dietary fibre and their possible role for colonic neoplasia. SCFA production and absorption are closely related to the nourishment of the colonic mucosa and sodium and water absorption, and mechanisms of diarrhoea. Patients with severe malabsorption compensate by the fermentation of otherwise osmotic active saccharides to SCFAs, which are readily absorbed and used as energy fuels in the organism. SCFA production from dietary carbohydrates is a mechanism whereby considerable amounts of calories can be salvaged in short-bowel patients with remaining colonic function if dietary treatment is adjusted. SCFA enemas are a new and promising treatment modality for patients with ulcerative colitis. The effect has been attributed to the oxidation of SCFAs in the colonocytes. An impressive number of papers have described the effects of butyrate on various cell functions, the significance of which is still unknown. Up until now, attention has been related especially to cancer prophylaxis and treatment. Diminished production of SCFAs appears to be involved in antibiotic-associated diarrhoea, diversion colitis, and possibly in pouchitis. The interaction between bacterial fermentation, ammonia metabolism, and bacterial growth and protein synthesis appears to be the main mechanism of action of lactulose treatment in hepatic coma. Pathological and extremely high rates of saccharide fermentation explain the severe deterioration in patients with D-lactate acidosis. Hence, this scientific field has come late to clinical working gastroenterologists, but as work is progressing the production of SCFAs in the large bowel becomes involved in several well-known intestinal disorders.
Increased glucose transport is a common characteristic of most tumors. To examine the role of elevated glucose uptake in lung cancer, we performed PCR amplification of 2 facilitative glucose transporter genes (GLUT1 and GLUT3) and immunohistochemical staining for GLUT1, proliferating cell nuclear antigen (PCNA), and sialyl Lewis x (sLe(x)) on tumor specimens from 327 patients with lung cancer who underwent surgical resection from 1980 to 1993. To evaluate the relationship between GLUT, alpha-2,3-sialyltransferase (ST), and alpha-1,3-fucosyltransferase (Fuc-T) genes, PCR amplification of the ST3N and Fuc-TVII also was performed. Amplification of GLUT1 was significantly greater than that of GLUT3. GLUT1 and GLUT3 amplification correlated with PCNA staining (p < 0.01). In addition, GLUT1 amplification correlated with the grading of sLe(x) staining as well as with the grading of GLUT1 staining (p < .03, p < 0.01). GLUT1 was co-amplified with ST3N and Fuc-TVII genes, which are involved in the synthesis of sLe(x) (p < 0.01). The survival of patients whose tumors showed GLUT1 amplification was significantly shorter than that of patients whose tumors did not (p < 0.01). In a multivariate analysis of survival, GLUT1 remained a statistically significant prognostic factor. Our results suggest that GLUT1 amplification may participate in sLe(x) synthesis as well as in proliferation, and may be of prognostic value in lung cancer.
The increased glucose uptake seen in cancer cells correlates with the expression of human erythrocyte glucose transporter (Glut1) protein in certain human malignancies.
Our purpose was to determine Glut1 expression in cutaneous neoplasms.
A polyclonal anti-Glut1 antibody (MYM) and a standard ABC immunoperoxidase technique were used to determine Glut1 expression in invasive squamous cell carcinomas (SCCs), SCC in situ, basal cell carcinomas (BCCs), melanomas, actinic keratoses (AKs), seborrheic keratoses, common acquired nevi, and scars with regenerative epidermal hyperplasia.
All of the cases of SCC in situ, 14 of 15 (93%) of the SCC, and 13 of 15 AKs (87%) showed intense membranous staining for Glut1. Glut1 staining was present in the epidermis of 8 of 15 scars (53%) but was not detected in any BCC, even in areas of focal keratinization and squamous metaplasia. Glut1 reactivity was absent in the melanomas and seborrheic keratoses.
Glut1 expression in a cutaneous lesion strongly suggests a proliferative lesion of the squamous cell type.
We have investigated the functional characteristics of the human amino acid transporter ATB degree using the Xenopus laevis oocyte expression system. When expressed in oocytes, ATB degree mediates the uptake of neutral amino acids in an Na(+)-dependent manner. In addition, this transporter is able to mediate the efflux of intracellular neutral amino acids in exchange with extracellular neutral amino acids. This homo- and hetero-exchange of amino acids is absolutely Na(+)-dependent and conforms strictly to the substrate specificity of ATB degree. Kinetic analysis indicates that the affinity of ATB degree for a given amino acid substrate is similar whether ATB degree catalyzes the influx of the amino acid or the amino acid-induced efflux of intracellular amino acids. These results demonstrate for the first time the ability of ATB degree to function as a homo- and hetero-exchanger for its substrates.
Malignant cells exhibit increased glycolytic metabolism, and in many cases increased glucose transporter gene expression. The authors hypothesized that GLUT1 glucose transporter expression is increased in colorectal carcinoma, and that the degree of expression might have prognostic significance.
GLUT1 glucose transporter immunostaining was studied in normal colon and benign colon adenomas and in 112 colorectal carcinomas from patients for whom long term clinical outcome was known.
GLUT1 immunostaining was absent in normal colorectal epithelium and tubular adenomas, and absent or only weakly apparent in tubulovillous adenomas. The majority of carcinomas (101 of 112; 90%) had GLUT1 immunostaining. Tumors from 92 patients had low GLUT1 expression (< 50% of cells were GLUT1 positive) and 19 of these patients (21%) died of disease during follow-up. In contrast, tumors from 20 patients had high GLUT1 expression (> 50% of cells were GLUT1 positive) and 9 of these patients (45%) died of disease during follow-up. Disease specific mortality was greater in patients with high GLUT1 tumors (relative risk of 2.4; P=0.02). In a multivariate analysis to assess whether high GLUT1 staining correlated with increased mortality independently of Dukes stage, the risk of death from colon carcinoma in the group with high GLUT1 staining was 2.3 times that in the group with low GLUT1 staining, a difference that approached statistical significance (P=0.07).
GLUT1 glucose transporter expression is associated strongly with neoplastic progression in the colon, and assessment of the extent of GLUT1 immunostaining in colorectal carcinoma identifies patients with a poorer prognosis.
Owing to the high efficacy of L-asparaginase in the treatment of acute lymphatic leukaemia the enzyme was introduced into the chemotherapy schedules for remission induction of this disease shortly after results of large-scale clinical trials had become available. Since asparaginase monotherapy was associated with a high response rate but short remission duration, the enzyme is currently employed within the framework of combination chemotherapy schedules which achieve treatment response in about 90% and long-term remissions in the majority of patients. Recently initiated clinical trials have still confirmed the eminent value of asparaginase in the combination chemotherapy of acute lymphatic leukaemia and of some subtypes of non-Hodgkin lymphoma, and its important role as an essential component of multimodal treatment protocols. Despite the unique mechanism of action of this cytotoxic substance which shows relative selectivity with regard to the metabolism of malignant cells, some patients experience toxic effects during asparaginase therapy. Immunological reactions toward the foreign protein include enzyme inactivation without any clinical manifestations as well as anaphylactic shock. Severe functional disorders of organ systems result from the impaired homeostasis of the amino acids asparagine and glutamine. The changes affecting the proteins of the coagulation system have considerable clinical impact as they may induce bleeding as well as thromboembolic events and may be associated with life-threatening complications when the central nervous system is involved. Risk factors predisposing to thromboembolic complications are hereditary resistance against activated protein C and any other hereditary thrombophilia. Other organ systems potentially affected by relevant functional disorders are the central nervous system, the liver, and the pancreas, with patients who have a history of pancreatic disorders carrying an especially high risk of developing pancreatitis. Studies on the mechanisms of action and the occurrence of resistance phenomena have shown that a treatment response may only be expected if the malignant cells are unable to increase their asparagine synthetase activity to an extent providing enough asparagine to the cell; one may thus conclude that the enzyme-induced asparagine depletion of the serum constitutes the decisive cytotoxic mechanism. Independent of the asparagine depletion related cytotoxicity however, there are other mechanisms of clinical relevance like induction of apoptosis. Besides this, further influences on signal transduction cannot be excluded. Only few publications have dealt with the question of minimum trough activities to be ensured before each subsequent asparaginase dose in order to maintain uninterrupted asparagine depletion under treatment, and answers to this problem are not definitive. Clinical studies using enzymes from E. coli strains indicate that a trough activity of 100 U/l will suffice for complete asparagine depletion of the fluid body compartments with the preparations studied. These findings have been transferred to enzymes from other E. coli strains as well as those isolated from Erwinia chrysanthemi and to the PEG-conjugated E. coli asparaginases. It might be desirable to countercheck the results for confirmation or correction. The dosage and administration schedule of the various enzyme preparations required for complete asparagine depletion over a period of time have been insufficiently defined. While pharmacokinetic studies showed clinically relevant differences in biological activity and activity half-lives for enzymes from different biological sources, the findings of recently published clinical trials indicate that the therapeutic efficacy is affected when different asparaginase preparations are given by identical therapy schedules. (ABSTRACT TRUNCATED)