Regulation Of Polyamine Metabolism Research Articles

Spermine deficiency shifts the balance between jasmonic acid and salicylic acid-mediated defence responses in Arabidopsis.

Polyamines are small aliphatic polycations present in all living organisms. In plants, the most abundant polyamines are putrescine (Put), spermidine (Spd) and spermine (Spm). Polyamine levels change in response to different pathogens, including Pseudomonas syringae pv. tomato DC3000 (Pst DC3000). However, the regulation of polyamine metabolism and their specific contributions to defence are not fully understood. Here we report that stimulation of Put biosynthesis by Pst DC3000 is dependent on coronatine (COR) perception and jasmonic acid (JA) signalling, independently of salicylic acid (SA). Conversely, lack of Spm in spermine synthase (spms) mutant stimulated galactolipids and JA biosynthesis, and JA signalling under basal conditions and during Pst DC3000 infection, whereas compromised SA-pathway activation and defence outputs through SA-JA antagonism. The dampening of SA responses correlated with COR and Pst DC3000-inducible deregulation of ANAC019 expression and its key SA-metabolism gene targets. Spm deficiency also led to enhanced disease resistance to the necrotrophic fungal pathogen Botrytis cinerea and stimulated endoplasmic reticulum (ER) stress signalling in response to Pst DC3000. Overall, our findings provide evidence for the integration of polyamine metabolism in JA- and SA-mediated defence responses, as well as the participation of Spm in buffering ER stress during defence.

Spermidine as a promising anticancer agent: Recent advances and newer insights on its molecular mechanisms.

Spermidine is a naturally occurring polyamine compound found in semen. It is also found in several plant sources and boasts a remarkable biological profile, particularly with regards to its anticancer properties. Spermidine specifically interferes with the tumour cell cycle, resulting in the inhibition of tumor cell proliferation and suppression of tumor growth. Moreover, it also triggers autophagy by regulating key oncologic pathways. The increased intake of polyamines, such as spermidine, can suppress oncogenesis and slow the growth of tumors due to its role in anticancer immunosurveillance and regulation of polyamine metabolism. Spermidine/spermine N-1-acetyltransferase (SSAT) plays a critical role in polyamine homeostasis and serves as a diagnostic marker in human cancers. Chemically modified derivatives of spermidine hold great potential for prognostic, diagnostic, and therapeutic applications against various malignancies. This review discusses in detail the recent findings that support the anticancer mechanisms of spermidine and its molecular physiology.

Investigation of The Roles of Hydrogen Peroxide and NADPH Oxidase in The Regulation of Polyamine Metabolism in Maize Plants under Drought Stress Conditions

The relationship between hydrogen peroxide and polyamine metabolism and the role of NADPH oxidase (NOX) in this relationship under drought conditions has not yet been clarified. To reveal the relationship, expression levels of the genes in polyamine metabolism, such as arginine decarboxylase (ADC), agmatine aminohydrolase (AIH), spermidine synthase (SPDS), S-adenosyl methionine decarboxylase (SAMDC), diamine oxidase (DAO) and polyamine oxidase (PAO) were determined by RT PCR under drought stress combined with exogenous hydrogen peroxide (H2O2) and diphenyleneiodonium chloride (DPI) treatments in maize seedlings. Also, some basic stress parameters (leaf water potential, lipid peroxidation), polyamine levels (putrescine, spermidine, and spermine), and gene expression of NOX were measured under drought stress. Exogenous H2O2 induced polyamine contents by up-regulating polyamine-synthesizing genes and downregulating polyamine oxidizing genes. Moreover, when the NOX enzyme was inhibited by DPI, the polyamine metabolic pathway tended toward degradation rather than production. Exogenous H2O2 regulated polyamine metabolism to promote their synthesis, and NOX had a key role in this regulation.

Application of γ-aminobutyric acid induces disease resistance in apples through regulation of polyamine metabolism, GABA shunt and reactive oxygen species metabolism

Apples were treated with γ-aminobutyric acid (GABA) to investigate its effect on blue mold, reactive oxygen species (ROS) metabolism and GABA shunt. The results showed that GABA significantly inhibit blue mold of apples. Further studies demonstrated that GABA remarkably increased H2O2, ascorbic acid, and glutathione contents; superoxide dismutase, NADPH oxidase, catalase, glutathione reductase, ascorbate peroxidase, dehydroascorbate reductase, and monodehydroascorbate reductase activities in apple fruit. GABA also activated glutamate decarboxylase, glutamate dehydrogenase and glutamate synthase, while inhibited GABA transaminase and succinate hemialdehyde dehydrogenase in apples. GABA treatment increased endogenous GABA, pyruvic acid and glutamate contents in the fruit. In addition, exogenous GABA dipping up-regulated MdMT, MdMS, MdSAMS, MdSAMDC, MdODC, MdADC and MdSPDS, and down-regulated MdPAO and MdDAO in apples. These findings indicate that GABA enhances disease resistance in apples through regulation of GABA shunt, ROS and polyamine metabolism.

4-Hydroxymelatonin alleviates nickel stress, improves physiochemical traits of Solanum melongena: Regulation of polyamine metabolism and antioxidative enzyme

The core objective of current research was to reveal the potential of 4-hydroxymelatonin (4-OHMT) in alleviation of nickel (Ni) toxicity in Solanum melongena L. Nickel toxicity reduced growth, leaf relative water content (LRWC), chlorophyll (Chl) content, intercellular carbon dioxide concentration, stomatal conductivity and transpiration rate in S. melongena seedlings. However, exogenous application of 4-OHMT enhanced growth, net photosynthesis rate and gas exchange characteristics in treated seedlings. Moreover, 4-OHMT amplified the activity of superoxide dismutase, catalase and ascorbate peroxidase. Additionally, 4-OHMT applied plants exhibited higher level of glutathione (GSH). The stress alleviation capability of 4-OHMT in S. melongena seedlings is attributed to the reduced level of hydrogen peroxide, malondialdehyde and electrolyte leakage. 4-Hydroxymelatonin reduced Ni content in root and shoot of S. melongena seedlings. Further research with reference to potential of 4-OHMT for stress mitigation in other crops will assist in increasing the crop productivity.

Antizyme Inhibitors in Polyamine Metabolism and Beyond: Physiopathological Implications.

The intracellular levels of polyamines, cationic molecules involved in a myriad of cellular functions ranging from cellular growth, differentiation and apoptosis, is precisely regulated by antizymes and antizyme inhibitors via the modulation of the polyamine biosynthetic and transport systems. Antizymes, which are mainly activated upon high polyamine levels, inhibit ornithine decarboxylase (ODC), the key enzyme of the polyamine biosynthetic route, and exert a negative control of polyamine intake. Antizyme inhibitors (AZINs), which are proteins highly homologous to ODC, selectively interact with antizymes, preventing their action on ODC and the polyamine transport system. In this review, we will update the recent advances on the structural, cellular and physiological functions of AZINs, with particular emphasis on the action of these proteins in the regulation of polyamine metabolism. In addition, we will describe emerging evidence that suggests that AZINs may also have polyamine-independent effects on cells. Finally, we will discuss how the dysregulation of AZIN activity has been implicated in certain human pathologies such as cancer, fibrosis or neurodegenerative diseases.

Regulation of Polyamine Metabolism by Curcumin for Cancer Prevention and Therapy.

Curcumin (diferuloylmethane), the natural polyphenol responsible for the characteristic yellow pigment of the spice turmeric (Curcuma longa), is traditionally known for its antioxidant, anti-inflammatory, and anticarcinogenic properties. Capable of affecting the initiation, promotion, and progression of carcinogenesis through multiple mechanisms, curcumin has potential utility for both chemoprevention and chemotherapy. In human cancer cell lines, curcumin has been shown to decrease ornithine decarboxylase (ODC) activity, a rate-limiting enzyme in polyamine biosynthesis that is frequently upregulated in cancer and other rapidly proliferating tissues. Numerous studies have demonstrated that pretreatment with curcumin can abrogate carcinogen-induced ODC activity and tumor development in rodent tumorigenesis models targeting various organs. This review summarizes the results of curcumin exposure with regard to the modulation of polyamine metabolism and discusses the potential utility of this natural compound in conjunction with the exploitation of dysregulated polyamine metabolism in chemopreventive and chemotherapeutic settings.

Effect of Oaz1 overexpression on goose ovarian granulosa cells.

Ornithine decarboxylase antizyme 1 (OAZ1) regulates both polyamine biosynthesis and transport and acts as a tumor suppressor because of its functions in the regulation of cell proliferation, apoptosis and growth. However, the roles of OAZ1 in the regulation of polyamine metabolism, cell proliferation, apoptosis and hormone receptor expression in granulosa cells (GCs) from geese have not been determined. To define the roles of OAZ1 in primary GCs, we constructed and transfected an Oaz1 overexpression vector with a thymidine deletion into primary GCs. Oaz1 overexpression induced 65 and 30% decreases in the putrescine and spermidine contents, respectively, 225% increase in the spermine content and up-regulated the expression levels of genes associated with polyamine metabolism (but Oaz2). The Ccnd1, Pcna, Bax and Caspase 3 mRNA expression levels in GCs overexpressing Oaz1 were significantly higher than the expression levels in the vehicle group (p<0.05). Additionally, a slight increase in the E2 concentration in the culture medium and enhanced Er, Fshr and Lhr expression were observed in the GCs 24h after Oaz1 overexpression (p<0.05). These data suggested that Oaz1 up-regulated the expression of genes related to polyamine metabolic enzymes and reproductive hormone receptors, disturbed intracellular polyamine homeostasis, and affected the transcription of genes associated to cell proliferation and apoptosis in goose primary GCs. Taken together, our results indicate that modifying polyamine metabolism by Oaz1 may interfere with the expression of genes involved in cell proliferation, apoptosis and responsiveness of ovary toward hormones in goose GCs.

Activation of SAT1 engages polyamine metabolism with p53-mediated ferroptotic responses

Although p53-mediated cell-cycle arrest, senescence, and apoptosis remain critical barriers to cancer development, the emerging role of p53 in cell metabolism, oxidative responses, and ferroptotic cell death has been a topic of great interest. Nevertheless, it is unclear how p53 orchestrates its activities in multiple metabolic pathways into tumor suppressive effects. Here, we identified the SAT1 (spermidine/spermine N1-acetyltransferase 1) gene as a transcription target of p53. SAT1 is a rate-limiting enzyme in polyamine catabolism critically involved in the conversion of spermidine and spermine back to putrescine. Surprisingly, we found that activation of SAT1 expression induces lipid peroxidation and sensitizes cells to undergo ferroptosis upon reactive oxygen species (ROS)-induced stress, which also leads to suppression of tumor growth in xenograft tumor models. Notably, SAT1 expression is down-regulated in human tumors, and CRISPR-cas9-mediated knockout of SAT1 expression partially abrogates p53-mediated ferroptosis. Moreover, SAT1 induction is correlated with the expression levels of arachidonate 15-lipoxygenase (ALOX15), and SAT1-induced ferroptosis is significantly abrogated in the presence of PD146176, a specific inhibitor of ALOX15. Thus, our findings uncover a metabolic target of p53 involved in ferroptotic cell death and provide insight into the regulation of polyamine metabolism and ferroptosis-mediated tumor suppression.

Inhibition of polyamine oxidase reduces hyperoxia‐mediated neurovascular injury in retina

Retinopathy of prematurity (ROP) is a potentially blinding eye disease that affects premature infants and often leads to lifelong vision impairment. Hyperoxia treatment is shown to induce neurovascular damage in the developing retina. Using an oxygen‐induced retinopathy (OIR) mouse model for ROP, we have demonstrated that deletion of the arginase 2 reduced hyperoxia–induced retinal neurodegeneration through the regulation of polyamine metabolism. The present study was undertaken to study the impact of polyamine oxidase (PAO) inhibitor, MDL 72527, on hyperoxia‐induced retinal neurovascular injury in the OIR retina. Newborn wild‐type (WT) mice pups were exposed to 70% oxygen from postnatal day (P) 7 to P12. Pups were treated daily starting at P6 with either vehicle or MDL 72527 (39 mg/kg of body weight, in saline, i.p.) and sacrificed at different stages of hyperoxia treatment. Retinal cryostat sections were used for neurodegeneration studies using immunolabeling of retinal neurons. Analysis of vaso‐obliteration (N=10–12), vessel sprouting (N=7–9) and microglial activation (N=6) were performed on retinal flat mounts. Mixed rat retinal neuronal cultures (7 DIV) were subjected to normoxia (21%) or hyperoxia (50%) with or without MDL 72527 at different time points (N=6–8). These were processed for cellular signaling analysis using Western blotting. Immunolabeling studies using synaptophysin antibody showed the presence of improved synaptic contacts in MDL 72527‐treated OIR retina (P12) compared to vehicle treated groups. Analysis of retinal flat mounts showed significantly reduced a vascular area at P9 and P12( p&lt;0.05) and increased vascular sprouting at P9 (p&lt;0.01) in MDL 72527‐treated compared to vehicle‐treated OIR retinas after 2 or 5 days of hyperoxia. These results demonstrate reduced vascular injury in OIR retina in response to PAO inhibition. Immunolabeling studies using Iba‐1 antibody showed that activation of microglia was significantly reduced and number of resting microglia were increased in the OIR retina with MDL treatment (p&lt;0.05). Analysis of signaling pathways in retinal neurons in response to hyperoxia showed activation of neuro‐inflammatory signaling studied by increased p‐NFkB, p‐ERK1/2 and cytochrome‐C levels, which were significantly reduced by MDL treatment. Our data suggest that PAO inhibition offers a new therapeutic target towards hyperoxia‐induced retinal neurovascular injury.Support or Funding InformationThese studies are supported by AHA 11SDG7440088, Culver VDI Pilot Grant Award and NEI‐R01‐EY011766.

Chemical hypoxia-induced stimulation of polyamine biosynthesis and ornithine decarboxylase gene transcription in C6 glioma cells

Polyamines, aliphatic polycationic compounds distributed ubiquitously in the living organisms, are considered to be implicated in various cellular processes, and furthermore speculated to play a possible pathophysiological role in physical and mental disorders. In particular, the regulation of polyamine metabolism in damaged glial cells seems interesting and important to understand a possible connection of polyamines with the brain functions. On the other hand, previous studies have shown that ornithine decarboxylase (ODC), the enzyme catalyzing the rate-limiting step of polyamine biosynthesis, can be activated during the period of recovery from various damage in the brain, but little is known about the effects of toxic insults on the expression of ODC gene in neuronal and glial cells. Based on the hypothesis that noxious impacts on the brain might be able to increase polyamine production in glial cells to protect the neuronal cell functions against toxic injuries, the effects of chemical hypoxia on polyamine production and the transcription of ODC gene in C6 glioma cells were investigated using a HPLC assay method and a one-step RT-PCR analysis. In consequence, both cobalt and azide were shown to increase the cellular polyamine contents through elevating ODC mRNA levels under the conditions in which hypoxia-inducible factor-1 (Hif-1) gene was expressed. Thus, chemical hypoxia is speculated to induce ODC gene transcription, thus resulting in the increment of polyamine contents in glial cells, thereby contributing to the recovery of brain function from hypoxic injury.

Changes in free polyamine levels, expression of polyamine biosynthesis genes, and performance of rice cultivars under salt stress: a comparison with responses to drought.

Soil salinity affects a large proportion of rural area and limits agricultural productivity. To investigate differential adaptation to soil salinity, we studied salt tolerance of 18 varieties of Oryza sativa using a hydroponic culture system. Based on visual inspection and photosynthetic parameters, cultivars were classified according to their tolerance level. Additionally, biomass parameters were correlated with salt tolerance. Polyamines have frequently been demonstrated to be involved in plant stress responses and therefore soluble leaf polyamines were measured. Under salinity, putrescine (Put) content was unchanged or increased in tolerant, while dropped in sensitive cultivars. Spermidine (Spd) content was unchanged at lower NaCl concentrations in all, while reduced at 100 mM NaCl in sensitive cultivars. Spermine (Spm) content was increased in all cultivars. A comparison with data from 21 cultivars under long-term, moderate drought stress revealed an increase of Spm under both stress conditions. While Spm became the most prominent polyamine under drought, levels of all three polyamines were relatively similar under salt stress. Put levels were reduced under both, drought and salt stress, while changes in Spd were different under drought (decrease) or salt (unchanged) conditions. Regulation of polyamine metabolism at the transcript level during exposure to salinity was studied for genes encoding enzymes involved in the biosynthesis of polyamines and compared to expression under drought stress. Based on expression profiles, investigated genes were divided into generally stress-induced genes (ADC2, SPD/SPM2, SPD/SPM3), one generally stress-repressed gene (ADC1), constitutively expressed genes (CPA1, CPA2, CPA4, SAMDC1, SPD/SPM1), specifically drought-induced genes (SAMDC2, AIH), one specifically drought-repressed gene (CPA3) and one specifically salt-stress repressed gene (SAMDC4), revealing both overlapping and specific stress responses under these conditions.

Regulation of polyamine metabolism in Pyropia cinnamomea (W.A. Nelson), an important mechanism for reducing UV-B-induced oxidative damage.

It is generally accepted that ultraviolet (UV) radiation can have adverse affects on phototrophic organisms, independent of ozone depletion. The red intertidal seaweed Pyropia cinnamomea W.A. Nelson (previously Porphyra cinnamomea Sutherland etal. 2011), similar to many other intertidal macrophytes, is exposed to high levels of UV radiation on a daily basis due to emersion in the upper littoral zone. It has been shown that seaweeds, like higher plants, respond to an increased activity of antioxidative enzymes when exposed to stress. However, earlier investigations have shown that P.cinnamomea also compensates for stress due to UV radiation by increasing polyamine (PA) levels, especially bound-soluble and bound-insoluble PAs. The PA precursor putrescine (PUT) can be synthesized via two enzymatic pathways: arginine decarboxylase (ADC) and ornithine decarboxylase (ODC). Both of these enzymes showed increased activity in P.cinnamomea under UV stress. In higher plants, ADC is the enzyme responsible for increased PA levels during stress exposure, while ODC is correlated with cell division and reproduction. However, there are contrary findings in the literature. Using two irreversible inhibitors, we identified the enzyme most likely responsible for increased PUT synthesis and therefore increased stress tolerance in P.cinnamomea. Our results show that changes in the PA synthesis pathway in P.cinnamomea under UV stress are based on an increased activity of ADC. When either inhibitor was added, lipid hydroperoxide levels increased even under photosynthetically active radiation, suggesting that PAs are involved in protection mechanisms under normal light conditions as well. We also show that under optimum or low-stress conditions, ODC activity is correlated with PUT synthesis.

Arginase 2 deficiency reduces hyperoxia-mediated retinal neurodegeneration through the regulation of polyamine metabolism

Hyperoxia treatment has been known to induce neuronal and glial death in the developing central nervous system. Retinopathy of prematurity (ROP) is a devastating disease in premature infants and a major cause of childhood vision impairment. Studies indicate that, in addition to vascular injury, retinal neurons are also affected in ROP. Using an oxygen-induced retinopathy (OIR) mouse model for ROP, we have previously shown that deletion of the arginase 2 (A2) significantly reduced neuro-glial injury and improved retinal function. In the current study, we investigated the mechanism of A2 deficiency-mediated neuroprotection in the OIR retina. Hyperoxia treatment has been known to induce neuronal death in neonates. During the hyperoxia phase of OIR, a significant increase in the number of apoptotic cells was observed in the wild-type (WT) OIR retina compared with A2-deficient OIR. Mass spectrometric analysis showed alterations in polyamine metabolism in WT OIR retina. Further, increased expression level of spermine oxidase was observed in WT OIR retina, suggesting increased oxidation of polyamines in OIR retina. These changes were minimal in A2-deficient OIR retina. Treatment using the polyamine oxidase inhibitor, N, N′-bis (2, 3-butadienyl)-1, 4-butanediamine dihydrochloride, significantly improved neuronal survival during OIR treatment. Our data suggest that retinal arginase is involved in the hyperoxia-induced neuronal degeneration in the OIR model, through the regulation of polyamine metabolism.

Regulation of polyamine metabolism and biosynthetic gene expression during olive mature-fruit abscission

Exogenous ethylene and some inhibitors of polyamine biosynthesis can induce mature-fruit abscission in olive, which could be associated with decreased nitric oxide production as a signaling molecule. Whether H₂O₂ also plays a signaling role in mature-fruit abscission is unknown. The possible involvement of H₂O₂ and polyamine in ethylene-induced mature-fruit abscission was examined in the abscission zone and adjacent cells of two olive cultivars. Endogenous H₂O₂ showed an increase in the abscission zone during mature-fruit abscission, suggesting that accumulated H₂O₂ may participate in abscission signaling. On the other hand, we followed the expression of two genes involved in the polyamine biosynthesis pathway during mature-fruit abscission and in response to ethylene or inhibitors of ethylene and polyamine. OeSAMDC1 and OeSPDS1 were expressed differentially within and between the abscission zones of the two cultivars. OeSAMDC1 showed slightly lower expression in association with mature-fruit abscission. Furthermore, our data show that exogenous ethylene or inhibitors of polyamine encourage the free putrescine pool and decrease the soluble-conjugated spermidine, spermine, homospermidine, and cadaverine in the olive abscission zone, while ethylene inhibition by CoCl₂ increases these soluble conjugates, but does not affect free putrescine. Although the impact of these treatments on polyamine metabolism depends on the cultivar, the results confirm that the mature-fruit abscission may be accompanied by an inhibition of S-adenosyl methionine decarboxylase activity, and the promotion of putrescine synthesis in olive abscission zone, suggesting that endogenous putrescine may play a complementary role to ethylene in the normal course of mature-fruit abscission.

A combined model of hepatic polyamine and sulfur amino acid metabolism to analyze S-adenosyl methionine availability

Many molecular details remain to be uncovered concerning the regulation of polyamine metabolism. A previous model of mammalian polyamine metabolism showed that S-adenosyl methionine availability could play a key role in polyamine homeostasis. To get a deeper insight in this prediction, we have built a combined model by integration of the previously published polyamine model and one-carbon and glutathione metabolism model, published by different research groups. The combined model is robust and it is able to achieve physiological steady-state values, as well as to reproduce the predictions of the individual models. Furthermore, a transition between two versions of our model with new regulatory factors added properly simulates the switch in methionine adenosyl transferase isozymes occurring when the liver enters in proliferative conditions. The combined model is useful to support the previous prediction on the role of S-adenosyl methionine availability in polyamine homeostasis. Furthermore, it could be easily adapted to get deeper insights on the connections of polyamines with energy metabolism.

Regulation of polyamine metabolism by translational control

Polyamines are low molecular weight, positively charged compounds that are ubiquitous in all living cells. They play a crucial role in many biochemical processes including regulation of transcription and translation, modulation of enzyme activities, regulation of ion channels and apoptosis. A strict balance between synthesis, catabolism and excretion tightly controls the cellular concentration of polyamines. The concentrations of rate-limiting enzymes in the polyamine synthesis and degradation pathways are regulated at different levels, including transcription, translation and degradation. Polyamines can modulate the translation of most of the enzymes required for their synthesis and catabolism through feedback mechanisms that are unique for each enzyme. Translational control is associated with cis-acting and trans-acting factors that can be influenced by the concentration of polyamines through mechanisms that are not completely understood. In this review, we present an overview of the translational control mechanisms of the proteins in the polyamine pathway, including ornithine decarboxylase (ODC), ODC antizyme, S-adenosylmethionine decarboxylase and spermidine/spermine N(1) acetyltransferase, highlighting the areas where more research is needed. A better understanding of the translational control of these enzymes would offer the possibility of a novel pharmacological intervention against cancer and other diseases.

Entamoeba histolytica, heterologous expression and in situ immunolocalization of ornithine decarboxylase (EhODC)

Previous studies from this laboratory have dealt with the purification and biochemical characterization of ornithine decarboxylase (ODC) from Entamoeba histolytica. Enzyme compartmentalization has been described as a major mechanism in the regulation of polyamine metabolism. However, the subcellular location of ODC in the human parasite has remained unresolved. To examine this issue, we cloned the full-length gene ( Ehodc) encoding for the parasite enzyme, whose open reading frame encodes for a peptide of 412 amino acids with an estimated molecular mass of 46 kDa that exhibits similarity to other ODCs. Heterologous overexpression of the gene allowed us to purify the recombinant protein (rEhODC) by metal affinity chromatography. The purified polypeptide was used to raise heteroclonal antibodies that were utilized to localize the enzyme in situ by immunofluorescence and confocal microscopy. EhODC was observed to be associated with the plasma membrane, in vesicles close to the plasma membrane and in the EhkOs organelle.

Methylated analogs of spermine and spermidine as tools to investigate cellular functions of polyamines and enzymes of their metabolism

Biogenic amines spermine and spermidine are essential factors of cellular growth. Polyamine analogues are widely used to investigate and to regulate the enzymes of polyamine metabolism and functions of spermine and spermidine in vitro and in vivo. Recently, it was demonstrated that alpha-methylated derivatives of spermine and spermidine are capable to fulfill key cellular functions of polyamines, moreover in some cases of (R)- and (S)-isomers are actually different. Using these alpha-methylated spermine and spermidine analogues it turned possible to prevent the development of acute pancreatitis of SSAT-transgenic rats and to demostrate for the first time that polyamine oxidase, spermine oxidase and deoxyhypusine synthase have dormant stereospecificity. An original approach to regulate the stereospecificity of polyamine oxidase was suggested. It was also demonstrated that the depletion of the intracellular polyamine pool has both hypusine-related consequences and also the consequences unrelated to the posttranslational modification of eukaryotic initiation translation factor eIF5A. Possible applications of a new family of C-methylated polyamine analogues for the investigation and regulation of polyamine metabolism in vitro and in vivo are discussed.

Loss of Methylthioadenosine Phosphorylase and Elevated Ornithine Decarboxylase Is Common in Pancreatic Cancer

Loss of the methylthioadenosine phosphorylase (MTAP) gene at 9p21 is observed frequently in a variety of human cancers. We have shown previously that MTAP can act as a tumor suppressor gene and that its tumor suppressor function is related to its effect on polyamine homeostasis. Ornithine decarboxylase is a key enzyme in the regulation of polyamine metabolism. The aim of this study is to analyze MTAP and ornithine decarboxylase (ODC) expression in primary pancreatic tumor specimens. We measured MTAP and ODC activity in protein extracts derived from 30 surgically resected tumor samples and eight normal pancreas samples. In a subset of six samples, we also examined MTAP DNA using interphase fluorescence in situ hybridization. In addition, we examined the effect of the ODC inhibitor difluoromethylornithine on two pancreatic adenocarcinoma-derived cell lines. MTAP activity was 2.8-fold reduced in adenocarcinomas and 6.3-fold reduced in neuroendocrine tumors compared with control pancreas. Conversely, ODC activity was 3.6-fold elevated in adenocarcinomas and 3.9-fold elevated in neuroendocrine tumors compared with control pancreas. Using interphase fluorescence in situ hybridization, we found in tumor samples that 43 to 75% of the nuclei had lost at least one copy of MTAP locus, indicating that loss of MTAP activity was at least partially because of deletion of the MTAP locus. We also show that inhibition of ODC by difluoromethylornithine caused decreased cell growth and increased apoptosis in two MTAP-deleted pancreatic adenocarcinoma-derived cell lines. MTAP activity is frequently lost, and ODC activity is frequently elevated in both pancreatic adenocarcinoma and neuroendocrine tumors. Inhibition of ODC activity caused decreased cell growth and increased apoptosis in pancreatic tumor-derived cell lines. These findings suggest that MTAP and polyamine metabolism could be potential therapeutic targets in the treatment of pancreatic cancer.