Novel N-Acylethanolamide Derivatives Affect Body Weight and Energy Balance.

Introduction - The obesity pandemic is multifactorial. Nutritional, pharmacologic and surgical interventions are limited in reach and efficacy, raising need for new therapeutics. Aims - Characterization of anorexigenic and cognitive effect and central mechanism of action of novel N-acylethanolamide derivatives. Methods - Sabra mice divided to similar experimental groups, injected IP with: oleyl-L-leucinolamide (1 A), linoleyl-L-leucinolamide (4 A), linoleyl-L-valinolamide (5 A), oleyl-oxycarbonyl-L-valinolamide (1 B), oleyl-oxycarbonyl-D-valinolamide (2 B), oleylamine-carbonyl-L-valinolamide (3 B), oleylamine-carbonyl-D-valinolamide (4 B), and oleyl-L-hydroxyvalineamide (5 B). Control group with vehicle. Body weight and food consumption followed for 39 days. Motor activity and cognitive function by open field test and eight-arm maze. Mice sacrificed and mechanism of action investigated by qPCR. The genes analyzed involved in energy balance and regulation of appetite. Catecholamines and serotonin evaluated. Results - Compounds 1 A, 5 A, 1 B-4 B, caused significant weight loss of 4.2-5.6 % and 5 A, 1 B-4 B, improved cognitive function following 8 i. p. injections of 1 mg/kg during 39 days, by different mechanisms. 5 A, 3 B and 4 B decreased food consumption, whereas 1 A, 5 A and 2 B increased motor activity. 1 A, 4 A, 1 B and 3 B elevated SIRT-1, associated with survival. POMC upregulated by 1 B and 2 B, CART by 1 B, 2 B and 1 A. NPY and CAMKK2 downregulated by 5 A. 4 B enhanced 5-HT levels. 4 A, 5 A, 1 B, 4 B, 5 B decreased FAAH, showing long lasting effect. Conclusions - These new compounds might be developed for the treatment of obesity and for improved cognitive function.

CD5 blockade enhances ex vivo CD8+ T cell activation and tumour cell cytotoxicity.

CD5 is expressed on T cells and a subset of B cells (B1a). It can attenuate TCR signalling and impair CTL activation and is a therapeutic targetable tumour antigen expressed on leukemic T and B cells. However, the potential therapeutic effect of functionally blocking CD5 to increase T cell anti-tumour activity against tumours (including solid tumours) has not been explored. CD5 knockout mice show increased anti-tumour immunity: reducing CD5 on CTLs may be therapeutically beneficial to enhance the anti-tumour response. Here, we show that ex vivo administration of a function-blocking anti-CD5 MAb to primary mouse CTLs of both tumour-naïve mice and mice bearing murine 4T1 breast tumour homografts enhanced their capacity to respond to activation by treatment with anti-CD3/anti-CD28 MAbs or 4T1 tumour cell lysates. Furthermore, it enhanced TCR signalling (ERK activation) and increased markers of T cell activation, including proliferation, CD69 levels, IFN-γ production, apoptosis and Fas receptor and Fas ligand levels. Finally, CD5 function-blocking MAb treatment enhanced the capacity of CD8+ T cells to kill 4T1-mouse tumour cells in an ex vivo assay. These data support the potential of blockade of CD5 function to enhance T cell-mediated anti-tumour immunity.

Platinum (IV)-fatty acid conjugates overcome inherently and acquired Cisplatin resistant cancer cell lines: an in-vitro study.

BACKGROUND: Platinum-based drugs are used as cancer chemotherapeutics for the last 40 years. However, drug resistance and nephrotoxicity are the major limitations of the use of platinum-based compounds in cancer therapy. Platinum (IV) complexes are believed to act as platinum prodrugs and are able to overcome some of platinum (II) limitations. METHODS: A number of previously sensitized platinum (IV) complexes were evaluated for their anti-cancer activity by monitoring ability to affect proliferation, clonigenicity and apoptosis induction of Cisplatin sensitive and resistant cancer cells. In addition, the uptake of Cisplatin and the platinum (IV) derivatives to Cisplatin sensitive and resistant cancer cells was monitored. RESULTS: The bis-octanoatoplatinum (IV) complex (RJY13), a Cisplatin derivative with octanoate as axial ligand, exhibited strong anti-proliferative effect on the Cisplatin resistant and sensitive ovarian cells, A2780cisR and A2780, respectively. Moreover, RJY13 exhibited good activity in inhibiting clonigenicity of both cells. Anti-proliferative activity of RJY13 was mediated by induction of apoptosis. Interestingly, a bis-lauratopaltinum (IV) complex (RJY6) was highly potent in inhibiting clonigenicity of both Cisplatin sensitive and Cisplatin resistant cells, however, exhibited reduced activity in assays that utilize cells growing in two dimensional (2D) conditions. The uptake of Cisplatin was reduced by 30% in A2780 in which the copper transporter-1 (Ctr1) was silenced. Moreover, uptake of RJY6 was marginally dependent on Ctr1, while uptake of RJY13 was Ctr1-independent. CONCLUSIONS: Our data demonstrated the potential of platinum (IV) prodrugs in overcoming acquired and inherited drug resistance in cancer cell lines. Moreover, our data demonstrated that the uptake of Cisplatin is partially dependent on Ctr1 transporter, while uptake of RJY6 is marginally dependent on Ctr1 and RJY13 is Ctr1-independent. In addition, our data illustrated the therapeutic potential of platinum (IV) prodrugs in cancer therapy.

Facile preparation of mono-, di- and mixed-carboxylato platinum(IV) complexes for versatile anticancer prodrug design.

Facile strategies were developed for the versatile functionalization of platinum(IV) axial sites, allowing for easy accessibility to unsymmetric mono- and mixed-carboxylato, as well as symmetric di-substituted platinum(IV) complexes. The first method involves the direct oxidation and carboxylation of the platinum(II) center using an appropriate peroxide and the carboxylate of choice to firstly yield a monocarboxylato monohydroxido platinum(IV) complex. This platinum(IV) intermediate can undergo further carboxylation to give rise to a mixed-carboxylato platinum(IV) complex. The second method involves the activation of the carboxylate of choice by a common carbodiimide coupling reagent, and its reaction with a dihydroxido platinum(IV) precursor to give the monocarboxylato platinum(IV) complex. Uronium salts can be employed to promote efficient dicarboxylation of the dihydroxido platinum(IV) precursor. Lastly, an axial azide pendant group was demonstrated to be suitable for orthogonal "click" conjugation reactions.

Oleylamine-carbonyl-valinol inhibits auto-phosphorylation activity of native and T315I mutated Bcr-Abl, and exhibits selectivity towards oncogenic Bcr-Abl in SupB15 ALL cell lines.

Chronic myeloid leukemia (CML) is characterized by the presence of p210(Bcr-Abl) which exhibits an abnormal kinase activity. Selective Abl kinase inhibitors have been successfully established for the treatment of CML. Despite high rates of clinical response, CML patients can develop resistance against these kinase inhibitors mainly due to point mutations within the Abl protein kinase domain. Previously, we have identified oleic acid as the active component in the mushroom Daedalea gibbosa that inhibited the kinase activity of Bcr-Abl. Here, we report that the oleyl amine derivatives, S-1-(1-Hydroxymethyl-2-methyl-propyl)-3-octadec-9-enyl-urea [oleylaminocarbonyl-L-N-valinol,oroleylaminocarbonyl-S-2-isopropyl-N-ethanolamine,oleylamine-carbonyl-L-valinol] (cpd 6) and R-1-(1-Hydroxymethyl-2-methyl-propyl)-3-octadec-9-enyl-urea [oleylamineocarbonyl-D-N-valinol, oleylaminocarbonyl-R-2-isopropyl-N-ethanolamine, or oleylamine-carbonyl-D-valinol] (cpd 7), inhibited the activity of the native and T315I mutated Bcr-Abl. Furthermore, cpd 6 and 7 exhibited higher activity towards the oncogenic Bcr-Abl in comparison to native c-Abl in SupB15 Ph-positive ALL cell line.

p185(BCR/ABL) has a lower sensitivity than p210(BCR/ABL) to the allosteric inhibitor GNF-2 in Philadelphia chromosome-positive acute lymphatic leukemia.

BACKGROUND: The t(9;22) translocation leads to the formation of the chimeric breakpoint cluster region/c-abl oncogene 1 (BCR/ABL) fusion gene on der22, the Philadelphia chromosome. The p185(BCR/ABL) or the p210(BCR/ABL) fusion proteins are encoded as a result of the translocation, depending on whether a "minor" or "major" breakpoint occurs, respectively. Both p185(BCR/ABL) and p210(BCR/ABL) exhibit constitutively activated ABL kinase activity. Through fusion to BCR the ABL kinase in p185(BCR/ABL) and p210(BCR/ABL) "escapes" the auto-inhibition mechanisms of c-ABL, such as allosteric inhibition. A novel class of compounds including GNF-2 restores allosteric inhibition of the kinase activity and the transformation potential of BCR/ABL. Here we investigated whether there are differences between p185(BCR/ABL) and p210(BCR/ABL) regarding their sensitivity towards allosteric inhibition by GNF-2 in models of Philadelphia chromosome-positive acute lymphatic leukemia. DESIGN AND METHODS: We investigated the anti-proliferative activity of GNF-2 in different Philadelphia chromosome-positive acute lymphatic leukemia models, such as cell lines, patient-derived long-term cultures and factor-dependent lymphatic Ba/F3 cells expressing either p185(BCR/ABL) or p210(BCR/ABL) and their resistance mutants. RESULTS: The inhibitory effects of GNF-2 differed constantly between p185(BCR/ABL) and p210(BCR/ABL) expressing cells. In all three Philadelphia chromosome-positive acute lymphatic leukemia models, p210(BCR/ABL)-transformed cells were more sensitive to GNF-2 than were p185BCR/ABL-positive cells. Similar results were obtained for p185(BCR/ABL) and the p210(BCR/ABL) harboring resistance mutations. CONCLUSIONS: Our data provide the first evidence of a differential response of p185(BCR/ABL)- and p210(BCR/ABL)- transformed cells to allosteric inhibition by GNF-2, which is of importance for the treatment of patients with Philadelphia chromosome-positive acute lymphatic leukemia.

Overcoming Bcr-Abl T315I mutation by combination of GNF-2 and ATP competitors in an Abl-independent mechanism.

BACKGROUND: Philadelphia positive leukemias are characterized by the presence of Bcr-Abl fusion protein which exhibits an abnormal kinase activity. Selective Abl kinase inhibitors have been successfully established for the treatment of Ph (+) leukemias. Despite high rates of clinical response, Ph (+) patients can develop resistance against these kinase inhibitors mainly due to point mutations within the Abl protein. Of special interest is the 'gatekeeper' T315I mutation, which confers complete resistance to Abl kinase inhibitors. Recently, GNF-2, Abl allosteric kinase inhibitor, was demonstrated to possess cellular activity against Bcr-Abl transformed cells. Similarly to Abl kinase inhibitors (AKIs), GNF-2 failed to inhibit activity of mutated Bcr-Abl carrying the T315I mutation. METHODS: Ba/F3 cells harboring native or T315I mutated Bcr-Abl constructs were treated with GNF-2 and AKIs. We monitored the effect of GNF-2 with AKIs on the proliferation and clonigenicity of the different Ba/F3 cells. In addition, we monitored the auto-phosphorylation activity of Bcr-Abl and JAK2 in cells treated with GNF-2 and AKIs. RESULTS: In this study, we report a cooperation between AKIs and GNF-2 in inhibiting proliferation and clonigenicity of Ba/F3 cells carrying T315I mutated Bcr-Abl. Interestingly, cooperation was most evident between Dasatinib and GNF-2. Furthermore, we showed that GNF-2 was moderately active in inhibiting the activity of JAK2 kinase, and presence of AKIs augmented GNF-2 activity. CONCLUSIONS: Our data illustrated the ability of allosteric inhibitors such as GNF-2 to cooperate with AKIs to overcome T315I mutation by Bcr-Abl-independent mechanisms, providing a possibility of enhancing AKIs efficacy and overcoming resistance in Ph+ leukemia cells.

Allosteric inhibition enhances the efficacy of ABL kinase inhibitors to target unmutated BCR-ABL and BCR-ABL-T315I.

BACKGROUND: Chronic myelogenous leukemia (CML) and Philadelphia chromosome-positive (Ph+) acute lymphatic leukemia (Ph + ALL) are caused by the t(9;22), which fuses BCR to ABL resulting in deregulated ABL-tyrosine kinase activity. The constitutively activated BCR/ABL-kinase "escapes" the auto-inhibition mechanisms of c-ABL, such as allosteric inhibition. The ABL-kinase inhibitors (AKIs) Imatinib, Nilotinib or Dasatinib, which target the ATP-binding site, are effective in Ph + leukemia. Another molecular therapy approach targeting BCR/ABL restores allosteric inhibition. Given the fact that all AKIs fail to inhibit BCR/ABL harboring the 'gatekeeper' mutation T315I, we investigated the effects of AKIs in combination with the allosteric inhibitor GNF2 in Ph + leukemia. METHODS: The efficacy of this approach on the leukemogenic potential of BCR/ABL was studied in Ba/F3 cells, primary murine bone marrow cells, and untransformed Rat-1 fibroblasts expressing BCR/ABL or BCR/ABL-T315I as well as in patient-derived long-term cultures (PDLTC) from Ph + ALL-patients. RESULTS: Here, we show that GNF-2 increased the effects of AKIs on unmutated BCR/ABL. Interestingly, the combination of Dasatinib and GNF-2 overcame resistance of BCR/ABL-T315I in all models used in a synergistic manner. CONCLUSIONS: Our observations establish a new approach for the molecular targeting of BCR/ABL and its resistant mutants using a combination of AKIs and allosteric inhibitors.

DNA interactions of new cytotoxic tetrafunctional dinuclear platinum complex trans,trans-[{PtCl2(NH3)}2(piperazine)].

A new tetrafunctional dinuclear platinum complex trans,trans-[{PtCl2(NH3)}2(piperazine)] with sterically rigid linking group was designed, synthesized and characterized. In this novel molecule, the DNA-binding features of two classes of the platinum compounds with proven antitumor activity are combined, namely trans oriented bifunctional mononuclear platinum complexes with a heterocyclic ligand and polynuclear platinum complexes. DNA-binding mode of this new complex was analyzed by various methods of molecular biology and biophysics. The complex coordinates DNA in a unique way and interstrand and intrastrand cross-links are the predominant lesions formed in DNA in cell-free media and in absence of proteins. An intriguing aspect of trans,trans-[{PtCl2(NH3)}2(piperazine)] is that, using a semi-rigid linker, interstrand cross-linking is diminished relative to other dinuclear platinum complexes with flexible linking groups and lesions that span several base pairs, such as tri- and tetrafunctional adducts, become unlikely. In addition, in contrast to the inability of trans,trans-[{PtCl2(NH3)}2(piperazine)] to cross-link two DNA duplexes, the results of the present work convincingly demonstrate that this dinuclear platinum complex forms specific DNA lesions which can efficiently cross-link proteins to DNA. The results substantiate the view that trans,trans-[{PtCl2(NH3)}2(piperazine)] or its analogues could be used as a tool for studies of DNA properties and their interactions or as a potential antitumor agent. The latter view is also corroborated by the observation that trans,trans-[{PtCl2(NH3)}2(piperazine)] is a more effective cytotoxic agent than cisplatin against human tumor ovarian cell lines.

Cationic nonsymmetric transplatinum complexes with piperidinopiperidine ligands. Preparation, characterization, in vitro cytotoxicity, in vivo toxicity, and anticancer efficacy studies.

A series of complexes of the general formula trans-[PtCl2(Am)(pip-pip)] x HCl where pip-pip is 4-piperidinopiperidine and Am is NH3, methylamine (MA), dimethylamine (DMA), n-propylamine (NPA), isopropylamine (IPA), n-butylamine (NBA), or cyclohexylamine (CHA) were prepared and characterized, and their cytotoxic properties against ovarian and colon cancer cells were evaluated. The trans-[PtCl2(NH3)(pip-pip)] x HCl was significantly more potent than cisplatin in all the cisplatin-resistant ovarian cancer cell lines and was nearly as cytotoxic as cisplatin against colon cancer cells. In vivo studies in mice showed that the pip-pip complexes are significantly less toxic than cisplatin. Cisplatin was more efficacious than both trans-[PtCl2(NH3)(pip-pip)] x HCl and trans-[PtCl2(NBA)(pip-pip)] x HCl in the A2780 and A2780cisR tumor xenograft models, consistent with its lower IC50 values in A2780 cells but contrary to the higher IC50 values in A2780cisR cells. In the colon cancer cell studies, trans-[PtCl2(NH3)(pip-pip)] x HCl was slightly less potent than cisplatin in the in vitro studies but had efficacy comparable to that of cisplatin in the in vivo xenograft model.

Interactions of platinum complexes containing cationic, bicyclic, nonplanar piperidinopiperidine ligands with biological nucleophiles.

The determination of the structures and DNA interactions and the reactions with GSH and ubiquitin of complexes of the general formula trans-[PtCl2(Am)(pip-pip)] x HCl, where pip-pip is 4-piperidinopiperidine and Am is NH3, methylamine (MA), dimethylamine (DMA), n-propylamine (NPA), isopropylamine (IPA), n-butylamine (NBA), or cyclohexylamine (CHA), were performed. X-ray structures and NMR studies of the NH3 and MA complexes showed that both pip rings were in the chair conformation and that the second pip ring is fluxional. The DNA binding studies showed that these complexes bind to calf thymus DNA nearly an order of magnitude more quickly than cisplatin and form covalent adducts that stabilize the double helix. The binding of the pip-pip complexes to DNA results in high unwinding angles (approximately 30 degrees) and in the formation of approximately 25% interstrand cross-links. The pip-pip complexes reacted with GSH more quickly than cisplatin and transplatin, and the rate of reaction decreased with increasing steric bulk of the ligand trans to the pip-pip. The reactions with ubiquitin resulted in monofunctional binding to Met1. Only the NH3, MA, and DMA complexes reacted with ubiquitin in a slower and less efficient fashion than cisplatin.

Preparation, cytotoxicity and interactions with nucleophiles of three isomeric transplatinum complexes containing methylpiperidine ligands.

Three isomeric complexes, trans-[PtCl2(NH3)(2-methylpiperidine)], trans-[PtCl2(NH3)(3-methylpiperidine)] and trans-[PtCl2(NH3)(4-methylpiperidine)], were prepared and their cytotoxicities against six ovarian cancer cell lines, three sensitive and three resistant to cisplatin, were measured. There were no significant differences in the cytotoxicities of the three isomers against these cell lines. The interactions of the three complexes with reduced glutathione (GSH) and with ubiquitin (Ub), as a model protein, were studied. The trans-[PtCl2(NH3)(2-methylpiperidine)] reacted approximately twice as slowly with GSH as did the other two isomers. In the 1:1 interactions of the three complexes with ubiquitin (Mr = 8565 amu), trans-[PtCl2(NH3)(3-methylpiperidine)] and trans-[PtCl2(NH3)(4-methylpiperidine)] attained 100% modification while trans-[PtCl2(NH3)(2-methylpiperidine)] reached only less than 50% modification. Trans-[PtCl2(NH3)(2-methylpiperidine)] reacts significantly less efficiently with GSH and proteins than the other two isomers yet this is not reflected in the cytotoxicity values. These results indicate that for these complexes, in these cell lines, cytosolic detoxification probably does not play a dominant role in determining the cytotoxicity of the complexes.

Water soluble cationic trans-platinum complexes which induce programmed cell death in the protozoan parasite Leishmania infantum.

We have evaluated the cytotoxic properties against the protozoan Leishmania infantum of four water soluble cationic trans-Pt(II)Cl(2) compounds containing as inert groups NH3 and piperazine (1), 4-picoline and piperazine (2), n-butylamine and piperazine (3), and NH3 and 4-piperidino-piperidine (4). The leishmanicidal activity of compounds 3 and 4 against promastigotes of the parasite Leishmania infantum was 2.5- and 1.6-times higher than that of the cytotoxic drug cis-diamminedichloroplatinum(II), respectively. Interestingly, compounds 3 and 4 produce in Leishmania infantum promastigotes a higher amount of programmed cell death than cisplatin, which is associated with cell cycle arrest in G2/M. In contrast to cis-diamminedichloroplatinum(II), binding of compounds 3 and 4 to calf thymus DNA induces conformational changes more similar to those of trans-diamminedichloroplatinum(II) that may be attributed to denaturation of the double helix. Similarly to cis-diamminedichloroplatinum(II) and trans-diamminedichloroplatinum(II), the interaction of compounds 3 and 4 with ubiquitin results in an increase of the alpha-helix content of the protein as observed by circular dichroism spectroscopy. However, fluorescence studies indicate that compounds 3 and 4 produce a decrease in the fluorescence of the tyrosine 59 residue of ubiquitin higher than both cis-diamminedichloroplatinum(II) and trans-diamminedichloroplatinum(II). Altogether, our results suggest that the biochemical mechanism of cytotoxic activity of compounds 3 and 4 against Leishmania infantum must be different from that of cis-diamminedichloroplatinum(II). To the best of our knowledge, compounds 3 and 4 are the first reported trans-platinum complexes that show antiparasitic activity.

Novel apoptosis-inducing trans-platinum piperidine derivatives: synthesis and biological characterization.

The synthesis, chemical characterization, and interaction with cells of new sterically hindered trans- and cis-diaminedichloroplatinum(II) complexes are described. The amine ligands include monofunctional piperidine (pip) and piperazine (pz). The poor solubility of trans-diaminedichloroplatinum complexes was overcome by introducing the positively charged pz ligand, which allows retaining of the classic platinum coordination sphere. In vitro evaluation in OV-1063 and C-26 tumor cells revealed that replacing one NH3 of the inactive transplatin by an aromatic planar ligand (4-picoline, 4-pic) or by an aliphatic nonplanar heterocyclic ligand (pip) or replacing both NH3 groups with a 4-pic ligand and a pip or pz ligand significantly increases the cytotoxic activity of these complexes. The unsymmetric complexes trans-[PtCl2(4-pic)(pip)] and trans-[PtCl2(4-pic)(pz)]HCl were the most cytotoxic compounds against the cisplatin-sensitive tumor cell line C-26 (IC50 = 4.5 and 5.5 microM, respectively) and the cisplatin-sensitive tumor cell line OV-1063 (IC50 = 6.5 and 7.4 microM, respectively). In contrast, replacing one NH3 of the cis isomer by an aromatic planar ligand (4-pic) or by an aliphatic amine lowered their cytotoxiciy in comparison to cisplatin. Cell penetration and Pt-DNA adduct formation were also evaluated, and it was clearly shown that both trans-[PtCl2(4-pic)(pip)] and trans-[PtCl2(4-pic)(pz)]HCl penetrate efficiently the cellular membrane of the tumor cells and platinate the cellular DNA. When comparing cellular DNA platination, positively charged trans-[PtCl2(4-pic)(pz)]HCl was 7-fold higher than both cisplatin and its neutral analogue trans-[PtCl2(4-pic)(pip)]. Moreover, in contrast to cisplatin, in the cell lines used, cell death caused by both complexes appeared to be apoptotic according to several criteria including early phosphatidylserine exposure, activation of caspases, and characteristic morphological changes. Our results suggest that these novel mixed nonclassical trans-Pt(II) complexes are biologically and mechanistically distinct from known Pt complexes and deserve evaluation of their efficacy in tumor-bearing animals.

Novel soluble cationic trans-diaminedichloroplatinum(II) complexes that are active against cisplatin resistant ovarian cancer cell lines.

Positively charged, water soluble cis/trans-[PtCl(2)(piperazine)(Am1)] (where Am1 = NH(3), n-butylamine, isopropylamine, 4-picoline, piperidine, and piperazine) has significant cytotoxic activity against cisplatin resistant ovarian cancer cells. The charged complexes are taken up by cancer cells much more rapidly than cisplatin and bind to cellular DNA and to calf thymus DNA much faster than cisplatin or transplatin. The platinum-piperazine complexes bind proteins (ubiquitin and myoglobin) very slowly as compared to cisplatin and to their neutral piperidine analogues. Altogether, the results reported here suggest that combination of positively charged ligands with a trans-Pt(II)Cl(2) center may lead to the discovery of platinum complexes that are able to circumvent cisplatin resistance.

Decreased levels of osteopontin and bone sialoprotein II are correlated with reduced proliferation, colony formation, and migration of GFP-MDA-MB-231 cells.

MDA-MB-231 human breast cancer cells transfected with GFP were used as model to determine the reduction in proliferation, colony formation, and migration in response to agents with anti-metastatic properties. These agents consisted of antisense oligonucleotides (ASOs) directed against osteopontin (OPN), bone sialoprotein II (BSP II), and osteonectin (ON), as well as an antibody directed against BSP II. A bisphosphonate derivative (ibandronate) and an alkylphosphocholine (erucylphospho-NNN-trimethylpropanolamine; ErPC3) were used as positive controls. The ASOs directed against OPN, BSP II and ON suppressed the expression of their respective target proteins by 81%, 74% and 69%, respectively. They were barely but significantly active in inhibiting the proliferation, but intermediately to highly active in inhibiting the colony formation and migration of GFP-MDA-MB-231 breast cancer cells. The antibody against human BSP II was significantly more active than all ASOs used and was equally active or even surpassed the activity of ibandronate and ErPC3 in all three assays. The results obtained suggest a specific anti-metastatic activity of this antibody as well as of the ASOs found effective in decreasing OPN and BSP II expression.

Interactions of cisplatin and transplatin with proteins. Comparison of binding kinetics, binding sites and reactivity of the Pt-protein adducts of cisplatin and transplatin towards biological nucleophiles.

In this manuscript we report on the interactions of cis-DDP (cisplatin, cis-diamminedichloroplatinum(II)) and trans-DDP (transplatin, trans-diamminedichloroplatinum(II)) with two model proteins, ubiquitin (Ub) and horse heart myoglobin (Mb), and attempt to answer the question whether proteins that have methionine-Pt adducts can transfer the platinum to biological nucleophiles and particularly to DNA. Our study shows that cisplatin and transplatin form different adducts with ubiquitin: transplatin forms one major adduct, trans-[Pt(Ub)(NH(3))(2)Cl], while cisplatin forms four distinct adducts, [Pt(Ub)(NH(3))(2)Cl], [Pt(Ub)(NH(3))(2)(H(2)O)], [Pt(Ub)(NH(3))(2)], and [Pt(Ub)(NH(3))]. When binding ubiquitin, Met1 is the preferred binding site of cisplatin, but not of transplatin. Cisplatin binds faster than transplatin to both ubiquitin and horse heart myoglobin. Both cisplatin and transplatin adducts form stable ternary adducts when reacted with 5'-guanosine monophosphate (5'-GMP) or a tetranucleotide. No transfer of the Pt moiety from the proteins to the nucleotides was observed. Glutathione efficiently removes the platinum from preformed adducts of both cisplatin and transplatin with ubiquitin.

Novel acylethanolamide derivatives that modulate body weight through enhancement of hypothalamic pro-opiomelanocortin (POMC) and/or decreased neuropeptide Y (NPY).

Newly synthesized acylethanolamide derivatives oleoyl-L-valinolamide (1), oleoyl-D-valinolamide (2), elaidoyl-L-valinolamide (3), elaidoyl-D-valinolamide (4) stearoyl-L-valinolamide (5), and palmitoyl-L-valinolamide (6) were investigated in mice as antiobesity compounds. Compounds 1, 2, 5, 6 significantly decreased body weight by 6.57% following eight injections of 1 mg/kg i.p. during 39 days, while 3 and 4 showed no such activity. Receptor binding indicated that no compound activated CB1, CB2, PPARα, or TRPV1 receptors. Hypothalamic RT-PCR showed that mRNA expression of the anorexigenic genes POMC and CART was up-regulated by 1, 2, 5 and 1, 2, respectively, while that of the orexigenic genes NPY and CaMKK2 was down-regulated by the respective compounds 1, 5, 6 and 1, 2, 5. Oleoyl-L-valinolamide enhances anorectic pathways and lead to decreased glucose levels, enhanced locomotor activity, and improved cognition. Effects of oleoyl-L-valinolamide on weight were dose-dependent, and it could be given orally. 1, 2, 4, 5 down-regulated FAAH mRNA expression.

Oleic acid is the active component in the mushroom Daedalea gibbosa inhibiting Bcr-Abl kinase autophosphorylation activity.

The hallmark of chronic myeloid leukemia (CML) is the abnormal activity of p210(Bcr-Abl) kinase. Selective kinase inhibitors such as imatinib or nilotinib have been established successfully for the treatment of CML. Despite high rates of clinical response, CML patients can develop resistance to these kinase inhibitors mainly due to point mutations within the Abl kinase domain of the fusion protein. Previously, we reported that a crude extract of the mushroom Daedalea gibbosa inhibited kinase activity of Bcr-Abl kinase. Here we report on the identification of the active component of Daedalea gibbosa, oleic acid, which inhibited Bcr-Abl kinase autophosphorylation in Ba/F3 cells and exhibited anti-CML activity in a BCR/ABL-positive mouse model.