Synthesis and biological evaluation in vitro of selective,high affinity peptide antagonists of human melanin-concentrating hormone action at human melanin-concentrating hormone receptor 1

Human melanin-concentrating hormone (hMCH) and many of its analogues are potent but nonspecific ligands for human melanin-concentrating hormone receptors 1 and 2 (hMCH-1R and hMCH-2R). To differentiate between the physiological functions of these receptors, selective antagonists are needed. In this study, analogues of Ac-Arg(6)-cyclo(S-S)(Cys(7)-Met(8)-Leu(9)-Gly(10)-Arg(11)-Val(12)-Tyr(13)-Arg(14)-Pro(15)-Cys(16))-NH(2), a high affinity but nonselective agonist at hMCH-1R and hMCH-2R, were prepared and tested in binding and functional assays on cells expressing these receptors. In the new analogues, 5-aminovaleric acid (Ava) was incorporated in place of the Leu(9)-Gly(10) and/or Arg(14)-Pro(15) segments of the disulfide ring. Several of these compounds turned out to be high affinity antagonists selective for hMCH-1R. Moreover, even at micromolar concentrations, they were devoid of agonist potency at both hMCH receptors and not effective as hMCH-2R antagonists. For example, peptide 14, Gva(6)- cyclo(S-S)(Cys(7)-Met(8)-Leu(9)-Gly(10)-Arg(11)-Val(12)-Tyr(13)-Ava(14,15)-Cys(16))-NH(2), (Gva = 5-guanidinovaleric acid), was a full competitive hMCH-1R antagonist (IC(50) = 14 nM, K(B) = 0.9 nM) with more than 1000-fold selectivity over hMCH-2R. Examination of various compounds with Ava in positions 9,10 and/or 14,15 revealed that the Leu(9)-Gly(10) and Arg(14)-Pro(15) segments of the disulfide ring are the principal structural elements determining hMCH-1R selectivity and ability to act as a hMCH-1R antagonist.     

Synthesis and evaluation of pyridone-phenoxypropyl-R-2-methylpyrrolidine analogues as histamine H3 receptor antagonists

6-{4-[3-(R)-2-Methylpyrrolidin-1-yl)propoxy]-phenyl}-2H-pyridazin-3-one 6 (Irdabisant; CEP-26401) was recently reported as a potent H(3)R antagonist with excellent drug-like properties and in vivo activity that advanced into clinical evaluation. A series of pyridone analogs of 6 was synthesized and evaluated as H(3)R antagonists. Structure-activity relationships revealed that the 5-pyridone regiomer was optimal for H(3)R affinity. N-Methyl 9b showed excellent H(3)R affinity, acceptable pharmacokinetics and pharmaceutical properties. In vivo evaluation of 9b showed potent activity in the rat dipsogenia model and robust wake-promoting activity in the rat EEG model.     

Induction of B1 receptors in streptozotocin diabetic rats: possible involvement in the control of hyperglycemia-induced glomerular Erk 1 and 2 phosphorylation

We investigated the effects of a 3-week treatment with various combinations of angiotensin-converting enzyme inhibitor (ACEI) and B1 and B2 bradykinin receptor (B1R and B2R) antagonists (B1A and B2A) and AT1 receptor antagonist on ERK 1 and 2 phosphorylation in isolated glomeruli from streptozotocin-treated diabetic rats (STZ rats). Body weight, glycemia, and blood pressure were monitored. The rats were divided into nine groups: (1) control; and groups 2-9 were STZ treated with (3) insulin, (4) ACEI, (5) ACEI + B1A, (6) ACEI + B2A, (7) B2A, (8) B1A, (9) AT1 antagonist. ERK 1 and 2 phosphorylation and expression of B1R and B2R were assessed by Western blot analysis. ERK 1 and 2 phosphorylation was higher in STZ rats; this activation was normalized by insulin and reduced by ACEI but not by AT1 antagonist. The reduction of ERK 1 and 2 phosphorylation by the ACEI was reversed by B1A and B2A. The induction of B1R was confirmed by increased expression of mRNA and B1 receptor protein. Since ERK 1 and 2 phosphorylation is an early event in the induction of matrix secretion and hyperproliferation associated with diabetic nephropathy, activation of B1R and B2R appears to be a useful pharmacological target in the management of this pathology.     

Two small molecule agonists of glucagon-like peptide-1 receptor modulate the receptor activation response differently

The glucagon-like peptide-1 receptor (GLP-1R) is a target for type 2 diabetes treatment. Due to the inconvenience of peptide therapeutics, small-molecule GLP-1R agonists have been studied. Compound 2 (6,7-dichloro-2-methylsulfonyl-2-N-tert-butylaminoquinoxaline) and compound B (4-(3-(benzyloxy)phenyl)-2-(ethylsulfinyl)-6-(trifluoromethyl)pyrimidine) have been described as small molecule, ago-allosteric modulators of GLP-1R. However, their modes of action at the GLP-1R have not been elucidated. Thus, in this study, we compared the mechanisms of action between these two compounds. When compound 2 was treated with endogenous or exogenous peptide agonists (GLP-1 and exenatide) or fragments of peptide agonists (GLP-1(9-36), Ex3, Ex4, and Ex5), the response curve of these peptide agonists shifted left without a change in maximum efficacy. In contrast, compound B potentiated the response and increased maximum efficacy. However, N-terminal truncated orthosteric antagonists including Ex7, Ex9, and Ex10, augmented the response of compound 2 at the GLP-1R but did not alter compound B activity. Intriguingly, when we co-treated compound 2 with compound B in CHO cells expressing full-length hGLP-1R or N-terminal extracellular domain-truncated GLP-1R, the activation of both types of receptors increased additively, implying that the N-terminus of the receptor is not involved in the modulation by compound agonists. We confirmed that these two compounds increased calcium influx by different patterns in CHO cells expressing GLP-1R. Taken together, our findings suggest that compounds 2 and B have different modes of action to activate GLP-1R. Further study to identify the putative binding sites will help in the discovery of orally available GLP-1R agonists.     

The discovery of potent antagonists of NPBWR1 (GPR7)

The synthesis and evaluation of small molecule antagonists of the G protein-coupled receptor NPBWR1 (GPR7) are reported for the first time. [4-(5-Chloropyridin-2-yl)piperazin-1-yl][(1S,2S,4R)-4-{[(1R)-1-(4-methoxyphenyl)ethyl]amino}-2-(thiophen-3-yl)cyclohexyl]methanone (1) emerged as a hit from a high-throughput screen. Examination of substituents that focused on replacing the 5-chloropyridine and 4-methoxybenzylamino groups of 1 led to the identification of compounds that exhibited subnanomolar potencies as low as 660pM (9k) in the functional assay and 200pM in the binding assay (9i).     

Design, synthesis, and structure-activity relationships of a series of 2-Ar-8-methyl-5-alkylaminoquinolines as novel CRF(1) receptor antagonists

We designed and synthesized a series of 2-Ar-8-methyl-5-alkylaminolquinolines as potent corticotropin-releasing factor 1 (CRF(1)) receptor antagonists. The structure-activity relationships of substituents at each position (R(3), R(5), R(5'), and R(8)) was investigated. By derivatization, three compounds (6, 14b, and 14c) were identified as orally active CRF(1) receptor antagonists.     

N-oleoyldopamine, a novel endogenous capsaicin-like lipid, protects the heart against ischemia-reperfusion injury via activation of TRPV1

N-oleoyldopamine (OLDA), a bioactive lipid originally found in the mammalian brain, is an endovanilloid that selectively activates the transient receptor potential vanilloid type 1 (TRPV1) channel. This study tests the hypothesis that OLDA protects the heart against ischemia and reperfusion (I/R) injury via activation of the TRPV1 in wild-type (WT) but not in gene-targeted TRPV1-null mutant (TRPV1(-/-)) mice. Hearts of WT or TRPV1(-/-) mice were Langendorffly perfused with OLDA (2 x 10(-9) M) in the presence or absence of CGRP8-37 (1 x 10(-6) M), a selective calcitonin gene-related peptide (CGRP) receptor antagonist; RP-67580 (1 x 10(-6) M), a selective neurokinin-1 receptor antagonist; chelerythrine (5 x 10(-6) M), a selective protein kinase C (PKC) antagonist; or tetrabutylammonium (TBA, 5 x 10(-4) M), a nonselective K(+) channel antagonist, followed by 35 min of global ischemia and 40 min of reperfusion (I/R). Left ventricular end-diastolic pressure (LVEDP), left ventricular developed pressure (LVDP), coronary flow (CF), and left ventricular peak positive dP/dt (+dP/dt) were evaluated after I/R. OLDA improved recovery of cardiac function after I/R in WT but not TRPV1(-/-) hearts by increasing LVDP, CF, and +dP/dt and by decreasing LVEDP. CGRP8-37, RP-67580, chelerythrine, or TBA abolished the protective effect of OLDA in WT hearts. Radioimmunoassay showed that the release of substance P (SP) and CGRP after OLDA treatment was higher in WT than in TRPV1(-/-) hearts, which was blocked by chelerythrine or TBA. Thus OLDA exerts a cardiac protective effect during I/R injury in WT hearts via CGRP and SP release, which is abolished by PKC or K(+) channel antagonists. The protective effect of OLDA is void in TRPV1(-/-) hearts, supporting the notion that TRPV1 mediates OLDA-induced protection against cardiac I/R injury.     

Synthesis and biological evaluation of disubstituted N6-cyclopentyladenine analogues: the search for a neutral antagonist with high affinity for the adenosine A1 receptor

Novel 3,8- and 8,9-disubstituted N(6)-cyclopentyladenine derivatives were synthesised in moderate overall yield from 6-chloropurine. The derivatives were made in an attempt to find a new neutral antagonist with high affinity for adenosine A(1) receptors. N(6)-Cyclopentyl-9-methyladenine (N-0840) was used as a lead compound. Binding affinities of the new analogues were determined for human adenosine A(1) and A(3) receptors. Their intrinsic activity was assessed in [35S]GTPgammaS binding experiments. Elongation of the 9-methyl of N-0840 to a 9-propyl substituent was very well tolerated. A 9-benzyl group, on the other hand, caused a decrease in adenosine A(1) receptor affinity. Next, the 8-position was examined in detail, and affinity was increased with appropriate substitution. Most derivatives were A(1)-selective and 20 of the new compounds (6-9, 15-21, 23-26, 28, 31, 33, 35, and 36) had higher adenosine A(1) receptor affinity than the reference substance, N-0840. Compound 31 (N(6)-cyclopentyl-8-(N-methylisopropylamino)-9-methyladenine, LUF 5608) had the highest adenosine A(1) receptor affinity, 7.7 nM. In the [35S]GTPgammaS binding experiments, derivatives 5, 14, 22, 23, 25, 26, 33 and 34 did not significantly change basal [35S]GTPgammaS binding, thus behaving as neutral antagonists. Moreover, four of these compounds (23, 25, 26, and 33) displayed a 4- to 10-fold increased adenosine A(1) receptor affinity (75-206 nM) compared to N-0840 (852 nM). In summary, we synthesised a range of N-0840 analogues with higher affinity for adenosine A(1) receptors. In addition, four new derivatives, LUF 5666 (23), LUF 5668 (25), LUF 5669 (26) and LUF 5674 (33), behaved as neutral antagonists when tested in [35S]GTPgammaS binding studies. Thus, these compounds have improved characteristics as neutral adenosine A(1) receptor antagonists.     

Synthesis of novel N1-substituted bicyclic pyrazole amino acids and evaluation of their interaction with glutamate receptors

N1-substituted bicyclic pyrazole amino acids (S)-9a-9c and (R)-9a-9c, which are conformationally constrained analogues of glutamic acid, were prepared via a strategy based on a 1,3-dipolar cycloaddition. The new amino acids were tested for activity at ionotropic and metabotropic glutamate receptors. Some of them turned out to be selective for the NMDA receptors, where they behaved as weak antagonists. The biological activity is mainly due to the interaction with the glutamate binding site, and not with the glycine co-agonist site.     

Melanocortin-3 receptor activates MAP kinase via PI3 kinase

HEK 293 cells stably expressing human melanocortin-3 receptor (MC3R) were exposed to melanocortin receptor agonist, NDP-MSH (10(-)(10)-10(-)(6) M). ERK1/2 was phosphorylated in a dose-dependent manner with an EC(50) of 3.3+/-1.5 x 10(-)(9) M, similar to the IC(50) of NDP-MSH binding to the MC3R. ERK1/2 phosphorylation was blocked by the melanocortin receptor antagonists SHU9119. NDP-MSH-induced ERK1/2 phosphorylation was sensitive to pertussis toxin and the PI3K inhibitor, wortmannin. Rp-cAMPS, BAPTA-AM and Myr-PKC did not inhibit the NDP-MSH-induced ERK1/2 phosphorylation. NDP-MSH stimulated cellular proliferation in a dose-dependent manner with a similar EC(50) to ERK1/2 phosphorylation, 2.1+/-0.6 x 10(-)(9) M. Cellular proliferation was blocked by AGRP (86-132) and by the MEK inhibitor, PD98059. The NDP-MSH did not inhibit serum deprivation-induced apoptosis. MC3R activation induces ERK1/2 phosphorylation via PI3K and this pathway is involved in cellular proliferation in HEK cells expressing MC3R.     

Doxorubicin cardiomyopathy-induced inflammation and apoptosis are attenuated by gene deletion of the kinin B1 receptor

Clinical use of the anthracycline doxorubicin (DOX) is limited by its cardiotoxic effects, which are attributed to the induction of apoptosis. To elucidate the possible role of the kinin B1 receptor (B1R) during the development of DOX cardiomyopathy, we studied B1R knockout mice (B1R(-/-)) by investigating cardiac inflammation and apoptosis after induction of DOX-induced cardiomyopathy. DOX control mice showed cardiac dysfunction measured by pressure-volume loops in vivo. This was associated with a reduced activation state of AKT, as well as an increased bax/bcl2 ratio in Western blots, indicating cardiac apoptosis. Furthermore, mRNA levels of the proinflammatory cytokine interleukin 6 were increased in the cardiac tissue. In DOX B1R(-/-) mice, cardiac dysfunction was improved compared to DOX control mice, which was associated with normalization of the bax/bcl-2 ratio and interleukin 6, as well as AKT activation state. These findings suggest that B1R is detrimental in DOX cardiomyopathy in that it mediates the inflammatory response and apoptosis. These insights might have useful implications for future studies utilizing B1R antagonists for treatment of human DOX cardiomyopathy.     

Presynaptic CB(1) cannabinoid receptors control frontocortical serotonin and glutamate release--species differences

Both the serotonergic and endocannabinoid systems modulate frontocortical glutamate release; thus they are well positioned to participate in the pathogenesis of psychiatric disorders. With the help of fluorescent and confocal microscopy, we localized the CB(1) cannabinoid receptor (CB(1)R) in VGLUT1- and 2- (i.e. glutamatergic) and serotonin transporter- (i.e. serotonergic) -positive fibers and nerve terminals in the mouse and rat frontal cortex. CB(1)R activation by the synthetic agonists, WIN55212-2 (1 μM) and R-methanandamide (1 μM) inhibited the simultaneously measured evoked Ca(2+)-dependent release of [(14)C]glutamate and [(3)H]serotonin from frontocortical nerve terminals of Wistar rats, in a fashion sensitive to the CB(1)R antagonists, O-2050 (1 μM) and LY320135 (5 μM). CB(1)R agonists also inhibited the evoked release of [(14)C]glutamate in C57BL/6J mice in a reversible fashion upon washout. Interestingly, the evoked release of [(14)C]glutamate and [(3)H]serotonin was significantly greater in the CB(1)R knockout CD-1 mice. Furthermore, CB(1)R binding experiments revealed similar frontocortical CB(1)R density in the rat and the CD-1 mouse. Still, the evoked release of [(3)H]serotonin was modulated by neither CB(1)R agonists nor antagonists in wild-type CD-1 or C57BL/6J mice. Altogether, this is the first study to demonstrate functional presynaptic CB(1)Rs in frontocortical glutamatergic and serotonergic terminals, revealing species differences.     

Receptor-antagonist interactions in the complexes of agouti and agouti-related protein with human melanocortin 1 and 4 receptors

The molecular interactions between human melanocortin receptor-1 and -4 (hMC1R and hMC4R) and their endogenous antagonists, agouti signaling protein (ASIP) and agouti-related protein (AGRP), were assessed by studying the effects of site-directed mutations on the binding affinity of (125)I-ASIP[90-132(L89Y)] and (125)I-AGRP(86-132). Mutations of homologous residues from transmembrane helices (TMHs) 3 and 6 and extracellular loop (EL) 3 (D121A, T124A, F257A, and F277M in hMC1R and D126A, I129A F261A, and M281F in hMC4R) impaired binding of both antagonists to hMC4R and binding of the ASIP fragment to hMC1R. However, the mutations in TMH2 (E94A in hMC1R and E100A in hMC4R), TMH7 (F280A in hMC1R and F284A in hMC4R), and EL2 (Y183S, H184S, and D184H in hMC1R) only significantly affected binding of the ASIP fragment. The dependence of agonist binding on the dithiothreitol concentration followed a monophasic curve for wild-type hMC4R and its C40A, C271A, and C279A mutants and a biphasic curve for hMC1R, suggesting the presence of at least one structurally and functionally essential disulfide bond in both wild-type receptors and the hMC4R mutants. Models of complexes of both receptors with the ASIP fragment and hMC4R with the AGRP fragment were calculated using constraints from the experimental structures of rhodopsin and AGRP fragments, a set of deduced hydrogen bonds, supplemented by two proposed disulfide bridges and receptor-ligand contacts, derived from our mutagenesis data. In the models of the ASIP fragment complexed with both receptors, the core ligand tripeptide, Arg-Phe-Phe, positioned between TMHs 3 and 6, is shifted toward TMHs 2 and 7 relative to its position in the AGRP-hMC4R model, while the N-terminal loop and two central disulfides of the antagonists interact with EL2 of the receptors.     

Discovery of novel phenethylpyridone derivatives as potent melanin-concentrating hormone 1 receptor antagonists

Novel phenethylpyridone derivatives were identified as potent human melanin-concentrating hormone 1 receptor (MCH-1R) antagonists. A search for surrogates for the 4-(2-aminoethoxy)phenyl moiety of 1 resulted in discovery of 2-[4-(aminomethyl)phenyl]ethyl substructure as in 6a. Successive optimization of the right-hand moiety led to the identification of a number of potent derivatives.     

Identification of a key region of kinin B(1) receptor for high affinity binding of peptide antagonists

To investigate the molecular basis for the specificity of ligand recognition in human kinin B(1) (B(1)R) and B(2) (B(2)R) receptors, we constructed a series of chimeric receptors by progressively replacing, from the N to the C terminus, the human B(2)R domains by their B(1) counterparts. The chimeric construct possessing the C-terminal tail and the transmembrane domain VII (TM VII) of the B(2)R (construct 6) displayed 7- and 20- fold decreased affinities for the B(1) agonist [(3)H]desArg(10)-kallidin (desArg(10)-KD) and the B(1) antagonist [(3)H]desArg(10)-[Leu(9)]-KD respectively, as compared with the wild-type B(1)R. Moreover, the substitution of the B(1) TM VII by its B(2) homologue TM increased the affinity for the pseudopeptide antagonists, Hoe140 and NPC 567. High affinity for desArg(10)-KD binding was fully regained when the B(2) residue Thr(287) was replaced in construct 6 by the corresponding B(1) Leu(294) residue. When the B(2) residue Tyr(295) was exchanged with the corresponding B(1) Phe(302), high affinity binding for both agonist and antagonist was recovered. Moreover, the L294T and F302Y mutant B(1)R exhibited 69- and 6.5-fold increases, respectively, in their affinities for the B(2) receptor antagonist, Hoe140. Therefore we proposed that Leu(294) and Phe(302) residues, which may not be directly involved in the binding of B(1)R ligands and, hence, their Thr(287) and Tyr(295) B(2) counterparts, are localized in a receptor region, which plays a pivotal role in the binding selectivity of the peptide or pseudopeptide kinin ligands.     

Characterization of a Human NK1 Tachykinin Receptor in the Astrocytoma Cell Line U 373 MG

Abstract: The human NK₁ tachykinin receptor in the astrocytoma cell line U 373 MG was characterized using selective agonists and antagonists described for this receptor in the rat. Specific [³H]substance P binding sites were present on cell homogenates, whereas [³H]neurokinin A or [³H]-senktide binding sites were absent. The binding was saturable and reversible. The binding of [³H]substance P was inhibited by very low concentrations of [L-Pro⁹]substance P and [Sar⁹,Met(O₂)¹¹]substance P; septide was ∼ 1,000-fold less potent. The most potent peptide antagonist was trans -4-hydroxy-1-(1 H -indol-3-ylcarbonyl)-L-prolyl- N -methyl- N -(phenylmethyl)-L-tyrosineamide. The rank order of potency for the nonpeptide antagonists was ( S , S )-CP 96,345 > (±)-CP 96,345 > (±)-2-chlorobenzylquinuclidinone > ( R , R )-CP 96,345 > RP 67580 > RP 68651. In [³H]-inositol-labeled cells, substance P stimulated phosphatidylinositol turnover. A good correlation was found when the abilities of NK₁ receptor agonists for stimulating inositol phosphate production and for inhibiting [³H]substance P binding were compared. Similarly, the binding and functional assays were well correlated for the antagonists. As a result of its high sensitivity and selectivity, the U 373 MG cell line thus appears an excellent tool for investigating the pharmacology of the human NK₁ receptor.     

Cyclic analogs of alpha-melanocyte-stimulating hormone (alphaMSH) with high agonist potency and selectivity at human melanocortin receptor 1b

Alpha-melanotropin (alphaMSH), Ac-Ser1-Tyr2-Ser3-Met4-Glu5-His6-Phe7-Arg8-Trp9-Gly10-Lys11-Pro12-Val13-NH2,(1) has been long recognized as an important physiological regulator of skin and hair pigmentation in mammals. Binding of this peptide to the melanocortin receptor 1 (MC1R) leads to activation of tyrosinase, the key enzyme of the melanin biosynthesis pathway. In this study, interactions of the human MC1bR (an isoform of the receptor 1a) with the synthetic cyclic analogs of alphaMSH were studied. These ligands were analogs of MTII, Ac-Nle4-cyclo-(Asp5-His6-D-Phe7-Arg8-Trp9-Lys10)-NH2, a potent pan-agonist at the human melanocortin receptors (hMC1,3-5R). In the structure of MTII, the His6-D-Phe7-Arg8-Trp9 segment has been recognized as "essential" for molecular recognition at the human melanocortin receptors (hMC1,3-5R). Herein, the role of the Trp9 in the ligand interactions with the hMC1b,3-5R has been reevaluated. Analogs with various amino acids in place of Trp9 were synthesized and tested in vitro in receptor affinity binding and cAMP functional assays at human melanocortin receptors 1b, 3, 4 and 5 (hMC1b,3-5R). Several of the new peptides were high potency agonists (partial) at hMC1bR (EC50 from 0.5 to 20 nM) and largely inactive at hMC3-5R. The bulky aromatic side chain in position 9, such as that in Trp, was found not to be essential to agonism (partial) of the studied peptides at hMC1bR.     

Optimization of CRF1R binding affinity of 2-(2,4,6-trichlorophenyl)-4-trifluoromethyl-5-aminomethylthiazoles through rapid and selective parallel synthesis

An efficient approach was developed to synthesize 2-(2,4,6-trichlorophenylamino)-4-trifluoromethyl-5-aminomethylthiazoles, corticotropin-releasing factor type 1 receptor (CRF(1)R) antagonists, by monoalkylation of amines with chloromethyl intermediate 5. The effect of variations in aminomethyl side chain of 6 on binding affinity is discussed.     

The substance P/neurokinin-1 receptor system in lung cancer: Focus on the antitumor action of neurokinin-1 receptor antagonists

The last decades have seen no significant progress in extending the survival of lung cancer patients and there is an urgent need to improve current therapies. The substance P (SP)/neurokinin-1 receptor (NK-1R) system plays an important role in the development of cancer: SP and NK-1R antagonists respectively induce cell proliferation and inhibition in human cancer cell lines. No study of the involvement of this system in non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) cells has been carried out in depth. Here, we demonstrate the involvement of the SP/NK-1R system in human H-69 (SCLC) and COR-L23 (NSCLC) cell lines: (1) they express isoforms of the NK-1R and mRNA for the NK-1R; (2) they overexpress the tachykinin 1 gene; (3) the NK-1R is involved in their viability; (4) SP induces their proliferation; (5) NK-1R antagonists (Aprepitant (Emend), L-733,060, L-732,138) inhibit the growth of both cell lines in a concentration-dependent manner; (6) the specific antitumor action of these antagonists against such cells occurs through the NK-1R; and (7) lung cancer cell death is due to apoptosis. We also demonstrate the presence of NK-1Rs and SP in all the human SCLC and NSCLC samples studied. Our findings indicate that the NK-1R may be a promising new target in the treatment of lung cancer and that NK-1R antagonists could be new candidate antitumor drugs in the treatment of SCLC and NSCLC.     

Total synthesis of the mollu-series glycosyl ceramides alpha-D-Manp-(1----3)-beta-D-Manp-(1----4)-beta-D-Glcp-(1----1)-Cer and alpha-D-Manp-(1----3)-[beta-D-Xylp-(1----2)]-beta-D-Manp-(1----4)-beta- D-Glcp-(1----1)-Cer

The mollu-series glycosphingolipids, O-alpha-D-mannopyranosyl-(1----3)-O-beta-D-mannopyranosyl-(1----4)-O-bet a-D-glucopyranosyl-(1----1)-2-N-tetracosanoyl-(4E)-sphingeni ne and O-alpha-D-mannopyranosyl-(1----3)-O-[beta-D-xylopyranosyl-(1----2])-O- beta-D-mannopyranosyl-(1----4)-O-beta-D-glucopyranosyl-(1----1)-2-N- tetracosanoyl-(4E)-sphingenine, were synthesized for the first time by using 2,3,4-tri-O-acetyl-D-xylopyranosyl trichloroacetimidate, methyl 2,3,4,6-tetra-O-acetyl-1-thio-alpha-D-mannopyranoside, benzyl O-(4,6-di-O-benzyl-beta-D-mannopyranosyl)-(1----4)-2,3,6-tri-O-benzyl-be ta-D- glucopyranoside 9, and (2S,3R,4E)-2-azido-3-O-(tert-butyldiphenylsilyl)-4-octade cene-1,3-diol 6 as the key intermediates. The hexa-O-benzyl disaccharide 9 was prepared by coupling two monosaccharide synthons, namely, 2,3-di-O-allyl-4,6-di-O-benzyl-alpha-D-mannopyranosyl bromide and benzyl 2,3,6-tri-O-benzyl-beta-D-glucopyranoside. It was demonstrated that azide 6 was highly efficient as a synthon for the ceramide part in the coupling with both glycotriaosyl and glycotetraosyl donors, particularly in the presence of trimethylsilyl triflate.