A new general solid-phase method for the synthesis of backbone-to-backbone cyclized peptides

A model peptide with the sequences Ala-Pro-Lys(2ClZ)-Tyr(2BrZ) was synthesized on a 4-methylbenzhydryl amine (MBHA) polystyrene resin using conventional Boc/benzyl protective group strategy. The amino acid aldehyde Boc-valinal was coupled by reductive alkylation with NaCNBH₃ in acidified DMF for 1 h. The secondary amine in the peptide-resin Boc-Valψ[CH₂NH]Ala-Pro-Lys(2CIZ)-Tyr(2BrZ)-MBHA was reductively alkylated by 3(4-methylbenzylthio)-propanal at 40 °C for 6 h, resulting the peptide-resin Boc-Valψ[CH₂N(CH₂CH²CH₂-S-pMeBzl)]Ala-Pro-Lys(2ClZ)-Tyr(2BrZ)-MBHA. After the removal of the Boc group the synthesis was continued employing the above-mentioned methods, which led to the resin-bound peptide Leuψ[CH₂N(CH₂CH₂CH₂S-pMeBzl)]Ser-Pro-Gly-Lys(2ClZ)-Valψ[CH₂N(CH₂CH₂CH₂-S-pMeBzl)]Ala-Pro-Lys(2ClZ)-Tyr(2BrZ)-MBHA. The peptide was cleaved from the resin with hydrogen fluoride. Reversed-phase HPLC and plasma desorbtion mass spectrometry analysis showed that the expected peptide Leuψ[CHIN(CH₂CH₂CH₂SH)ISer-Pro-Gly-Lys-Valψ[CH₂N(CH₂CH₂CH₂-SH)]Ala-Pro-Lys-Tyr-NH₂ was obtained as the major product with low levels of side products. Intramolecular oxidation of the thiols gave the backbone to backbone cyclized peptide Leuψ[CH₂N(CH₂CH₂CH₂S)]Ser-Pro-Gly-Lys-Valψ[CH₂N(CH₂CH₂CH₂-S)]A1a-Pro-Lys-Tyr-NH₂.     

Thyrotropin-Releasing Hormone (TRH) Uptake in Intestinal Brush-Border Membrane Vesicles: Comparison with Proton-Coupled Dipeptide and Na+-Coupled Glucose Transport

The mechanism of thyrotropin-releasing hormone (pGlu-His-Pro-NH₂; TRH) uptake across the luminal membrane of intestinal enterocytes was investigated using brush-border membrane vesicles (BBMV) from rabbit duodenum and jejunum and rat upper small intestine. [¹⁴C]Glucose accumulated within the intestinal vesicles (at 10 sec), in the presence of an inwardly directed Na⁺ gradient, 7- to 14-fold higher than equilibrium values (65 min). The vesicles also accumulated the dipeptide [¹⁴C]Gly-Sar. Dipeptide uptake was greatest in the presence of both an inwardly directed proton gradient and an inside negative membrane potential. The H⁺-dependent Gly-Sar transport was not affected by the presence of an excess (46-fold) of cold TRH. In contrast to the observations with glucose and Gly-Sar, the uptake of [³H]TRH after 10 or 60 sec (in each of the vesicle preparations) was not enhanced by either Na⁺ or H⁺ gradient conditions. The absence of vesicular accumulation of TRH was not due to peptide hydrolysis. For example, after a 60-sec incubation with rabbit jejunal BBMV no degradation of the tripeptide was evident. After 65 min, 6% of [³H]TRH had undergone degradation, by deamidation, to form TRH-OH. These studies provide no evidence for the oral absorption of TRH by a Na⁺- or H⁺-dependent carrier system in the brush-border membrane. Previous observations of TRH absorption in vivo may be accounted for by passive absorption of the peptide combined with its relative resistance to luminal hydrolysis.     

Potent pseudopeptide bombesin-like agonists and antagonists

Bombesin-like pseudopeptides have been synthesized, and certain physicochemical properties and biological activities have been examined. Bombesin and the related peptide litorin were modified at positions 13–14 and 8–9, respectively, with ψ[CH₂S] and ψ[CH₂N(CH₃)]. [Phe¹³ψ[CH₂S]Leu¹⁴]bombesin and [Phe⁸ψ[CH₂S]-Leu⁹]litorin bound to the murine pancreatic bombesin gastrin releasing peptide receptor with similar dissociation constants (K_d= 3.9 and 3.4 nM. respectively). Increased potency was achieved by oxidation of the thiomethylene ether to two diastereomeric sulfoxides (isomer I, K_d= 1.6 nM and isomer II, K_d= 0.89nM. Further oxidation to the sulfone decreased potency ([Phe⁸ψ[CH₂SO₂]Leu⁹]litoin, K_d= 9.9nM). All five analogs were receptor antagonists as determined by phosphatidylinositol turnover in murine pancreas. In contrast to these peptide backbone substitutions, a ψCH₂N(CH₃)] at the 8–9 amide bond position resulted in an agonist. The analogs were compared with those of litorin (K_d= 0.1 nM) and [Leu⁹]litorin (K_d= 0.17 nM) by CD and fluorescence spectroscopy. The CD spectra demonstrated ordered conformation for all the peptides in TFE. Different conformations corresponding to agonist and antagonist peptides were suggested by CD. Based on the pH-dependence of the fluorescence spectra of the peptides in a zwitterionic detergent, two titratable groups were identified (pK_a= 6.3 and 8.5). The lower pK_a is found in the agonist analogs but not in the ψ[CH₂S]-containing antagonist.     

Synthesis of diketopiperazines with on‐resin N‐methylation and cyclative release

Abstract: Enantiomerically pure N‐methylated diketopiperazines (DKP) can be obtained by treating a N‐methylated resin‐bound dipeptide with 20% piperidine in dimethylformamide via a process known as cyclative release. N‐methylated resin‐bound dipeptides can be formed from N‐methylated precursors or N‐methylation can be selectively performed on the resin. When on‐resin N‐methylation was performed on the C‐terminal side of the dipeptide, diastereomers were formed. Yet the cyclative release is shown to be a stereoselective process, as seen using preformed N‐methylated amino acids. The procedure was also applied to synthesize the pseudodiketopiperazine cyclo(Pheψ[CH₂NH]Leu). When comparing nonmethylated, monomethylated and bismethylated derivatives, we find that N‐methylation results in a dramatic increase in solubility.     

Effects of thioamide substitution for the enkephalin conformation

The crystals of Boc-Tyr-Gly-Gly-PheΨ[CSNH]Leu-OBzl monohydrate (C₄₀H₅₁N₅O₈S H₂O), a monothionated Leu-enkephalin analogue, were obtained with space group P2₁ a = 12.616(3), b= 9.347(2), c= 18.548(5) Å, β= 96.31(4)°. The structure was elucidated by X-ray diffraction analysis, and refined to the R value of 0.091 for the observed 3294 reflections. Two antiparallel molecules related by a pseudo twofold symmetry were stabilized to each other by four intermolecular hydrogen bonds. The molecular conformation was bent at the Phe residue, and the extended moiety of the Tyr-Gly-Gly fragment was almost perpendicular to that of the Phe-Leu residues. Consequently the molecule, as a whole, formed an L-shape conformation with a slightly left-handed helicity.     

Synthesis of Potentially Antineoplastic Derivatives of N-[N-(2-Chloroethyl)-N-nitrosocarbamoyl]amino Acids

The synthesis of N-[N-(2-chloroethyl)-N-nitrosocarbamoyl]amino acids and their anilides, congeners of N-(2-chloroethyl)-N-nitrosoureas, as potential antineoplastic substances is reported. N-[N-(2-chloroethyl)-N-nitrosocarbamoyl]amino acids are prepared by reaction of amino acids with N-(2-chloroethyl)-N-nitrosocarbamoyl azide. Corresponding anilides and toluidides are obtained by condensation of the primary reaction products with aniline and toluidine using dicyclohexylcarbodiimide (DCC).     

Importance of the leucine side-chain to the spasmogenic activity and binding of Substance P analogues

Previous studies with Substance P (SP) antagonists (GR 71251, [d Pro⁹, Pro¹⁰, Trp¹¹]SP and d Pro⁹, MeLeu¹⁰, Trp¹¹]SP) have suggested the existence in the guinea-pig ileum (GPI) of two distinct tachykinin receptors associated with the contractile responses of [Pro⁹]SP and septide. In addition [Apa^9-10]SP, a glycine-substituted analogue of SP with a carba bond between residues 9 and 10, [Gly⁹-ψ(CH₂-CH₂)-Gly¹⁰SP = [Apa^9-10]SP, was shown to belong to the ‘septide family’ (low affinity for NK-1 specific binding sites and high potency in the GPI). In order to establish the importance of the isopropyl side-chain in position 10, the binding potencies and activities of [Gly⁹-ψ(CH₂-CH₂)-Gly¹⁰]SP, [Ala¹⁰]SP, [Gly⁹-ψ(CH₂-CH₂)-Leu¹⁰]SP and [Gly⁹-ψ(CH₂-CH₂)-d Leu¹⁰]SP were compared. Conformational behaviour of active peptides with a carba bond was analyzed by NMR and modelisation studies. This study with agonists demonstrated that undecapeptides substituted in position 10 in the SP sequence also enabled discrimination of NK-1 receptors from receptors responsible for the spasmogenic activities of peptides belonging to the ‘septide family’. [Gly⁹-ψ(CH₂-CH₂)- Leu¹⁰]SP is ahighly potent NK-1 agonist, [Gly⁹-ψ(CH₂-CH₂)-Gly¹⁰]SP acts on the septide-sensitive receptor, and [Ala¹⁰]SP is a mixed agonist.     

The Cardiovascular Effects of Thyrotropin-Releasing Hormone (TRH) are Attenuated by Cimetidine in Rats

Abstract: The modulation of cardioventilator effects of thyrotropin-releasing hormone (TRH) by histaminergic mechanisms was studied in anaesthetized rats pretreated with histamine receptor antagonists. TRH (1–10. nmol/kg) into the lateral cerebral ventricle dose-dependently elevated mean arterial pressure, heart rate and stimulated respiration. The respiratory stimulating effect of TRH remained unchanged after pretreatments with histamine H₁-receptor antagonist diphenhydramine or H₂-receptor antagonists cimetidine and ranitidine, while the TRH-induced hypertension and tachycardia were attenuated by cimetidine. This antagonism was not due to an interation between TRH and cimetidine at their central binding sites, since there was no displacement of [³H]MeTRH binding in the presence of cimetidine nor did TRH displace [³H]cimetidine in rat brain homogenates. Inability of diphenhydramine to modify the cardiovascular effects of TRH indicates that these effects are not due to histamine liberation, as cardiovascular stimulation after central administration of histamine is mainly mediated via H₁-receptors. The antagonism of the cardiovascular responses to TRH by cimetidine was not due to blockade of H₂-receptors, since another potent H₂-receptor antagonist ranitidine was unable to affect the cardiovascular effects of TRH. Therefore, we suggest that cimetidine exerted antagonism of TRH by some non-specific action.     

Crystal-state structural analysis of two γ-lactam-restricted analogs of Pro-Leu-Gly-NH2

The crystal structures of two analogs of Pro-Leu-Gly-NH, (1), containing a γ-lactam conformational constraint in place of the -Leu-Gly- sequences, are described. The highly biologically active (S,R)-diastere-omer 2a is semi-extended at the C-terminus, with the N-terminal Pro residue in the unusual “C₅” conformation [ψ₁=– 0.8(15)°] stabilized by a (peptide)N-H…N(amino) intramolecular H-bond [the N(3)…N(4) separation is 2.687(11)Å]. Conversely, the N,N′-isopropylidene aminal trihydrate of the (S,S)-diastereomer 2b, compound 3, adopts a β-bend conformation at the C-terminus, as already reported for 1. However, the backbone torsion angles [φ= 57.4(4), ψ₂=– 129.9(3)°; ψ₃= - 92.3(4), ψ₃= 6.4(5)°] lie close to the values expected for the corner residues of an ideal type-II β-bend. A weak intramolecular 4 → 1 H-bond is seen between the Gly carboxyamide anti-NH and Pro C=O groups. In the newly formed 2,2,3,4-tetraalkyl-5-oxo-imidazolidin-1-yl moiety the ψ₁ torsion angle is 12.9(4)° and the intramolecular N(3)…N(4) separation is 2.321(4)Å.     

Synthesis of 14C‐ and 14C3‐labelled Org 37462

Org 37462 (1) is the active ingredient in Orgalutran^®, an innovative product that reduces the time of treatment in in vitro fertilization from four to less than two weeks. Org 37462 is a synthetic decapeptide containing several amino acids that are unnatural in stereochemistry and/or in structure. The synthesis, starting with a ProtectingGroup‐D‐Ala‐resin, is a typical solid state synthesis. For the conduction of several metabolism studies, Org 37462 had to be labelled with carbon 14. It was decided to label D‐3‐(2‐naphthyl)alanine, the last amino acid to be coupled to the resin. We report the synthesis of [¹⁴C]‐ and [¹⁴C₃]‐Org 37462, starting from 2‐bromo‐[¹⁴C‐methyl]‐naphthalene and [¹⁴C₂]‐tert‐butyl glycinate. Copyright © 2008 John Wiley & Sons, Ltd.     

肿瘤化学治疗的研究 Ⅴ.某些具有双(2-氯乙基)氨基的苯丙氨酸的合成

本文报导了溶肉瘤素的三种异构体,卽β-[间双(2-氯乙基)氢基苯基]-α-氨基丙酸(Ⅰ_b)、β-[对双(2-氯乙基)氨基苯基]-β-氨基丙酸(Ⅱ_α)及β-[间双(2-氯乙基)氨基苯基]-β-氨基丙酸(Ⅱ_b)的合成和它们的抗肿瘤作用,并对它们的化学结构与生理作用的关系进行了讨论。上述化合物对多种动物实验肿瘤的生长皆具显著的、不同程度的抑制作用。化合物Ⅱ_a(编号合14)已在临床研究中。     

Substitution of aromatic and nonaromatic amino acids for the Phe3 residue in the δ-selective opioid peptide deltorphin I: Effects on binding affinity and selectivity

Deltorphins I and II (Tyr-D-Ala-Phe-Asp-Val-Val-Gly NH₂ and Tyr-D-Ala-Phe-Glu-Val-Val-Gly NH₂) display a high degree of 6-opioid receptor selectivity. Since they lack the intervening Gly3 residue found between the Tyr and Phe aromatic moieties in pentapeptide enkephalins, deltorphins I and II resemble a previously described series of cyclic tetrapep-tides based on Tyr-c[D-Cys-Phe-D-Pen] (JOM-13). With the goal of development of structure-activity relationships for deltorphins and comparison with that of the cyclic tetrapep-tides, ten analogs of deltorphin I were synthesized in which Phe³ was replaced with specific aromatic and nonaromatic amino acids with varying physicochemical properties. Results indicated that analogs containing the bicyclic aromatic amino acids 3-(l-naphthyl)-L-alanine [1-Nal; K_i(μ) = 767 nM, K_i(§) = 7.70 nM], 3-(2-naphthyl)-L-alanine [2-Nal; K_i(μ)= 1910 nM, K_i(§) = 49.2 nM], tryptophan [K_i(μ)= 1250 nM, K_i(§) = 23.9nM], and 3-(3-benzothienyl)-L-alanine [Bth; K_i(μ)= 112nM, K_i(§) = 3.36 nM] were fairly well tolerated at μ- and §-receptors, though affinity was compromised to varying degrees relative to deltorphin I. Shortening the Phe side chain by incorporation of phenylglycine (Pgl) was detrimental to both μ (K_i= 4710 nM) and § (K_i= 15.6 nM) binding, while extension of the side chain with homophenylalanine (Hfe) enhanced μ binding (K_i= 67.8 nM), leaving § affinity unaffected (K_i= 2.64 nM). Substitution with nonaromatic amino acids valine and isoleucine led expectedly to poor opioid binding [K_i(μ) =≥ 10000 nM for each, K_i(§) = 160 and 94.7 nM, respectively], while peptides containing cyclohexylalanine (Cha) and leucine surprisingly retained affinity at both μ (K_i= 322 and 1240 nM, respectively) and § (K_i= 10.5 and 12.4 nM, respectively) sites. In general, these trends mirror those observed for similar modification in Tyr-c[D-Cys-Phe-D-Pen].     

From peptide libraries to optimized nonpeptide ligands in the search for S‐farnesyltransferase inhibitors

Abstract: A complete 331 776‐member library of tetrapeptides made of 24 amino acid building blocks was synthesized robotically on solid phase and subjected to a deconvolution based on the inhibitory potency of the sublibraries in a HPLC assay of the S‐farnesyltransferase activity in vitro. One of the non‐natural peptide and noncysteine‐containing leads Nip‐Trp‐Phe‐His (Nip = p‐nitrophenyl‐l ‐alanine) was optimized chemically to give a proteolytically stable pseudopeptide with a 200‐fold potency compared with the original lead. The final compound was converted to the C‐terminal ethyl ester: p‐F‐C₆H₄‐CO(CH₂)₂‐CO‐Bta‐d ‐Pheψ[CH2NH]His‐OEt (Bta = benzothienyl‐l ‐alanine) and shown to behave as a prodrug which was hydrolyzed back to the C‐terminal acid following cell penetration. The method confirmed that several structurally original leads can be discovered in large libraries when deconvolution relies upon a highly specific assay and that these leads can be optimized by chemical modification to impart the final compound the desired pharmacological and pharmacokinetic properties.     

Synthesis and biological activities of ψ(CH2NH) pseudopeptide analogues of the C-terminal hexapeptide of neurotensin

Each peptide CO-NH function in the biologically important C-terminal 8-13 sequence of neurotensin was replaced by the reduced CH₂-NH isostere using the rapid in situ solid phase procedure developed by Sasaki & Coy. In general this modification resulted in a drop in receptor affinity except for the [Argψ(CH₂NH)Arg]-NT_8–13 analogue (PIC₅₀ 9.23 vs. NT_8–13 PIC₅₀ 8.03). This analogue also showed enhanced enzymatic stability, but acted as a full agonist as shown by the observation of relaxations of guinea-pig colon ascendens.     

Immunostimulants and Toll‐like receptor ligands obtained by screening combinatorial lipopeptide collections*

Abstract: Synthetic lipopeptides carrying the head group of bacterial lipoproteins are specific ligands of Toll‐like receptors (TLR). The three fatty acids containing lipopeptides with the tripalmitoyl‐S‐glyceryl‐cysteinyl N‐terminus (Pam₃Cys) are agonists of TLR2. The structurally related lipopeptides with a head group lacking the fatty acyl residue at the amino‐terminus (Pam₂Cys) stimulate TLR2 and 6. To investigate the influence of the peptide chain of lipohexapeptides with a free N‐terminus with regard to their ability to enhance B‐cell proliferation, a randomized S‐[2,3‐bis(palmitoyloxy)‐(2RS)‐propyl]‐(R)‐cysteinyl‐pentapeptide amide collection Pam₂CysXXXXX and 5 × 19 subcollections (Pam₂CysOXXXX, Pam₂CysXOXXX, Pam₂CysXXOXX, Pam₂CysXXXOX, Pam₂CysXXXXO, O: all protein amino acids except Cys) were prepared by parallel solid‐phase synthesis. The collection represents synthetic lipopeptide analogues of the numerous bacterial lipoproteins and of mycoplasma lipoprotein. Each of the 95 subcollections is characterized by one defined and four degenerated amino acid positions thus comprising 19⁴ individual lipopeptides with free N‐terminal amino groups. High‐performance liquid chromatography electrospray mass spectrometry (HPLC–ESI‐MS) was applied for the analytical characterization of the lipohexapeptide amide subcollections and for the individual lipohexapeptide amides. The subcollections were tested for polyclonal activation of murine spleen cells, deconvolution led to highly active single S‐[2,3‐bis(palmitoyloxy)‐(2RS)‐propyl]‐(R)‐cysteinyl‐pentapeptide amides.     

Synthesis of three isotopically labeled versions and a metabolite of Aurora A kinase inhibitor

Sodium ring‐[¹⁴C]‐4‐[[9‐chloro‐7‐(2,6‐difluorophenyl)‐5H‐pyrimido[5,4‐d][2]benzazepin‐2‐yl]amino]‐benzoate (1A, MLN8054), an Aurora A kinase inhibitor, was synthesized from [¹⁴C]‐cyanamide in two steps in an overall radiochemical yield of 7%. The intermediate, [¹⁴C]‐4‐guanidinobenzoic acid, was prepared by coupling [¹⁴C]‐cyanamide with 4‐aminobenzoic acid. Sodium carboxyl‐[¹⁴C]‐4‐[[9‐chloro‐7‐(2,6‐difluorophenyl)‐5H‐pyrimido[5,4‐d][2]benzazepin‐2‐yl]amino]‐benzoate (1B) was synthesized from carboxyl‐[¹⁴C]‐4‐guanidinobenzoic acid in one step in a radiochemical yield of 35%. [D₄,¹⁵N]‐4‐[[9‐chloro‐7‐(2,6‐difluorophenyl)‐5H‐pyrimido[5,4‐d][2]benzazepin‐2‐yl]amino]‐benzoic acid (1C) was synthesized from [¹⁵N₂]‐cyanamide and [D₄]‐4‐aminobenzoic acid in two steps in an overall yield of 37%. The major metabolite, β‐acyl glucuronide of 4‐[[9‐chloro‐7‐(2,6‐difluorophenyl)‐5H‐pyrimido[5,4‐d][2]benzazepin‐2‐yl]amino]‐benzoic acid (14), was synthesized from D‐glucuronic acid in three steps in an overall yield of 1%. The key intermediate for synthesis of glucuronide was prepared by HATU catalyzed coupling of 4‐[[9‐chloro‐7‐(2,6‐difluorophenyl)‐5H‐pyrimido[5,4‐d][2]benzazepin‐2‐yl]amino]‐benzoic acid with allyl glucuronate. Copyright © 2009 John Wiley & Sons, Ltd.     

Synthesis of [3H]‐labelled 4‐[Ethyl[2,5,6‐trimethyl‐7‐(2,4,6‐trimethylphenyl)pyrrolo[2,3‐d]pyrimidin‐4‐yl]amino]‐2,3‐[3H]‐butan‐1‐ol: a high affinity radioligand for the corticotropin‐releasing hormone type 1 receptor

[³H]‐Labelled 4‐[ethyl[2,5,6‐trimethyl‐7‐(2,4,6‐trimethylphenyl)pyrrolo[2,3‐d]pyrimidin‐4‐yl]amino]‐2,3‐[³H]‐butan‐1‐ol ( 3b ) was prepared as a novel non‐peptidic radiolabelled high affinity antagonist of the corticotropin‐releasing hormone type 1 receptor (CRHR₁) that could be useful as a more stable and receptor‐selective alternative to the radiolabelled peptides now used to label the CRHR₁ receptor for displacement studies in cell‐based binding assays. The precursor (Z)‐4‐[ethyl[2,5,6‐trimethyl‐7‐(2,4,6‐trimethylphenyl)pyrrolo[2,3‐d]pyrimidin‐4‐yl]amino]but‐2‐en‐1‐ol ( 2 ) was reduced with tritium gas using palladium as the catalyst. After HPLC purification 3b was obtained with a specific activity of 35 Ci/mmol in high radiochemical purity (>97%). Copyright © 2006 John Wiley & Sons, Ltd.     

Conformational preferences of oligopeptides rich in α-aminoisobutyric acid. III. Design,synthesis and hydrogen bonding in 310-helices

Two sterically constrained peptides {ⁱBoc-Aib-Aib-Aib-DkNap-Leu-Qx-Ala-Aib-Aib-F1, (Dk⁴Qx⁶[7/9]) and ⁱBoc-Aib-Aib-Aib-DkNap-Leu-Aib-Ala-Aib-Aib-Fl, (Dk⁴7/9)} containing α-aminoisobutyric acid (Aib) and Aib-class amino acids in conjunction with selected mono-α-alkyl amino acids were synthesized by an optimized TBTU/HOBt procedure. The use of Aib-class amino acids (e.g. DkNap and Qx), defined and discussed here, gives rise to the same overwhelmingly 3₁₀-helical backbone conformation as that provided by simpler Aib-rich peptides and homopeptides. The synthetic α,α-dialkylamino acids (DkNap, Qx) are aromatic homologues of the known alicyclic variants of Aib, the Ac₅c and Ac₆c amino acids. Two new organic solubilizing groups for peptides, ⁱBoc and 2-methoxyethylamine, are introduced. The ¹H nuclear magnetic resonance analyses of the Dk⁴s/p[7/9] and Dk⁴Qx⁶[7/9] peptides demonstrate the unambiguous 310s/b-helical hydrogen bonding pattern of these peptides, confirming the design objective of these sequence patterns containing greater than 50% Aib and Aib-class composition. © Munksgaard 1994.     

LABELING OF AMINO ACIDS AND PEPTIDES WITH ISOTOPIC OXYGEN AS FOLLOWED BY 17O-N.M.R.*

¹⁷O was introduced into the respective α- and γ-COOH groups of Boc-Gly and Boc-Glu by saponification of the corresponding O-methyl esters with 1 N NaOH in H₂¹⁷O. Other ¹⁷O enriched Boc-amino acids were prepared by acid catalyzed exchange into the amino acid α-COOH group followed by t-butyloxycarbonylation with t-butyl S-4, 6-dimethylpyrimidin-2-ylthio carbonate. Final enrichment, by approximately three orders of magnitude over natural abundance, was 60–100% of the possible maximum. The synthesis of [¹⁷O]-Gly-Ala, [¹⁷O]-Gly-Leu and [¹⁷O]-Gly-Glu by DCC/HBT mediated coupling of Boc-Gly-[¹⁷'O]-α-COOH with amino acid-O-t-butyl esters followed by deprotection with HCl/EtOAc proceeded without undue loss of the isotope. Boc-[¹⁷O]-Pro-Leu-Gly-NH₂ was prepared by a similar procedure. [Tyr^2–17O]-, [Pro^7–17O]- and [Gly^4–17O]-oxytocin were synthesized using solid phase support. ¹⁷O-chemical shifts of synthetic intermediates and of the final products were as expected for each functional group. Linewidth data correlate with the molecular weights of the compounds prepared.     

Syntheses of Potentially Antineoplastic Amides and Esters of N-[N'-(2-Chloroethyl)-N'-nitrosocarbamoyl]amino Acids,II

Syntheses of N-[N'-(2-chloroethyl)-N'-nitrosocarbamoyl]amino acid amides and esters as potential antineoplastic substances are reported. N-[N'-(2-chloroethyl)-N'-nitrosocarbamoyl]amino acids (with the exception of the glycine derivative) were prepared by reaction of 2-chloroethyl isocyanate with the sodium salt of an amino acid in a heterogenous medium followed by nitrosation with sodium nitrite under acidic conditions. Condensation with amines or alcohols using 1,1-carbonyldiimidazole led to the amides or esters.