Synthesis of MAPA Reagents and 2-Alkyl(aryl)aminopyridines from 2-Bromopyridine Using the Goldberg Reaction.

A short and economical synthesis of various 2-methylaminopyidine amides (MAPA) from 2-bromopyridine has been developed using the catalytic Goldberg reaction. The effective catalyst was formed in situ by the reaction of CuI and 1,10-phenanthroline in a 1/1 ratio with a final loading of 0.5-3 mol%. The process affords high yields and can accommodate multigram-scale reactions. A modification of this method provides a new preparation of 2-N-substituted aminopyridines from various secondary N-alkyl(aryl)formamides and 2-bromopyridine. The intermediate aminopyridine formamide is cleaved in situ through methanolysis or hydrolysis to give 2-alkyl(aryl)aminopyridines in high yields.

Synthesis of C-4 Substituted Amido Nicotine Derivatives via Copper(I)- and (II)-Catalyzed Cross-Coupling Reactions.

The syntheses of seven novel amido nicotine derivatives 12-18 from (S)-nicotine are presented. (S)-Nicotine and (S)-6-chloronicotine derivatives were cross-coupled with the corresponding amides 6-10 at the C-4 position of the pyridine ring via copper(I)-mediated reactions. Derivatives 16-18 were also obtained via copper(II)-mediated reactions from (S)-nicotine containing a C-4 boronic acid pinacol ester group. The optimization of reaction conditions for both routes provided a useful method for preparing C-4 amide-containing nicotine analogs.

Studies toward the Synthesis of Lepadiformine A.

Herein is described an original approach to access a tricyclic framework of the lepadiformine-type alkaloids. A Grignard/N-acylpyridinium salt reaction of a 4-methoxytetrahydroquinoline is the key carbon-carbon bond-forming step that was used to establish the desired absolute stereochemistry at the C2 position of the target alkaloid. The synthesis features an allylation reaction with an N-acyliminium ion to set the C10 quaternary stereocenter, a mild dissolving-metal cleavage of hindered phenyl carbamates, and an aminoiodocyclization to form the pyrrolidine ring. While this route does not provide the correct C10 stereochemistry, it showcases an efficient method to build analogues with the ring system of this class of alkaloids in 11 steps overall.

A twist on facial selectivity of hydride reductions of cyclic ketones: twist-boat conformers in cyclohexanone, piperidone, and tropinone reactions.

The role of twist-boat conformers of cyclohexanones in hydride reductions was explored. The hydride reductions of a cis-2,6-disubstituted N-acylpiperidone, an N-acyltropinone, and tert-butylcyclohexanone by lithium aluminum hydride and by a bulky borohydride reagent were investigated computationally and compared to experiment. Our results indicate that in certain cases, factors such as substrate conformation, nucleophile bulkiness, and remote steric features can affect stereoselectivity in ways that are difficult to predict by the general Felkin-Anh model. In particular, we have calculated that a twist-boat conformation is relevant to the reactivity and facial selectivity of hydride reduction of cis-2,6-disubstituted N-acylpiperidones with a small hydride reagent (LiAlH4) but not with a bulky hydride (lithium triisopropylborohydride).

Total synthesis of alkaloid 205B.

Concise and highly stereocontrolled total syntheses of racemic and enantiopure frog alkaloid 205B (1) were accomplished in 11 steps from 4-methoxypyridines 6 and 7 in overall yields of 8 and 8%, respectively. The assembly of the core of the natural product relies on a stereoselective Tsuji-Trost allylic amination reaction and a ring-closing metathesis. The synthesis features the use of an N-acylpyridinium salt reaction to introduce the first stereocenter and an unprecedented trifluoroacetic anhydride-mediated addition of an allylstannane to a vinylogous amide with complete facial selectivity. Deoxygenation of the C4 ketone proved difficult but was accomplished via a modified Barton-McCombie reaction in the presence of a catalytic amount of diphenyl diselenide.

Studies toward the total synthesis of dihydrolycolucine. Preparation of AB and CEF ring fragments.

A strategy for the synthesis of the lycopodium alkaloid dihydrolycolucine (1) has been investigated. Synthetic routes were developed based on N-acylpyridinium salt chemistry to prepare target fragments 3 and 4 that could ultimately converge to the natural product. Key reactions include IMDA cycloadditions and retro-Mannich ring-openings to form both the AB and the EF ring fragments. The ring C precursor was prepared using pyridine substitution and directed lithiation chemistry. A Suzuki cross-coupling of rings C and EF led to the CEF ring fragment. Initial attempts at closure of the seven-membered D ring were unsuccessful.

Hydrophobic derivatization of N-linked glycans for increased ion abundance in electrospray ionization mass spectrometry.

A library of neutral, hydrophobic reagents was synthesized for use as derivatizing agents in order to increase the ion abundance of N-linked glycans in electrospray ionization mass spectrometry (ESI MS). The glycans are derivatized via hydrazone formation and are shown to increase the ion abundance of a glycan standard more than 4-fold. Additionally, the data show that the systematic addition of hydrophobic surface area to the reagent increases the glycan ion abundance, a property that can be further exploited in the analysis of glycans. The results of this study will direct the future synthesis of hydrophobic reagents for glycan analysis using the correlation between hydrophobicity and theoretical non-polar surface area calculation to facilitate the development of an optimum tag for glycan derivatization. The compatibility and advantages of this method are demonstrated by cleaving and derivatizing N-linked glycans from human plasma proteins. The ESI-MS signal for the tagged glycans are shown to be significantly more abundant, and the detection of negatively charged sialylated glycans is enhanced.

Identification and replication of loci involved in camptothecin-induced cytotoxicity using CEPH pedigrees.

To date, the Centre d'Etude Polymorphism Humain (CEPH) cell line model has only been used as a pharmacogenomic tool to evaluate which genes are responsible for the disparity in response to a single drug. The purpose of this study was demonstrate the model's ability to establish a specific pattern of quantitative trait loci (QTL) related to a shared mechanism for multiple structurally related drugs, the camptothecins, which are Topoisomerase 1 inhibitors. A simultaneous screen of six camptothecin analogues for in vitro sensitivity in the CEPH cell lines resulted in cytotoxicity profiles and orders of potency which were in agreement with the literature. For all camptothecins studied, heritability estimates for cytotoxic response averaged 23.1 ± 2.6%. Nonparametric linkage analysis was used to identify a relationship between genetic markers and response to the camptothecins. Ten QTLs on chromosomes 1, 3, 5, 6, 11, 12, 16 and 20 were identified as shared by all six camptothecin analogues. In a separate validation experiment, nine of the ten QTLs were replicated at the significant and suggestive levels using three additional camptothecin analogues. To further refine this list of QTLs, another validation study was undertaken and seven of the nine QTLs were independently replicated for all nine camptothecin analogues. This is the first study using the CEPH cell lines that demonstrates that a specific pattern of QTLs could be established for a class of drugs which share a mechanism of action. Moreover, it is the first study to report replication of linkage results for drug-induced cytotoxicity using this model. The QTLs, which have been identified as shared by all camptothecins and replicated across multiple datasets, are of considerable interest; they harbor genes related to the shared mechanism of action for the camptothecins, which are responsible for variation in response.

Asymmetric synthesis of all the known phlegmarine alkaloids.

The asymmetric synthesis of all four of the known natural phlegmarines and one synthetic derivative has been accomplished in 19-22 steps from 4-methoxy-3-(triisopropylsilyl)pyridine. Chiral N-acylpyridinium salt chemistry was used twice to set the stereocenters at the C-9 and C-2' positions of the phlegmarine skeleton. Key reactions include the use of a mixed Grignard reagent for the second N-acylpyridinium salt addition, zinc/acetic acid reduction of a complex dihydropyridone, and a von Braun cyanogen bromide N-demethylation of a late intermediate. These syntheses confirmed the absolute stereochemistry of all of the known phlegmarines.

[2 + 2] Photochemical cycloaddition/ring opening of 6-alkenyl-2,3-dihydro-4-pyridones.

During the course of a study aimed at constructing azaspirocycles from 2,3-dihydro-4-pyridones, an unexpected product was obtained in the SET ring-opening reaction of photocycloadduct 1. Differences in reactivity between homologues 1 and 2 were observed in the presence of SmI(2). Tricyclic ketone 2 afforded azaspiro[5.5]undecane 15 when treated with SmI(2); however, when ketone 1 was submitted to similar reaction conditions a double ring-opening/reduction sequence gave cis-piperidinol 10.

A five-step synthesis of (S)-macrostomine from (S)-nicotine.

A concise synthesis of (S)-macrostomine has been accomplished in five steps from natural nicotine in 19% overall yield via a pyridyne Diels-Alder cycloaddition reaction as the key step. A Kumada cross-coupling reaction on a 1-chloroisoquinoline intermediate provided the natural product.

Interplay of permanent charge and hydrophobicity in the electrospray ionization of glycans.

The analysis of N-linked glycans by mass spectrometry (MS) has been characterized by low signal-to-noise ratios and high limits of detection due to their hydrophilicity and lack of basic sites able to be protonated. As a result, every step in glycan sample preparation must be thoroughly optimized in order to minimize sample loss, contamination, and analytical variability. Importantly, properties of glycans and their derivatized counterparts must be thoroughly studied in order to exploit certain characteristics for enhancing MS analysis. Herein, the effectiveness of the incorporation of a permanent charge is studied and determined to hamper glycan analysis. Also, a procedure for glycan hydrazone formation is optimized and outlined where a large number of variables were simultaneously analyzed using a fractional factorial design (FFD) in order to determine which conditions affected the reaction efficiency of the hydrazone formation reaction. Finally, the hydrophobic tagging of glycans is shown to be a viable opportunity to further increase the ion abundance of glycans in MS.

Synthesis of fused-ring nicotine derivatives from (S)-nicotine.

The synthesis of novel fused-ring bicyclic (4-6) and tricyclic (7-10) nicotine derivatives from natural (S)-nicotine are described. Enantiopure bicyclic dioxino, dihydrofuro, and dihydropyranol nicotine derivatives as well as tricylic benzofuro and benzopyran derivatives were synthesized from simple alkoxy or halonicotine intermediates. Attempts to synthesize furonicotines (11, 12) resulted in formation of the furonicotine dimers 42 and 49.

Improving limits of detection for B-type natriuretic peptide using PC-IDMS: an application of the ALiPHAT strategy.

Hydrophobic tagging of biomolecules has been reported by our group and others to increase their ionization efficiency during electrospray ionization and facilitate their detection by mass spectrometry. As such, hydrophobic tagging should provide a viable method for augmenting MS-based quantification of low abundance proteins by decreasing their detection limits. Herein we have evaluated two commercial alkylation reagents and several newly synthesized hydrophobic alkylation reagents for their utility in quantifying B-type Natriuretic Peptide, a low abundance cardiac biomarker, by protein cleavage isotope dilution mass spectrometry. For the cysteine containing tryptic peptide evaluated, a approximately 3.5-fold decrease in the detection limit was observed for the best performing hydrophobic reagent, 2-iodo-N-octylacetamide, relative to the commonly used alkylation reagent, iodoacetamide. Additionally, we have evaluated the use of nonpolar surface areas as a metric for assessing the effectiveness of the alkylation reagents in improving ESI response.

Increasing the hydrophobicity and electrospray response of glycans through derivatization with novel cationic hydrazides.

Novel tags are used to increase the hydrophobicity of glycans and impart a permanent charge yielding as great as a approximately 5-fold increase in electrospray response from both a standard and complex mixture.

Evaluation of the ALiPHAT method for PC-IDMS and correlation of limits-of-detection with nonpolar surface area.

PC-IDMS experiments for two peptides, laminin nonapeptide and the N-terminal tryptic peptide of prostate specific antigen, were performed utilizing a variety of alkylating reagents. These experiments were conducted to investigate how hydrophobicity influences the limits-of-detection (LOD) by altering their electrospray ionization response. Nonpolar surface areas were calculated for both peptides and all alkylating reagents to provide an estimate of the hydrophobicity of the differently alkylated peptides. Decreases in LOD by 2-fold were observed for both peptides between the best and worst performing combination of alkylating reagent. However, while an increase in hydrophobicity was found to aid in decreasing LOD to an extent, beyond a certain hydrophobicity, we observed a decrease.

Synthesis of 1,3-amino alcohol derivatives via a silicon-mediated ring-opening of substituted piperidines.

Multisubstituted piperidines containing a trimethylsilylmethyl group at C-2 can be opened regioselectively with TBAF and cyanogen bromide. The ring-opened products contain synthetically useful cyanamide and terminal alkene functional groups. This method is useful for the stereoselective synthesis of alkylamine derivatives containing multiple chiral centers.

N-acyldihydropyridones as synthetic intermediates. A stereoselective synthesis of acyclic amino alcohols containing multiple chiral centers.

Various multisubstituted piperidines containing a phenyl group at C-2 can be opened regio- and stereoselectively with cyanogen bromide. The ring-opened products contain useful cyanamide and benzylic bromide functional groups. The benzyl bromide can be cleanly reduced, or substituted with various nucleophiles via an S(N)2 process to add additional heteroatoms stereoselectively. This methodology is useful for the stereoselective synthesis of uniquely substituted alkylamine derivatives containing multiple chiral centers and various functionality. Diastereomerically pure amino alcohols containing three to five contiguous stereocenters were prepared using this strategy.

Stereoselective synthesis of acyclic amino alcohols via von Braun ring opening of chiral piperidines.

Multisubstituted piperidines containing a phenyl group at C-2 can be opened regio- and stereoselectively with cyanogen bromide. The ring-opened products contain useful cyanamide and benzylic bromide functional groups. This methodology is useful for the stereoselective synthesis of uniquely substituted alkylamine derivatives containing multiple chiral centers and various functionality.