Inhibitory Effect of Pyra-Metho-Carnil on Cancer Spheroid Growth Through Decrease in Glycolysis-associated Molecules.

BACKGROUND/AIM: Pyra-Metho-Carnil (PMC) has been identified as a novel candidate compound for treating numerous malignancies; however, its mechanism of action remains unknown. In this study, we conducted RNA-sequencing (RNA-seq) analyses to elucidate the mechanism of PMC against human colorectal cancer cells harboring mutant KRAS (mtKRAS). MATERIALS AND METHODS: RNA-seq analyses of the HKe3-wild-type KRAS and HKe3-mtKRAS spheroids treated with DMSO or PMC for 6 days were performed. RESULTS: RNA-seq data suggested that PMC treatment suppresses the aerobic glycolysis pathway in HKe3-mtKRAS spheroids through the down-regulation of the HIF1 pathway. Indeed, treatment with PMC markedly suppresses the absorption of glucose by spheroids and the secretion of lactate from them. CONCLUSION: PMC suppresses growth of cancer spheroid through down-regulation of cancer-specific glucose metabolism.

Cancer Spheroid Proliferation Is Suppressed by a Novel Low-toxicity Compound, Pyra-Metho-Carnil, in a Context-independent Manner.

BACKGROUND/AIM: In a screen of compounds to selectively suppress the growth of cancer spheroids, which contained mutant (mt) KRAS, NPD10621 was discovered and associated derivatives were investigated. MATERIALS AND METHODS: Spheroid areas from HCT116-derived HKe3 spheroids expressing wild type (wt) KRAS (HKe3-wtKRAS) and mtKRAS (HKe3-mtKRAS) were treated with 12 NPD10621 derivatives and measured in three-dimensional floating (3DF) cultures. Several cancers were treated with NPD1018 (pyra-metho-carnil: PMC) in 3DF cultures. In a nude mouse assay, 50% cell growth inhibition (GI50) values were determined. RESULTS: From these 12 derivatives, PMC was the most effective inhibitor of HKe3-mtKRAS spheroid growth with the least toxicity. Furthermore, PMC-mediated growth suppression was observed in all tested cancer cell lines, independent of tissue context, driver gene mutations, and drug resistance, suggesting that the PMC target(s) was crucial for cancer growth in a context-independent manner. The GI50 value of PMC in nude mice assay was 7.7 mg/kg and nude mice that were administered 40 mg/kg PMC for 7 days did not show any abnormal blood cell count values. CONCLUSION: PMC is a low-toxicity compound that inhibits the growth of different tumor cell types.

Catalytic Dehydrogenative ß-Alkylation of Amino Acid Schiff Bases with Hydrocarbon.

A synthetic method for the synthesis of a highly congested α,ß-dehydroamino acid through the ß-C-H bond activation of an amino acid Schiff base is described. Abundant hydrocarbon feedstock could be used as an alkylating reagent to afford an α,ß-dehydroamino acid bearing a quaternary carbon at the γ-position with an exclusively (Z)-geometry. Notably, a tetrasubstituted olefin could be constructed from saturated starting materials. The transformation of the synthesized α,ß-dehydroamino acid into unnatural α-amino acid derivatives was also demonstrated.

Growth Suppression of Cancer Spheroids With Mutated KRAS by Low-toxicity Compounds from Natural Products.

BACKGROUND/AIM: Among compounds from natural products selectively suppressing the growth of cancer spheroids, which have mutant (mt) KRAS, NP910 was selected and its derivatives explored. MATERIALS AND METHODS: The area of HKe3 spheroids expressing wild type (wt) KRAS (HKe3-wtKRAS) and mtKRAS (HKe3-mtKRAS) were measured in three-dimensional floating (3DF) cultures treated with 18 NP910 derivatives. The 50% cell growth inhibition (GI50) was determined by long-term 3DF (LT3DF) culture and nude mice assay. RESULTS: We selected NP882 (named STAR3) as the most effective inhibitor of growth of HKe3-mtKRAS spheroids with the least toxicity among NP910 derivatives. GI50s of STAR3 in LT3DF and nude mice assay were 6 µM and 30.75 mg/kg, respectively. However, growth suppression by STAR3 was observed in 50% of cell lines independent of KRAS mutation, suggesting that the target of STAR3 was not directly associated with KRAS mutation and KRAS-related signals. CONCLUSION: STAR3 is a low-toxicity compound that inhibits growth of certain tumour cells.

Development of Catalytic Reactions for Precise Control of Chemoselectivity.

Catalytic chemoselective reactions of innately less reactive functionalities over more reactive functionalities are described. A cooperative catalyst comprising a soft Lewis acid/hard Brønsted base enabled chemoselective activation of a hydroxyl group over an amino group, allowing for nucleophilic addition to electron-deficient olefins. The reaction could be applicable for a variety of amino alcohols, including pharmaceuticals, without requiring a tedious protection-deprotection process. Chemoselective enolization and subsequent α-functionalization of carboxylic acid derivatives were also achieved by a redox active catalyst through the radical process, providing unnatural α-amino/hydroxy acid derivatives bearing a complex carbon framework and a diverse set of functionalities. The present chemoselective catalysis described herein offers new opportunities to expand the chemical space for innovative drug discovery research.

Catalytic Aerobic Cross-Dehydrogenative Coupling of Azlactones en Route to α,α-Disubstituted α-Amino Acids.

We developed a catalytic aerobic method to synthesize α,α-disubstituted α-amino acids through cross-dehydrogenative coupling of azlactones. Combining an iron catalyst with a bisoxazolidine ligand resulted in high catalytic performance, and cross-coupling with an indole proceeded smoothly under aerobic conditions. A wide variety of α-aryl and aliphatic amino acid derived azlactones were applied to the present catalysis. In addition, a quaternary carbon could be constructed using oxindole and benzofuranone under aerobic conditions.

Amino Acid Schiff Base Bearing Benzophenone Imine As a Platform for Highly Congested Unnatural α-Amino Acid Synthesis.

Unnatural α-amino acids are invaluable building blocks in synthetic organic chemistry and could upgrade the function of peptides. We developed a new mode for catalytic activation of amino acid Schiff bases, serving as a platform for highly congested unnatural α-amino acid synthesis. The redox active copper catalyst enabled efficient cross-coupling to construct contiguous tetrasubstituted carbon centers. The broad functional group compatibility highlights the mildness of the present catalysis. Notably, we achieved successive ß-functionalization and oxidation of amino acid Schiff bases to afford dehydroalanine derivatives bearing tetrasubstituted carbon. A three-component cross-coupling reaction of an amino acid Schiff base, alkyl bromides, and styrene derivatives demonstrated the high utility of the present method. The diastereoselective reaction was also achieved using menthol derivatives as a chiral auxiliary, delivering enantiomerically enriched α-amino acid bearing α,ß-continuous tetrasubstituted carbon. The synthesized highly congested unnatural α-amino acid could be derivatized and incorporated into peptide synthesis.

Chemoselective Catalytic α-Oxidation of Carboxylic Acids: Iron/Alkali Metal Cooperative Redox Active Catalysis.

We developed a chemoselective catalytic activation of carboxylic acid for a 1e- radical process. α-Oxidation of a variety of carboxylic acids, which preferentially undergo undesired decarboxylation under radical conditions, proceeded efficiently under the optimized conditions. Chemoselective enolization of carboxylic acid was also achieved even in the presence of more acidic carbonyls. Extensive mechanistic studies revealed that the cooperative actions of iron species and alkali metal ions derived from 4 Å molecular sieves substantially facilitated the enolization. For the first time, catalytic enolization of unprotected carboxylic acid was achieved without external addition of stoichiometric amounts of Brønsted base. The formed redox-active heterobimetallic enediolate efficiently coupled with free radical TEMPO, providing synthetically useful α-hydroxy and keto acid derivatives.

Growth Suppression of Human Colorectal Cancer Cells with Mutated KRAS by 3-Deaza-cytarabine in 3D Floating Culture.

BACKGROUND/AIM: During screening for compounds that selectively suppress growth of human colorectal cancer (CRC) spheroids with mutant (mt) KRAS, the uridine analogue, 5-bromouridine (BrUrd) was identified and its derivatives were explored. MATERIALS AND METHODS: DNA incorporation in two-dimensional (2D) and three-dimensional floating (3DF) cultures was examined with the uridine analogue, 5-ethynyl-2'-deoxyuridine (EdU). The area of HKe3 CRC spheroids expressing wild type (wt) KRAS (HKe3-wtKRAS) and mtKRAS (HKe3-mtKRAS) were measured in 3DF culture with 11 BrUrd derivatives. RESULTS: EdU was strongly incorporated into newly-synthesized DNA from HKe3-mtKRAS cells compared to HKe3-wtKRAS in 2D and 3DF culture. 3-Deaza-cytarabine, which has properties of BrUrd and cytidine, was the most effective inhibitor of HKe3-mtKRAS spheroids with the least toxicity to HKe3-wtKRAS. Growth suppression of 3-deaza-cytarabine was stronger than cytarabine in 2D culture, and toxicity was lower than gemcitabine in long-term 3DF culture. CONCLUSION: 3-Deaza-cytarabine exhibits properties useful for the treatment of CRC patients with mtKRAS.

Strategy for Catalytic Chemoselective Cross-Enolate Coupling Reaction via a Transient Homocoupling Dimer.

A new strategy, a transient homocoupling dimer strategy, for direct catalytic oxidative cross-enolate coupling reactions is developed. Cross-enolate coupling products bearing a (contiguous) tetrasubstituted carbon center were obtained chemoselectively without the need for stoichiometric amounts of strong bases/metal oxidants, and thus, the present catalysis provides a general method for the synthesis of unnatural α,α-disubstituted amino acid motifs. The distinct transformation of azlactone and 2-acylimidazole units highlighted the synthetic utility of the present catalysis.

An Expeditious Route to trans-Configured Tetrahydrothiophenes Enabled by Fe(OTf)3 -Catalyzed [3+2] Cycloaddition of Donor-Acceptor Cyclopropanes with Thionoesters.

A synthetic route to trans-configured tetrahydrothiophenes (THTs) through Fe(OTf)3 -promoted [3+2] cycloaddition of donor-acceptor cyclopropanes with thionoesters was developed. The cycloaddition proceeded in high yield with high diastereoselectivity, affording transient α-alkoxy THTs. Not only aromatic and aliphatic thionoesters, but also thionolactone were applicable to the present iron catalysis. Further transformation of the S,O-ketal functionality of the product was achieved in a highly trans diastereoselective manner. Moreover, the utility of our methodology was clearly demonstrated by the synthesis of enantioenriched trans-configured THTs.

Catalytic Aerobic Chemoselective α-Oxidation of Acylpyrazoles en Route to α-Hydroxy Acid Derivatives.

Catalytic aerobic chemoselective α-oxidation of acylpyrazoles is described. Acylpyrazoles, carboxylic acid oxidation state substrates, were efficiently oxidized under aerobic conditions using TEMPO as an oxygenating agent. The mild catalytic conditions of the present catalysis were amenable to late-stage α-oxidation of various pharmaceutical agents and natural products, leading to previously unreported α-hydroxy acid derivatives in short steps. Preliminary mechanistic studies revealed that in situ generated copper(II) peroxo species served as a Lewis acid/Brønsted base cooperative catalyst.

Catalytic Chemoselective Conjugate Addition of Amino Alcohols to α,ß-Unsaturated Ester: Hydroxy Group over Amino Group and Conjugate Addition over Transesterification.

A highly chemoselective conjugate addition of amino alcohols to α,ß-unsaturated ester using a soft Lewis acid/hard Brønsted base cooperative catalyst was developed. This catalysis achieved chemoselective addition of a hydroxy group over an amino group. Moreover, soft metal alkoxide generation enabled chemoselective soft conjugate addition over hard transesterification. Various amino alcohols, including unprecedented cyclic ß-amino alcohol, were applicable to the present catalysis.

Chemo- and Regioselective Direct Functional Group Installation through Catalytic Hydroxy Group Selective Conjugate Addition of Amino Alcohols to α,ß-Unsaturated Sulfonyl Compounds.

A chemoselective functional group installation through catalytic hydroxy group selective conjugate addition of amino alcohols to a variety of functionalized α,ß-unsaturated sulfonyl derivatives was developed. Azide group installation for click chemistry and facile fluorescent labeling onto the less reactive hydroxy group demonstrated the synthetic utility of the present chemoselective catalysis. Moreover, chemo- and regioselective reaction of an unprotected amino diol was achieved for the first time.

µ-Oxo-Dinuclear-Iron(III)-Catalyzed O-Selective Acylation of Aliphatic and Aromatic Amino Alcohols and Transesterification of Tertiary Alcohols.

A highly chemoselective and reactive µ-oxo-dinuclear iron(III) salen catalyst for transesterification was developed. The developed iron complex catalyzed acylation of aliphatic amino alcohols with nearly perfect O-selectivity, even when using activated esters, for which chemoselectivity is more difficult to control. In addition, O-selective transesterification of aromatic amino alcohols was achieved for the first time. The high activity of the iron complex enabled the use of sterically congested tertiary alcohols, including unprecedented tert-butanol.

Direct Catalytic Chemoselective α-Amination of Acylpyrazoles: A Concise Route to Unnatural α-Amino Acid Derivatives.

A direct copper-catalyzed highly chemoselective α-amination is described. Acylpyrazole proved to be a highly efficient enolate precursor of a carboxylic acid oxidation state substrate, while preactivation by a stoichiometric amount of strong base has been used in catalytic α-aminations. The simultaneous activation of both coupling partners, enolization and metal nitrenoid formation, was crucial for obtaining the product, and wide functional group compatibility highlighted the mildness of the present catalysis. The bidentate coordination mode was amenable to highly chemoselective activation over ketone and much more acidic nitroalkyl functionality. Deuterium exchange experiments clearly demonstrated that exclusive enolization of acylpyrazole was achieved without the formation of a nitronate. The present catalysis was applied to late-stage α-amination, allowing for concise access to highly versatile α-amino acid derivatives. The product could be transformed into variety of useful building blocks.

Oxetanyl peptides: novel peptidomimetic modules for medicinal chemistry.

The synthesis of novel oxetanyl peptides, where the amide bond is replaced by a non-hydrolyzable oxetanylamine fragment, is reported. This new class of pseudo-dipeptides with the same H-bond donor/acceptor pattern found in proteins expands the repertoire of peptidomimetics.

Chemoselective catalytic conjugate addition of alcohols over amines.

A highly chemoselective conjugate addition of alcohols in the presence of amines is described. The cooperative nature of the catalyst enabled chemoselective activation of alcohols over amines, allowing the conjugate addition to soft Lewis basic α,ß-unsaturated nitriles. Divergent transformation of the nitrile functionality highlights the utility of the present catalysis.

Direct catalytic asymmetric alkynylation of ketoimines.

An efficient protocol for direct catalytic alkynylation of ketoimines is described. The simultaneous activation of a soft Lewis basic terminal alkyne and a ketoimine bearing a thiophosphinoyl group by soft Lewis acid Cu(I) is crucial for high conversion. The reaction can be rendered asymmetric with a chiral bisphosphine ligand (S,S)-Ph-BPE.

Concise enantioselective synthesis of duloxetine via direct catalytic asymmetric aldol reaction of thioamide.

Direct catalytic asymmetric aldol reaction of thioamide offers a new entry to the concise enantioselective synthesis of duloxetine. The direct aldol protocol was scalable (>20 g) to afford the aldol product in 92% ee after LiAlH(4) reduction, and 84% of the chiral ligand was recovered after recrystallization. The following four steps of transformation delivered duloxetine.