Carbocationoids, a concept for controlling highly reactive cationic species.

Carbocations, which are positively charged highly electrophilic intermediates, are efficacious for the direct alkylation of low-reactive nucleophiles. The utilization of carbocations in SN1 reactions relies on the activation of their precursors in the presence of a nucleophile. However, undesirable interactions between the nucleophile and the leaving group activator limit the scope of acceptable nucleophiles. Here we report a strategy to conduct SN1 reactions involving unstable carbocations in an alternative stepwise procedure, which was demonstrated by the benzylation of various neutral nucleophiles. In the first step, carbocations were accumulated in a nucleophile-free solution in the form of carbocationoids utilizing the coordinative stabilization of triazinediones. Subsequently, the addition of these solutions in the second step enabled room-temperature alkylation without the need for acidic additives. This methodology overcomes the inherent challenges of carbocations in SN1 reactions.

Drug-drug conjugates of MEK and Akt inhibitors for RAS-mutant cancers.

Controlling RAS mutant cancer progression remains a significant challenge in developing anticancer drugs. Whereas Ras G12C-covalent binders have received clinical approval, the emergence of further mutations, along with the activation of Ras-related proteins and signals, has led to resistance to Ras binders. To discover novel compounds to overcome this bottleneck, we focused on the concurrent and sustained blocking of two major signaling pathways downstream of Ras. To this end, we synthesized 25 drug-drug conjugates (DDCs) by combining the MEK inhibitor trametinib with Akt inhibitors using seven types of linkers with structural diversity. The DDCs were evaluated for their cell permeability/accumulation and ability to inhibit proliferation in RAS-mutant cell lines. A representative DDC was further evaluated for its effects on signaling proteins, induction of apoptosis-related proteins, and the stability of hepatic metabolic enzymes. These in vitro studies identified a series of DDCs, especially those containing a furan-based linker, with promising properties as agents for treating RAS-mutant cancers. Additionally, in vivo experiments in mice using the two selected DDCs revealed prolonged half-lives and anticancer efficacies comparable to those of trametinib. The PK profiles of trametinib and the Akt inhibitor were unified through the DDC formation. The DDCs developed in this study have potential as drug candidates for the broad inhibition of RAS-mutant cancers.

Phototriggered Butenolide Formation from a Cyclobutenedione and an Acidic Nucleophile.

A phototriggered conjugation reaction of an aminocyclobutenedione and an acidic nucleophile was discovered. Upon blue light irradiation of the materials, a butenolide derivative with substituents derived from the aminocyclobutenedione and the nucleophile was produced. The reaction proceeded efficiently under organic solvent or organic solvent/aqueous buffer (1/1) conditions. This reaction would be useful for the synthesis of unique butenolide derivatives and derivatization of acidic functional groups contained in aqueous biomolecules.

Epitope-Based Specific Antibody Modifications.

Modified antibodies have essential roles in analytic, diagnostic, and therapeutic uses, and thus, these antibodies are required to have optimal physical and biological properties. Consequently, the development of methods for site-selective antibody modification is crucial. Herein, we used epitope-based affinity labeling to introduce a Fab region-selective antibody modification method. Although labeling that exploits the high affinity between an antibody and its epitope may appear straightforward, it remains challenging probably because of the loss of target affinity caused by modification around the epitope-binding site. By thoroughly screening the modifying agent structure, reaction conditions, and purification methods, we developed an efficient method for the selective modification of the Fab region of the antibody while maintaining the high affinity for the epitope.

[The Human Resource Development Policy of the Faculty of Pharmaceutical Sciences at Kanazawa University and Our Initiatives toward Its Realization].

The environment surrounding pharmaceutical education in Japan has changed significantly with the establishment of many new pharmacy schools and the transition to six-year pharmacist education. Under these circumstances, various issues have been revealed in recent years. In particular, the decrease in the number of doctoral students responsible for future pharmaceutical education and research in drug discovery and life science is a concern. To address this issue, we at Kanazawa University have revised the human resource development policy of the Faculty of Pharmaceutical Sciences to "fostering leaders who are active in a variety of professions in pharmaceutical sciences" and have made various efforts toward the realization of this policy. Among the topics introduced at the symposium, this paper focuses on reforming the educational system and reorganizing the School of Pharmacy and Pharmaceutical Sciences at Kanazawa University.

Measurement of Hepatic CYP3A4 and 2D6 Activity Using Radioiodine-Labeled O-Desmethylvenlafaxine.

Drug metabolizing enzyme activity is affected by various factors such as drug-drug interactions, and a method to quantify drug metabolizing enzyme activity in real time is needed. In this study, we developed a novel radiopharmaceutical for quantitative imaging to estimate hepatic CYP3A4 and CYP2D6 activity. Iodine-123- and 125-labeled O-desmethylvenlafaxine (123/125I-ODV) was obtained with high labeling and purity, and its metabolism was found to strongly involve CYP3A4 and CYP2D6. SPECT imaging in normal mice showed that the administered 123I-ODV accumulated early in the liver and was excreted into the gallbladder, as evaluated by time activity curves. In its biological distribution, 125I-ODV administered to mice accumulated early in the liver, and only the metabolite of 125I-ODV was quickly excreted into the bile. In CYP3A4- and CYP2D6-inhibited model mice, the accumulation in bile decreased more than in normal mice, indicating inhibition of metabolite production. These results indicated that imaging and quantifying the accumulation of radioactive metabolites in excretory organs will aid in determining the dosages of various drugs metabolized by CYP3A4 and CYP2D6 for individualized medicine. Thus, 123/125I-ODV has the potential to direct, comprehensive detection and measurement of hepatic CYP3A4 and CYP2D6 activity by a simple and less invasive approach.

[Ideal Thesis Research and Clinical Training as Pharmaceutical Practice in 6-year Pharmaceutical Education and Study System].

Thesis research and clinical training are comprehensive areas of learning for cultivating the major abilities indicated in the professional competencies of the Model Core Curriculum for Pharmacy Education. They are also important subjects that can effectively reflect the educational philosophy and the human resource development policies of each university. In the School of Pharmacy and the School of Pharmaceutical Sciences at Kanazawa University, we have established a human resource development policy of "fostering leaders who are active in a variety of professions in the field of pharmaceutical sciences". We have been implementing various initiatives in line with this objective. In this symposium, we will present an analysis of the current situation in pharmaceutical education that led us to adopt such a human resource development policy. We will also introduce specific reforms that our university has promoted in response to this situation.

Synthesis and characterization of tetraphenylammonium salts.

The phenyl (Ph) group is a representative substituent in the field of organic chemistry as benzene (the parent molecule) is of fundamental importance. Simple Ph-substituted compounds of common chemical elements are well known. However, extensive structural characterization of tetraphenylammonium (Ph4N+) salts has not been reported. Herein, the synthesis of Ph4N+ salts and their characterization data including the 1H and 13C nuclear magnetic resonance (NMR) spectra and the single-crystal X-ray structure have been presented. An intermolecular radical coupling reaction between an aryl radical and a triarylammoniumyl radical cation was conducted to synthesize the target moieties. The Ph4N+ salts described herein are the simplest tetraarylammonium (Ar4N+) salts known. The results reported herein can potentially help access the otherwise inaccessible non-bridged Ar4N+ salts, a new class of rigid and sterically hindered organic cations.

Discovery of a Novel Aminocyclopropenone Compound That Inhibits BRD4-Driven Nucleoporin NUP210 Expression and Attenuates Colorectal Cancer Growth.

Epigenetic deregulation plays an essential role in colorectal cancer progression. Bromodomains are epigenetic "readers" of histone acetylation. Bromodomain-containing protein 4 (BRD4) plays a pivotal role in transcriptional regulation and is a feasible drug target in cancer cells. Disease-specific elevation of nucleoporin, a component of the nuclear pore complex (NPC), is a determinant of cancer malignancy, but BRD4-driven changes of NPC composition remain poorly understood. Here, we developed novel aminocyclopropenones and investigated their biological effects on cancer cell growth and BRD4 functions. Among 21 compounds developed here, we identified aminocyclopropenone 1n (ACP-1n) with the strongest inhibitory effects on the growth of the cancer cell line HCT116. ACP-1n blocked BRD4 functions by preventing its phase separation ability both in vitro and in vivo, attenuating the expression levels of BRD4-driven MYC. Notably, ACP-1n significantly reduced the nuclear size with concomitant suppression of the level of the NPC protein nucleoporin NUP210. Furthermore, NUP210 is in a BRD4-dependent manner and silencing of NUP210 was sufficient to decrease nucleus size and cellular growth. In conclusion, our findings highlighted an aminocyclopropenone compound as a novel therapeutic drug blocking BRD4 assembly, thereby preventing BRD4-driven oncogenic functions in cancer cells. This study facilitates the development of the next generation of effective and potent inhibitors of epigenetic bromodomains and extra-terminal (BET) protein family.

Synthesis of N-Aminated Salts of Aliphatic tert-Amines, (Trialkyl)amidines, and (Pentaalkyl)guanidines by Electrophilic Amination in an Ethereal Solvent.

The electrophilic amination of nitrogen-based nucleophiles, including strong organic bases, was conducted in an Et2O solvent using O-(mesitylenesulfonyl)hydroxylamine. Aliphatic tert-amines and N,N,N'-(trialkyl)amidines rapidly formed precipitates of the corresponding aminated salts in high yields. The amination of the highly basic and sterically hindered N,N,N',N',N″-(pentaalkyl)guanidines was achieved under modified conditions, although the yields were moderate because of a competing side reaction caused by the acid-base equilibrium.

A versatile iodo(III)etherification of terminal ethynylsilanes using BF3-OiPr2 and alkyl benzyl ethers.

A series of (E)-α-silyl-ß-alkoxyvinyl-λ3-iodanes was synthesized from iodosylbenzene, BF3-ether complexes, and terminal ethynylsilanes. The combined use of BF3-OiPr2 and benzyl ethers of primary alcohols (ROBn) allows the chemoselective transfer of primary alkoxy groups (RO) onto the ß-position of the terminal ethynylsilanes.

N-Acyltriazinedione; a Novel Acylating Reagent Synthesized from a Triazinone-Type Condensing Reagent.

In this paper, we report the synthesis of N-acyltriazinedione via the unexpected O-N acyl rearrangement of acyloxytriazinone and its utility as an acylating reagent. N-Acyltriazinedione can be isolated by silica gel column chromatography and reacts with amines in the absence of any base to give the corresponding amides in good yields.

Development of a triazinedione-based dehydrative condensing reagent containing 4-(dimethylamino)pyridine as an acyl transfer catalyst.

A new triazinedione-based reagent, (N,N'-dialkyl)triazinedione-4-(dimethylamino)pyridine (ATD-DMAP) was developed for the operationally simple dehydrative condensation of carboxylic acids. This reagent comprises an ATD core and DMAP as the leaving group, which is liberated into the reaction system to accelerate acyl transfer reactions. Upon adding ATD-DMAP to a mixture of carboxylic acids and alcohols in the presence of an amine base, the corresponding esters were formed rapidly at room temperature. Moreover, dehydrative condensation between carboxylic acids and amines using ATD-DMAP proceeded in high yield.

Efficiency Enhancement of a Photocatalytic Decarbonylation of an Aminocyclopropenone by Benzothiophene Substitution.

To improve the efficiency of the photocatalytic decarbonylation of cyclopropenones, the effects of substituents on cyclopropenone were explored. A benzothiophene-substituted aminocyclopropenone exhibited significantly improved decarbonylation efficiency to produce the corresponding ynamine, which worked as a potent dehydration condensation agent. The benzothiophene derivative was applicable to the photocatalytic reaction in the presence of potential excited-state quenchers such as oxygen and anilines. The high catalyst sensitivity would be attributed to the involvement of triplet energy transfer reaction pathway, which was not observed in the reaction with previously reported aminocyclopropenones.

6-Hydroxyindole is an endogenous long-lasting OATP1B1 inhibitor elevated in renal failure patients.

The hepatic uptake transporter organic anion transporting polypeptide (OATP) 1B1 is inhibited by some uremic toxins; however, direct inhibition can only partially explain the delayed systemic elimination of substrate drugs in renal failure patients. This study aimed to examine the long-lasting inhibition of OATP1B1 by uremic toxins and their metabolites. Preincubation of HEK293/OATP1B1 cells with 21 uremic toxins resulted in almost no change in the uptake of a typical substrate [3H]estrone-3-sulfate (E1S), although some directly inhibited [3H]E1S uptake. In contrast, preincubation with an indole metabolite, 6-hydroxyindole, reduced [3H]E1S uptake, even after the inhibitor was washed out before [3H]E1S incubation. Such long-lasting inhibition by 6-hydroxyindole was time-dependent and recovered after a 3-h incubation without 6-hydroxyindole. Preincubation with 6-hydroxyindole increased the Km for [3H]E1S uptake with minimal change in Vmax. This was compatible with no change in the cell-surface expression of OATP1B1, as assessed by a biotinylation assay. Preincubation with 6-hydroxyindole reduced [3H]E1S uptake in human hepatocytes without changes in OATP1B1 mRNA. Plasma concentration of 6-hydroxyindole in renal failure patients increased as renal function decreased, but might be insufficient to exhibit potent OATP1B1 inhibition. In conclusion, 6-hydroxyindole is an endogenous long-lasting OATP1B1 inhibitor with elevated plasma concentrations in renal failure patients.

Development of Hydrophilic Polyacrylamide Gel-Based Condensing Reagents Comprised of Chlorotriazine.

Hydrophilic polyacrylamide gel-based triazine-type condensing reagents, PAG-Trz-Cl, have been developed. PAG-Trz-Cls were synthesized using a chlorotriazine with an acrylamide moiety, acrylamide, and N,N'-methylenebisacrylamide via both precipitation and solution polymerization. Because PAG-Trz-Cls adequately swell in aqueous media, the amidation between polar carboxylic acids and amines afforded the corresponding amides in good yields.

Development of Triazinone-Based Condensing Reagents for Amide Formation.

Novel triazinone-based condensing reagents have been developed. The palladium-catalyzed O-N allylic rearrangement of 2-(allyloxy)-4,6-dichloro-1,3,5-triazine and subsequent regioselective substitution using alcohols and an amine afforded chlorotriazinones, which can be readily converted using N-methylmorpholine into the corresponding condensing reagents. The condensation of carboxylic acids and amines using these reagents proceeded to afford the desired amides in good yields. In comparison with 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride, the newly synthesized triazinone-based condensing reagents exhibited higher reactivity.

Substitution of the Dimethylamino Group in Gramines and One-Pot Cyclization to Tetrahydro-ß-carbolines Using a Triazine-Based Activating Agent.

A new method for the substitution of 3-[(dimethylamino)methyl]indoles (gramines) with malonate-based nucleophiles was developed using 2-chloro-4,6-dimethoxy-1,3,5-triazine (CDMT) as the activating agent for the dimethylamino group. The reaction was completed in 1.5-6 h at room temperature in the presence of a tert-amine base and lithium salt. CDMT afforded superior results to methyl iodide, a common activating agent for the dimethylamino group in Mannich bases, particularly in the reactions of 1-substituted gramines. The reactivity of the possible intermediates, bis(indol-3-ylmethyl)dimethylammonium salts, was examined to obtain mechanistic insights on the reaction. This substitution method with CDMT enabled the sequential transformation of gramines: substitution with ( N-alkylidene)aminomalonates followed by the Pictet-Spengler reaction under acidic conditions afforded 1,2,3,4-tetrahydro-ß-carboline derivatives in one pot.

Phototriggered Active Alkyne Generation from Cyclopropenones with Visible Light-Responsive Photocatalysts.

A photocatalytic active alkyne generation reaction was developed using cyclopropenone as a starting reagent. Visible light-responsive photocatalysts induced cyclopropenone decarbonylation. The resulting highly reactive alkyne could be used directly, without isolation, for further reactions, such as in a dehydration condensation reaction and alkyne-azide click chemistry.

Development of a Storable Triazinone-Based Reagent for O- p-Methoxybenzylation under Mild Heating Conditions.

A new triazinone-based reagent for O- p-methoxybenzylation has been developed. In spite of its stability in solid form, this reagent converts a free alcohol into the corresponding p-methoxybenzyl ether with mild heating (50-60 °C) in a solution. High functional group tolerance can be achieved because the reaction does not require the addition of an acidic or basic activator.