Cyclative Release Strategy to Obtain Pure Cyclic Peptides Directly from the Solid Phase.

The synthesis of large numbers of cyclic peptides─required, for example, in screens for drug development─is currently limited by the need of chromatographic purification of individual peptides. Herein, we have developed a strategy in which cyclic peptides are released from the solid phase in the pure form and do not need purification. Peptides with an N-terminal thiol group are synthesized on the solid phase via a C-terminal disulfide linker, their sidechain-protecting groups are removed while the peptides remain on the solid phase, and the peptides are finally released via a cyclative mechanism by the addition of a base that deprotonates the N-terminal thiol group and triggers an intramolecular disulfide-exchange reaction. The method yields disulfide-cyclized peptides, a format on which many important peptide drugs such as oxytocin, vasopressin, and octreotide are based. We demonstrate that the method is applicable for facile synthesis in 96-well plates and allows for synthesis and screening of hundreds of cyclic peptides.

A Raf-like kinase is required for smoke-induced seed dormancy in Arabidopsis thaliana.

Plants sense and integrate diverse stimuli to determine the timing for germination. A smoke compound, 3,4,5-trimethylfuran-2(5H)-one (trimethylbutenolide, TMB), has been identified to inhibit the seed germination of higher plants. To understand the mode of action, we examined various physiological and molecular aspects of the TMB-dependent inhibition of seed germination in Arabidopsis thaliana The results indicated that the effect of TMB is due to the enhanced physiological dormancy, which is modulated by other dormancy regulatory cues such as after-ripening, stratification, and ABA/GA signaling. In addition, gene expression profiling showed that TMB caused genome-wide transcriptional changes, altering the expression of a series of dormancy-related genes. Based on the TMB-responsive physiological contexts in Arabidopsis, we performed mutant screening to isolate genetic components that underpin the TMB-induced seed dormancy. As a result, the TMB-RESISTANT1 (TES1) gene in Arabidopsis, encoding a B2 group Raf-like kinase, was identified. Phenotypic analysis of the tes1 mutant implicated that TES1 has a critical role in the TMB-responsive gene expression and the inhibition of seed germination. Taken together, we propose that plants have been equipped with a TMB sensory pathway through which the TMB induces the seed dormancy in a TES1-dependent way.

Targeted Degradation of Transcription Coactivator SRC-1 through the N-Degron Pathway.

Aberrantly elevated steroid receptor coactivator-1 (SRC-1) expression and activity are strongly correlated with cancer progression and metastasis. Here we report, for the first time, the development of a proteolysis targeting chimera (PROTAC) that is composed of a selective SRC-1 binder linked to a specific ligand for UBR box, a unique class of E3 ligases recognizing N-degrons. We showed that the bifunctional molecule efficiently and selectively induced the degradation of SRC-1 in cells through the N-degron pathway. Importantly, given the ubiquitous expression of the UBR protein in most cells, PROTACs targeting the UBR box could degrade a protein of interest regardless of cell types. We also showed that the SRC-1 degrader significantly suppressed cancer cell invasion and migration in vitro and in vivo. Together, these results demonstrate that the SRC-1 degrader can be an invaluable chemical tool in the studies of SRC-1 functions. Moreover, our findings suggest PROTACs based on the N-degron pathway as a widely useful strategy to degrade disease-relevant proteins.

Oligomers of α-ABpeptoid/ß3 -peptide hybrid.

Peptoids, oligomers of N-substituted glycines, have been attracting increasing interest due to their advantageous properties as peptidomimetics. However, due to the lack of chiral centers and amide hydrogen atoms, peptoids, in general, do not form folding structures except that they have α-chiral side chains. We have recently developed "peptoids with backbone chirality" as a new class of peptoid foldamers called α-ABpeptoids and demonstrated that they could have folding conformations owing to the methyl groups on chiral α-carbons in the backbone structure. Here we report α-ABpeptoid/ß3 -peptide oligomers as a unique peptidomimetic structure with a heterogeneous backbone. This hybrid structure contains a mixed α-ABpeptoid and ß3 -peptide residues arranged in an alternate manner. These α-ABpeptoid/ß3 -peptide oligomers could form intramolecular hydrogen bonding and have better cell permeability relative to pure peptide sequences. These oligomers were shown to adopt ordered folding structures based on circular dichroism studies. Overall, α-ABpeptoid/ß3 -peptide oligomers may represent a novel class of peptidomimetic foldamers and will find a wide range of applications in biomedical and material sciences.

Solid-Phase Synthesis and Circular Dichroism Study of ß-ABpeptoids.

The development of peptidomimetic foldamers that can form well-defined folded structures is highly desirable yet challenging. We previously reported on α-ABpeptoids, oligomers of N-alkylated ß²-homoalanines and found that due to the presence of chiral methyl groups at α-positions, α-ABpeptoids were shown to adopt folding conformations. Here, we report ß-ABpeptoids having chiral methyl group at ß-positions rather than α-positions as a different class of peptoids with backbone chirality. We developed a facile solid-phase synthetic route that enables the synthesis of ß-ABpeptoid oligomers ranging from 2-mer to 8-mer in excellent yields. These oligomers were shown to adopt ordered folding conformations based on circular dichroism (CD) and NMR studies. Overall, these results suggest that ß-ABpeptoids represent a novel class of peptidomimetic foldamers that will find a wide range of applications in biomedical and material sciences.

Synthesis and biological evaluation of potent benzoselenophene and heteroaromatic analogues of (S)-1-(chloromethyl)-8-methoxy-2,3-dihydro-1H-benzo[e]indol-5-ol (seco-MCBI).

A diverse series of compounds (18a-x) were synthesized from (S)-1-(chloromethyl)-8-methoxy-2,3-dihydro-1H-benzo[e]indol-5-ol (seco-MCBI) and benzoselenophene or heteroaromatic acids. These new compounds were evaluated for their cytotoxicity against the human gastric NCI-N87 and human ovarian SK-OV3 cancer cell lines. The incorporation of a methoxy substituent at the C-7 position of the seco-CBI unit enhances the cytotoxicity through its additional van der Waals interaction and gave a much higher potency than the corresponding seco-CBI-based analogues. Similarly, the seco-MCBI-benzoselenophene conjugates (18h-x) exhibited substitution effects on biological activity, and the N-butyramido and N-methylthiopropanamido analogues are highly potent, possessing >77- and >24-fold better activity than seco-MCBI-TMI for the SK-OV3 and NCI-N87 cell lines, respectively.

Submonomer Strategy toward Divergent Solid-Phase Synthesis of α-ABpeptoids.

A novel submonomer solid-phase synthetic method for α-ABpeptoid oligomers is reported. Iterative submonomer coupling and Fukuyama-Mitsunobu alkylation enable facile, divergent synthesis of α-ABpeptoid oligomers substituted with chemically diverse side chains in excellent yields.

Solid-Phase Synthesis of Triostin A Using a Symmetrical Bis(diphenylmethyl) Linker System.

Triostin A is a symmetric bicyclic depsipeptide with very potent antitumoral activity because of its bisintercalation into DNA. In this study, we report a new synthetic strategy that exploits a structural symmetry of triostin A. First, we prepared a novel symmetric linker molecule that is labile under mildly acidic conditions and suitable for a solid-phase synthesis procedure. Two Cys units were attached to a linker-resin conjugate via their free thiol groups, and double deprotection and double coupling reactions were then applied to synthesize linear tetradepsipeptides. Subsequently, the key biscyclization of the tetradepsipeptides was performed on the resin, and the resulting cyclic octapeptide was detached from the linker-resin conjugate to give a peptide with two free thiols. Finally, triostin A was obtained by oxidizing the free thiols in solution to produce a disulfide. The yield was improved through exploration of two different solid-phase synthetic approaches under similar strategy. Mainly, this strategy was developed to enable the ease and rapid preparation of libraries of symmetric bicyclic depsipeptides. It also addresses several synthetic problems with our synthesis, including diketopiperazine (DKP) formation, poor cyclization yields and preparation of noncommercial N-methyl amino acids in good yields.