Plasticity of Extrachromosomal and Intrachromosomal BRAF Amplifications in Overcoming Targeted Therapy Dosage Challenges.

Focal amplifications (FA) can mediate targeted therapy resistance in cancer. Understanding the structure and dynamics of FAs is critical for designing treatments that overcome plasticity-mediated resistance. We developed a melanoma model of dual MAPK inhibitor (MAPKi) resistance that bears BRAFV600 amplifications through either extrachromosomal DNA (ecDNA)/double minutes (DM) or intrachromosomal homogenously staining regions (HSR). Cells harboring BRAFV600E FAs displayed mode switching between DMs and HSRs, from both de novo genetic changes and selection of preexisting subpopulations. Plasticity is not exclusive to ecDNAs, as cells harboring HSRs exhibit drug addiction-driven structural loss of BRAF amplicons upon dose reduction. FA mechanisms can couple with kinase domain duplications and alternative splicing to enhance resistance. Drug-responsive amplicon plasticity is observed in the clinic and can involve other MAPK pathway genes, such as RAF1 and NRAS. BRAF FA-mediated dual MAPKi-resistant cells are more sensitive to proferroptotic drugs, extending the spectrum of ferroptosis sensitivity in MAPKi resistance beyond cases of dedifferentiation. SIGNIFICANCE: Understanding the structure and dynamics of oncogene amplifications is critical for overcoming tumor relapse. BRAF amplifications are highly plastic under MAPKi dosage challenges in melanoma, through involvement of de novo genomic alterations, even in the HSR mode. Moreover, BRAF FA-driven, dual MAPKi-resistant cells extend the spectrum of resistance-linked ferroptosis sensitivity. This article is highlighted in the In This Issue feature, p. 873.

Compound Uptake into E. coli Can Be Facilitated by N-Alkyl Guanidiniums and Pyridiniums.

Multidrug-resistant Gram-negative bacterial infections are on the rise, and with no FDA approvals for new classes of broad-spectrum antibiotics in over 50 years, these infections constitute a major threat to human health. A significant challenge is the inability of most compounds to accumulate in Gram-negative bacteria. Recently developed predictive guidelines show that appending a primary amine to an appropriately shaped compound can enhance Gram-negative accumulation. Here, we report that other positively charged nitrogen functional groups, namely, N-alkyl guanidiniums and pyridiniums, can also facilitate compound uptake into Gram-negative bacteria. The accumulation of a set of 60 nonantibiotic compounds, consisting of 20 primary amines and their corresponding guanidiniums and pyridiniums, was assessed in Escherichia coli. We also installed these alternate functional groups onto antibiotic scaffolds and assessed their accumulation and antibacterial activity in Gram-negative bacteria. The results suggest that other positively-charged, nitrogen-containing functional groups should be considered when designing antibiotics with Gram-negative activity.

Re-engineering natural products to engage new biological targets.

Covering: up to 2020 Natural products have a long history in drug discovery, with their inherent biological activity often tailored by medicinal chemists to arrive at the final drug product. This process is illustrated by numerous examples, including the conversion of epothilone to ixabepilone, erythromycin to azithromycin, and lovastatin to simvastatin. However, natural products are also fruitful starting points for the creation of complex and diverse compounds, especially those that are markedly different from the parent natural product and accordingly do not retain the biological activity of the parent. The resulting products have physiochemical properties that differ considerably when compared to traditional screening collections, thus affording an opportunity to discover novel biological activity. The synthesis of new structural frameworks from natural products thus yields value-added compounds, as demonstrated in the last several years with multiple biological discoveries emerging from these collections. This Highlight details a handful of these studies, describing new compounds derived from natural products that have biological activity and cellular targets different from those evoked/engaged by the parent. Such re-engineering of natural products offers the potential for discovering compounds with interesting and unexpected biological activity.

Gram-Negative Antibiotic Active Through Inhibition of an Essential Riboswitch.

Multidrug-resistant Gram-negative (GN) infections for which there are few available treatment options are increasingly common. The development of new antibiotics for these pathogens is challenging because of the inability of most small molecules to accumulate inside GN bacteria. Using recently developed predictive guidelines for compound accumulation in Escherichia coli, we have converted the antibiotic Ribocil C, which targets the flavin mononucleotide (FMN) riboswitch, from a compound lacking whole-cell activity against wild-type GN pathogens into a compound that accumulates to a high level in E. coli, is effective against Gram-negative clinical isolates, and has efficacy in mouse models of GN infections. This compound allows for the first assessment of the translational potential of FMN riboswitch binders against wild-type Gram-negative bacteria.

Diverse compounds from pleuromutilin lead to a thioredoxin inhibitor and inducer of ferroptosis.

The chemical diversification of natural products provides a robust and general method for the creation of stereochemically rich and structurally diverse small molecules. The resulting compounds have physicochemical traits different from those in most screening collections, and as such are an excellent source for biological discovery. Herein, we subject the diterpene natural product pleuromutilin to reaction sequences focused on creating ring system diversity in few synthetic steps. This effort resulted in a collection of compounds with previously unreported ring systems, providing a novel set of structurally diverse and highly complex compounds suitable for screening in a variety of different settings. Biological evaluation identified the novel compound ferroptocide, a small molecule that rapidly and robustly induces ferroptotic death of cancer cells. Target identification efforts and CRISPR knockout studies reveal that ferroptocide is an inhibitor of thioredoxin, a key component of the antioxidant system in the cell. Ferroptocide positively modulates the immune system in a murine model of breast cancer and will be a useful tool to study the utility of pro-ferroptotic agents for treatment of cancer.

Synthesis and application of a novel bis-1,2,3-triazole ligand containing a 2,2'-bipyrrolidine core.

Herein, we describe the synthesis of a novel bis-1,2,3-triazole ligand which contains an internal N-alkylated 2,2'-bipyrrolidine linker. By using simple starting materials, the ligand could be generated in good yield through several synthetic steps. To investigate the potential for the application of this ligand in transition metal catalysis, we generated a bis-Au(I) complex in nearly quantitative yield and examined its reactivity in the context of alkyne hydration. Both alkyl and aryl terminal alkynes could be efficiently converted to their corresponding ketones in nearly quantitative yields with only 1% catalyst loading under mild conditions.

Gold Redox Catalysis through Base-Initiated Diazonium Decomposition toward Alkene, Alkyne, and Allene Activation.

The discovery of photoassisted diazonium activation toward gold(I) oxidation greatly extended the scope of gold redox catalysis by avoiding the use of a strong oxidant. Some practical issues that limit the application of this new type of chemistry are the relative low efficiency (long reaction time and low conversion) and the strict reaction condition control that is necessary (degassing and inert reaction environment). Herein, an alternative photofree condition has been developed through Lewis base induced diazonium activation. With this method, an unreactive AuI catalyst was used in combination with Na2 CO3 and diazonium salts to produce a AuIII intermediate. The efficient activation of various substrates, including alkyne, alkene and allene was achieved, followed by rapid AuIII reductive elimination, which yielded the C-C coupling products with good to excellent yields. Relative to the previously reported photoactivation method, our approach offered greater efficiency and versatility through faster reaction rates and broader reaction scope. Challenging substrates such as electron rich/neutral allenes, which could not be activated under the photoinitiation conditions (<5 % yield), could be activated to subsequently yield the desired coupling products in good to excellent yield.

Accessing alternative reaction pathways of the intermolecular condensation between homo-propargyl alcohols and terminal alkynes through divergent gold catalysis.

An intermolecular condensation of alkynols and terminal alkynes is reported. Using IPrAuNTf2, an efficient Au-catalyzed cyclization-alkynylation strategy furnishes (2-arylalkynyl) cyclic ethers in moderate to excellent yields (up to 94%). This strategy is extended to the synthesis of functionalized 2,3-dihydrooxepines via the sequential Au-catalyzed ring expansion of the cyclic ether substrates.

Regioselective Amine-Borane Cyclization: Towards the Synthesis of 1,2-BN-3-Cyclohexene by Copper-Assisted Triazole/Gold Catalysis.

The combination of triazole/gold (TA-Au) and Cu(OTf)2 is identified as the optimal catalytic system for promoting intramolecular hydroboration for the synthesis of a six-membered cyclic amine-borane. Excellent yields (up to 95 %) and regioselectivities (5-exo vs. 6-endo) were achieved through catalyst control and sequential dilution. Good functional-group tolerance was attained, thus allowing the preparation of highly functionalized cyclic amine-borane substrates, which could not be achieved using other methods. Deuterium-labeling studies support the involvement of a hydride addition to a gold-activated alkyne with subsequent C-B bond formation.

Ambient synthesis of dienals via triazole-gold and amine catalysis relay.

A highly efficient synthesis of substituted conjugated dienals was developed through a triazole-gold (TA-Au)-catalyzed propargyl vinyl ether rearrangement followed by an amine catalyzed allene-aldehyde tautomerization. Various substituted vinylogous aldehydes were prepared in one pot with good to excellent yields (up to 95%) under mild conditions with high atom economy.

Synthesis of cyclic amine boranes through triazole-gold(I)-catalyzed alkyne hydroboration.

The first catalytic alkyne hydroboration of propargyl amine boranecarbonitriles is accomplished with triazole-Au(I) complexes. While the typical [L-Au](+) species decomposes within minutes upon addition of amine boranecarbonitriles, the triazole-modified gold catalysts (TA-Au) remained active, and allowed the synthesis of 1,2-BN-cyclopentenes in one step with good to excellent yields. With good substrate tolerability and mild reaction conditions (open-flask), this new method provides an alternative route to reach the interesting cyclic amine borane with high efficiency.

The intramolecular allenolate Rauhut-Currier reaction.

An intramolecular Rauhut-Currier reaction utilizing alkynoates as the initial conjugate acceptor affords densely functionalized 5- and 6-membered rings from ynoate-enoate, ynoate-enenitrile, and alkynyl sulfone-enenitrile substrates. Trialkylphosphines catalyze the reaction, and TMSCN serves as a pronucelophile to effect turnover of the catalyst and the formation of a second C-C bond. Because of the highly electrophilic alkyne acceptor, this reaction yields products that cannot be easily accessed from the traditional Rauhut-Currier reaction.