Marcos Pires

Developing selective strategies to treat metastatic cancers remains a significant challenge. Herein, we report the first antibody-recruiting small molecule (ARM) that is capable of recognizing the urokinase-type plasminogen activator receptor (uPAR), a uniquely overexpressed cancer cell-surface marker, and facilitating the immune-mediated destruction of cancer cells. A co-crystal structure of the ARM-U2/uPAR complex was obtained, representing the first crystal structure of uPAR complexed with a non-peptide ligand. Finally, we demonstrated that ARM-U2 substantially suppresses tumor growth in vivo with no evidence of weight loss, unlike the standard-of-care agent doxorubicin. This work underscores the promise of antibody-recruiting molecules as immunotherapeutics for treating cancer.

Marked for death: A small-molecule immunotherapeutic strategy involves the selective tagging of cancer cells for immune-cell recognition and targeted destruction. This approach holds tremendous promise in guiding the development of selective treatments against highly aggressive metastatic cancers with potentially few side effects.

Pharmaceutical and agrochemical discovery programs are under considerable pressure to meet increasing global demand and thus require constant innovation. Classical hydrocarbon scaffolds have long assisted in bringing new molecules to the market place, but an obvious omission is that of the Platonic solid cubane. Eaton, however, suggested that this molecule has the potential to act as a benzene bioisostere. Herein, we report the validation of Eaton's hypothesis with cubane derivatives of five molecules that are used clinically or as agrochemicals. Two cubane analogues showed increased bioactivity compared to their benzene counterparts whereas two further analogues displayed equal bioactivity, and the fifth one demonstrated only partial efficacy. Ramifications from this study are best realized by reflecting on the number of bioactive molecules that contain a benzene ring. Substitution with the cubane scaffold where possible could revitalize these systems, and thus expedite much needed lead candidate identification.

Over 25 years ago, Eaton proposed that cubane could act as a benzene bioisostere. This hypothesis has now been confirmed with the synthesis and evaluation of cubane derivatives of five biologically important molecules: Two cubane analogues showed increased bioactivity compared to their benzene counterparts whereas two further analogues were of equal bioactivity.

Investigation of the physiological and pathological functions of formaldehyde (FA) are largely restricted by a lack of useful FA imaging agents, in particular, those that allow detection of FA in the context of living tissues. Herein, we present the rational design, synthesis, and photophysical property studies of the first two-photon fluorescent FA probe, Na-FA. Importantly, the highly desirable attributes of the probe Na-FA (such as a very large turn-on signal (up to 900-fold), a low detection limit, and a very fast onset imparted by the unique design aspects of the probe), make it possible to monitor endogenous FA in living tissues for the first time. Furthermore, sodium bisulfite was identified as a simple and convenient inhibitor of FA within biological environments.

Shine a little light: The first two-photon fluorescent formaldehyde probe, Na-FA, was engineered by condensation of a hydrazine moiety with FA. This unique strategy grants the probe highly favorable properties: a very large turn-on signal, a low detection limit, and a very fast onset, which collectively enable tracking of endogenous FA in living tissues for the first time.

Developing a monomeric form of an avidin-like protein with highly stable biotin binding properties has been a major challenge in biotin-avidin linking technology. Here we report a monomeric avidin-like protein—enhanced monoavidin—with off-rates almost comparable to those of multimeric avidin proteins against various biotin conjugates. Enhanced monoavidin (eMA) was developed from naturally dimeric rhizavidin by optimally maintaining protein rigidity during monomerization and additionally shielding the bound biotin by diverse engineering of the surface residues. eMA allowed the monovalent and nonperturbing labeling of head-group-biotinylated lipids in bilayer membranes. In addition, we fabricated an unprecedented 24-meric avidin probe by fusing eMA to a multimeric cage protein. The 24-meric avidin and eMA were utilized to demonstrate how artificial clustering of cell-surface proteins greatly enhances the internalization rates of assembled proteins on live cells.

Extremely high binding stability against various biotin conjugates has been observed in a monomeric avidin-like protein—enhanced monoavidin—that was developed from naturally dimeric rhizavidin by minimally disturbing the protein rigidity and additionally shielding the bound biotin. This stable monomeric biotin linker protein was further engineered to generate an unprecedented 24-meric avidin probe.

Chemical probes capable of reacting with KS (ketosynthase)-bound biosynthetic intermediates were utilized for the investigation of the model type I iterative polyketide synthase 6-methylsalicylic acid synthase (6-MSAS) in vivo and in vitro. From the fermentation of fungal and bacterial 6-MSAS hosts in the presence of chain termination probes, a full range of biosynthetic intermediates was isolated and characterized for the first time. Meanwhile, in vitro studies of recombinant 6-MSA synthases with both nonhydrolyzable and hydrolyzable substrate mimics have provided additional insights into substrate recognition, providing the basis for further exploration of the enzyme catalytic activities.

Chemical probes capable of reacting with KS (ketosynthase)-bound biosynthetic intermediates were utilized for the investigation of the model type I iterative polyketide synthase 6-methylsalicylic acid synthase (6-MSAS) in vivo and in vitro. From the fermentation of fungal and bacterial 6-MSAS hosts in the presence of chain termination probes, a full range of biosynthetic intermediates was isolated (see examples) and characterized.

Prodrugs that release hydrogen sulfide upon esterase-mediated cleavage of an ester group followed by lactonization are described herein. By modifying the ester group and thus its susceptibility to esterase, and structural features critical to the lactonization rate, H₂S release rates can be tuned. Such prodrugs directly release hydrogen sulfide without the involvement of perthiol species, which are commonly encountered with existing H₂S donors. Additionally, such prodrugs can easily be conjugated to another non-steroidal anti-inflammatory agent, leading to easy synthesis of hybrid prodrugs. As a biological validation of the H₂S prodrugs, the anti-inflammatory effects of one such prodrug were examined by studying its ability to inhibit LPS-induced TNF-α production in RAW 264.7 cells. This type of H₂S prodrugs shows great potential as both research tools and therapeutic agents.

Tunable H₂S supply: Prodrugs that release hydrogen sulfide upon esterase-mediated cleavage of an ester group followed by lactonization are described (see example). By modifying the ester group and thus its susceptibility to esterase H₂S release rates can be tuned. The anti-inflammatory effects of one candidate were examined by studying its ability to inhibit TNF-α production in RAW 264.7 cells.

The synthesis of all 20 common natural proteinogenic and 4 otherα-amino acid-isosteric α-amino tetrazoles has been accomplished, whereby the carboxyl group is replaced by the isosteric 5-tetrazolyl group. The short process involves the use of the key Ugi tetrazole reaction followed by deprotection chemistries. The tetrazole group is bioisosteric to the carboxylic acid and is widely used in medicinal chemistry and drug design. Surprisingly, several of the common α-amino acid-isosteric α-amino tetrazoles are unknown up to now. Therefore a rapid synthetic access to this compound class and non-natural derivatives is of high interest to advance the field.

The good, the bad, and the Ugi: 20 natural proteinogenic and 4 other α-amino acid-isosteric α-amino tetrazoles have been synthesized under benign conditions according to a concise and efficient isocyanide-based multicomponent synthetic route. Several previously unknown ones are firstly synthesized and reported.

Mechanophores, that is, molecules that show a defined response to force, are crucial building blocks of mechanoresponsive materials. The possibility of mechanically induced cycloreversion for a series of triazoles formed via strain-promoted azide–alkyne cycloaddition reactions was investigated by density functional theory calculations, and these triazoles were compared to the 1,4- and 1,5-regioisomers formed in the reaction of an azide with a terminal alkyne. We show that cycloreversion is in principal possible and that the pulling geometry is the most important parameter that determines the probability of cycloreversion. We further compared triazole stability to the mechanical stability of polymers that are frequently used as force transducers in mechanochemical experiments and identified DIBAC (azadibenzylcyclooctyne) as a promising mechanophore for future applications.

Broken by you: Mechanical cycloreversion of triazoles depends on the direction of the applied force (shearing vs. unzipping) acting on the triazole unit. Computational screening of a number of triazoles formed from strained alkynes shows that cycloreversion only occurs when the force acts in the unzipping geometry and identifies azadibenzylcyclooctyne (DIBAC) as a promising strained alkyne for mechanically reversible triazole structures.