MOESM1 of Spectrophores as one-dimensional descriptors calculated from three-dimensional atomic properties: applications ranging from scaffold hopping to multi-target virtual screening

Additional file 1. Ring fragments and their corresponding clusters

MOESM2 of Spectrophores as one-dimensional descriptors calculated from three-dimensional atomic properties: applications ranging from scaffold hopping to multi-target virtual screening

Additional file 2. Experimental procedures for the biochemical assay

Characterization of Structurally Diverse ¹⁸F‑Labeled d‑TCO Derivatives as a PET Probe for Bioorthogonal Pretargeted Imaging

Background: The pretargeted imaging strategy using inverse electron demand Diels–Alder (IEDDA) cycloaddition between a trans-cyclooctene (TCO) and tetrazine (Tz) has emerged and rapidly grown as a promising concept to improve radionuclide imaging and therapy in oncology. This strategy has mostly relied on the use of radiolabeled Tz together with TCO-modified targeting vectors leading to a rapid growth of the number of available radiolabeled tetrazines, while only a few radiolabeled TCOs are currently reported. Here, we aim to develop novel and structurally diverse 18F-labeled cis-dioxolane-fused TCO (d-TCO) derivatives to further expand the bioorthogonal toolbox for in vivo ligation and evaluate their potential for positron emission tomography (PET) pretargeted imaging. Results: A small series of d-TCO derivatives were synthesized and tested for their reactivity against tetrazines, with all compounds showing fast reaction kinetics with tetrazines. A fluorescence-based pretargeted blocking study was developed to investigate the in vivo ligation of these compounds without labor-intensive prior radiochemical development. Two compounds showed excellent in vivo ligation results with blocking efficiencies of 95 and 97%. Two novel 18F-labeled d-TCO radiotracers were developed, from which [18F]MICA-214 showed good in vitro stability, favorable pharmacokinetics, and moderate in vivo stability. Micro-PET pretargeted imaging with [18F]MICA-214 in mice bearing LS174T tumors treated with tetrazine-modified CC49 monoclonal antibody (mAb) (CC49-Tz) showed significantly higher uptake in tumor tissue in the pretargeted group (CC49-Tz 2.16 ± 0.08% ID/mL) when compared to the control group with nonmodified mAb (CC49 1.34 ± 0.07% ID/mL). Conclusions: A diverse series of fast-reacting fluorinated d-TCOs were synthesized. A pretargeted blocking approach in tumor-bearing mice allowed the choice of a lead compound with fast reaction kinetics with Tz. A novel 18F-labeled d-TCO tracer was developed and used in a pretargeted PET imaging approach, allowing specific tumor visualization in a mouse model of colorectal cancer. Although further optimization of the radiotracer is needed to enhance the tumor-to-background ratios for pretargeted imaging, we anticipate that the 18F-labeled d-TCO will find use in studies where increased hydrophilicity and fast bioconjugation are required

Rescue of <i>SPP1</i> RNAi growth defect by expression of recoded SPPI.

<p>(a) Expression of recoded HA-tagged SPP1 detected by Western blot. Cell lysates from RNAi cell line (-), the RNAi cell line expressing SPP1 from the recoded gene (<i>SPP1</i>^<i>R</i>), or the RNAi cell line expressing inactive SPP1 from a recodedgene (<i>SPP1</i>^{<i>R</i>,<i>I</i>}) were probed with anti-HA antibody (Roche). Detection of EF-1α was used as a loading control. (b, c and e) Parasite growth was measured in cell lines after inducing <i>SPP1</i> RNAi with tetracycline (open squares) or without treatment (closed squares) in the <i>SPP1</i> RNAi cell line expressing <i>SPP1</i> from: the recoded gene <i>SPP1</i>^<i>R</i> (b), the parental <i>SPP1</i> RNAi cell line (c), or the RNAi cell line expressing inactive SPP1 from a recoded gene, <i>SPP1</i>^{<i>R</i>,<i>I</i>} (e). (d) Quantitative PCR showing relative quantification (RQ) of endogenous <i>SPP1</i> transcript in the RNAi cell line expressing active SPP1 from the <i>SPP1</i>^<i>R</i> gene either without induction (control, black bars) or after induction by tetracycline (+ Tet, white bars).</p

Effect of RNAi on <i>T</i>. <i>brucei</i> cell growth <i>in vivo</i>.

<p>ICR mice were inoculated with RNAi cell lines targeting the two OPB-like genes (a), two POP-like genes (b), dipeptidyl peptidase-8 (c), or the type-I signal peptidase (d). Two mice in each experiment were left untreated (filled symbols) and two were given doxycycline (open symbols) to induce RNAi. The arrow indicates doxycycline administration. Parasitaemia in infected mice was counted at the times indicated.</p

Recombinant TbDPP8 activity and inhibition.

<p>(a) Rate of cleavage by TbDPP8 of H-Gly-Pro-AMC (grey bars) and Z-Gly-Pro-AMC (white bars). (b) Structures of inhibitors used against TbDPP8. (c) Inhibitory activity of compounds against <i>T</i>. <i>brucei</i> DPP8, human DPP IV, human DPP8, and bloodstream form <i>T</i>. <i>brucei</i> 427.</p

Discovery and SAR of Novel and Selective Inhibitors of Urokinase Plasminogen Activator (uPA) with an Imidazo[1,2‑<i>a</i>]pyridine Scaffold

Urokinase plasminogen activator (uPA) is a biomarker and therapeutic target for several cancer types. Its inhibition is regarded as a promising, noncytotoxic approach in cancer therapy by blocking growth and/or metastasis of solid tumors. Earlier, we reported the modified substrate activity screening (MSAS) approach and applied it for the identification of fragments with affinity for uPA’s S1 pocket. Here, these fragments are transformed into a novel class of uPA inhibitors with an imidazo­[1,2-<i>a</i>]­pyridine scaffold. The SAR for uPA inhibition around this scaffold is explored, and the best compounds in the series have nanomolar uPA affinity and selectivity with respect to the related trypsin-like serine proteases (thrombin, tPA, FXa, plasmin, plasma kallikrein, trypsin, FVIIa). Finally, the approach followed for translating fragments into small molecules with a decorated scaffold architecture is conceptually straightforward and can be expected to be broadly applicable in fragment-based drug design

Extended Structure–Activity Relationship and Pharmacokinetic Investigation of (4-Quinolinoyl)glycyl-2-cyanopyrrolidine Inhibitors of Fibroblast Activation Protein (FAP)

Fibroblast activation protein (FAP) is a serine protease related to dipeptidyl peptidase IV (DPPIV). It has been convincingly linked to multiple disease states involving remodeling of the extracellular matrix. FAP inhibition is investigated as a therapeutic option for several of these diseases, with most attention so far devoted to oncology applications. We previously discovered the <i>N</i>-4-quinolinoyl-Gly-(2<i>S</i>)-cyanoPro scaffold as a possible entry to highly potent and selective FAP inhibitors. In the present study, we explore in detail the structure–activity relationship around this core scaffold. We report extensively optimized compounds that display low nanomolar inhibitory potency and high selectivity against the related dipeptidyl peptidases (DPPs) DPPIV, DPP9, DPPII, and prolyl oligopeptidase (PREP). The log <i>D</i> values, plasma stabilities, and microsomal stabilities of selected compounds were found to be highly satisfactory. Pharmacokinetic evaluation in mice of selected inhibitors demonstrated high oral bioavailability, plasma half-life, and the potential to selectively and completely inhibit FAP in vivo

Selective Inhibitors of Fibroblast Activation Protein (FAP) with a (4-Quinolinoyl)-glycyl-2-cyanopyrrolidine Scaffold

Fibroblast activation protein (FAP) is a serine protease that is generally accepted to play an important role in tumor growth and other diseases involving tissue remodeling. Currently there are no FAP inhibitors with reported selectivity toward both the closely related dipeptidyl peptidases (DPPs) and prolyl oligopeptidase (PREP). We present the discovery of a new class of FAP inhibitors with a <i>N</i>-(4-quinolinoyl)-Gly-(2-cyanopyrrolidine) scaffold. We have explored the effects of substituting the quinoline ring and varying the position of its sp² hybridized nitrogen atom. The most promising inhibitors combined low nanomolar FAP inhibition and high selectivity indices (>10³) with respect to both the DPPs and PREP. Preliminary experiments on a representative inhibitor demonstrate that plasma stability, kinetic solubility, and log <i>D</i> of this class of compounds can be expected to be satisfactory

Searching for New Leads for Tuberculosis: Design, Synthesis, and Biological Evaluation of Novel 2‑Quinolin-4-yloxyacetamides

In this study, a new series of more than 60 quinoline derivatives has been synthesized and evaluated against Mycobacterium tuberculosis (H37Rv). Apart from the SAR exploration around the initial hits, the optimization process focused on the improvement of the physicochemical properties, cytotoxicity, and metabolic stability of the series. The best compounds obtained exhibited MIC values in the low micromolar range, excellent intracellular antimycobacterial activity, and an improved physicochemical profile without cytotoxic effects. Further investigation revealed that the amide bond was the source for the poor blood stability observed, while some of the compounds exhibited hERG affinity. Compound <b>83</b> which contains a benzoxazole ring instead of the amide group was found to be a good alternative, with good blood stability and no hERG affinity, providing new opportunities for the series. Overall, the obtained results suggest that further optimization of solubility and microsomal stability of the series could provide a strong lead for a new anti-TB drug development program